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Sample records for acid sequence indicating

  1. Distinguishing Proteins From Arbitrary Amino Acid Sequences

    PubMed Central

    Yau, Stephen S.-T.; Mao, Wei-Guang; Benson, Max; He, Rong Lucy

    2015-01-01

    What kinds of amino acid sequences could possibly be protein sequences? From all existing databases that we can find, known proteins are only a small fraction of all possible combinations of amino acids. Beginning with Sanger's first detailed determination of a protein sequence in 1952, previous studies have focused on describing the structure of existing protein sequences in order to construct the protein universe. No one, however, has developed a criteria for determining whether an arbitrary amino acid sequence can be a protein. Here we show that when the collection of arbitrary amino acid sequences is viewed in an appropriate geometric context, the protein sequences cluster together. This leads to a new computational test, described here, that has proved to be remarkably accurate at determining whether an arbitrary amino acid sequence can be a protein. Even more, if the results of this test indicate that the sequence can be a protein, and it is indeed a protein sequence, then its identity as a protein sequence is uniquely defined. We anticipate our computational test will be useful for those who are attempting to complete the job of discovering all proteins, or constructing the protein universe. PMID:25609314

  2. Composition for nucleic acid sequencing

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2008-08-26

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid, i.e. the activity of a nucleic acid polymerizing enzyme on the template nucleic acid molecule to be sequenced is followed in real time. The sequence is deduced by identifying which base is being incorporated into the growing complementary strand of the target nucleic acid by the catalytic activity of the nucleic acid polymerizing enzyme at each step in the sequence of base additions. A polymerase on the target nucleic acid molecule complex is provided in a position suitable to move along the target nucleic acid molecule and extend the oligonucleotide primer at an active site. A plurality of labelled types of nucleotide analogs are provided proximate to the active site, with each distinguishable type of nucleotide analog being complementary to a different nucleotide in the target nucleic acid sequence. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand at the active site, where the nucleotide analog being added is complementary to the nucleotide of the target nucleic acid at the active site. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The steps of providing labelled nucleotide analogs, polymerizing the growing nucleic acid strand, and identifying the added nucleotide analog are repeated so that the nucleic acid strand is further extended and the sequence of the target nucleic acid is determined.

  3. Method for identifying and quantifying nucleic acid sequence aberrations

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    1998-01-01

    A method for detecting nucleic acid sequence aberrations by detecting nucleic acid sequences having both a first and a second nucleic acid sequence type, the presence of the first and second sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. The method uses a first hybridization probe which includes a nucleic acid sequence that is complementary to a first sequence type and a first complexing agent capable of attaching to a second complexing agent and a second hybridization probe which includes a nucleic acid sequence that selectively hybridizes to the second nucleic acid sequence type over the first sequence type and includes a detectable marker for detecting the second hybridization probe.

  4. Method for identifying and quantifying nucleic acid sequence aberrations

    DOEpatents

    Lucas, J.N.; Straume, T.; Bogen, K.T.

    1998-07-21

    A method is disclosed for detecting nucleic acid sequence aberrations by detecting nucleic acid sequences having both a first and a second nucleic acid sequence type, the presence of the first and second sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. The method uses a first hybridization probe which includes a nucleic acid sequence that is complementary to a first sequence type and a first complexing agent capable of attaching to a second complexing agent and a second hybridization probe which includes a nucleic acid sequence that selectively hybridizes to the second nucleic acid sequence type over the first sequence type and includes a detectable marker for detecting the second hybridization probe. 11 figs.

  5. High speed nucleic acid sequencing

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2011-05-17

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid. Each type of labeled nucleotide comprises an acceptor fluorophore attached to a phosphate portion of the nucleotide such that the fluorophore is removed upon incorporation into a growing strand. Fluorescent signal is emitted via fluorescent resonance energy transfer between the donor fluorophore and the acceptor fluorophore as each nucleotide is incorporated into the growing strand. The sequence is deduced by identifying which base is being incorporated into the growing strand.

  6. The Draft Genome Sequence of Sphingomonas sp. Strain FukuSWIS1, Obtained from Acidic Lake Grosse Fuchskuhle, Indicates Photoheterotrophy and a Potential for Humic Matter Degradation

    PubMed Central

    Salka, Ivette; Srivastava, Abhishek; Allgaier, Martin

    2014-01-01

    Sphingomonas spp. are Alphaproteobacteria considered to be versatile bacteria that can utilize a variety of natural substrates available in terrestrial and aquatic systems. Sphingomonas sp. strain FukuSWIS1 was isolated from the eutrophic and acidic freshwater Lake Grosse Fuchskuhle in northeastern Germany. The strain has a genome size of 3.89 Mb, possesses a set of photosynthetic genes, and expresses photopigment BChl a under oxic conditions. Thus, this strain belongs to the aerobic anoxygenic phototrophic (AAP) bacteria, which are most likely involved in humic matter degradation as indicated by the presence of organic compound mineralizing genes. PMID:25395647

  7. Detection of nucleic acid sequences by invader-directed cleavage

    DOEpatents

    Brow, Mary Ann D.; Hall, Jeff Steven Grotelueschen; Lyamichev, Victor; Olive, David Michael; Prudent, James Robert

    1999-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The 5' nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based by charge.

  8. Chip-based sequencing nucleic acids

    DOEpatents

    Beer, Neil Reginald

    2014-08-26

    A system for fast DNA sequencing by amplification of genetic material within microreactors, denaturing, demulsifying, and then sequencing the material, while retaining it in a PCR/sequencing zone by a magnetic field. One embodiment includes sequencing nucleic acids on a microchip that includes a microchannel flow channel in the microchip. The nucleic acids are isolated and hybridized to magnetic nanoparticles or to magnetic polystyrene-coated beads. Microreactor droplets are formed in the microchannel flow channel. The microreactor droplets containing the nucleic acids and the magnetic nanoparticles are retained in a magnetic trap in the microchannel flow channel and sequenced.

  9. Method for sequencing nucleic acid molecules

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2006-05-30

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid, i.e. the activity of a nucleic acid polymerizing enzyme on the template nucleic acid molecule to be sequenced is followed in real time. The sequence is deduced by identifying which base is being incorporated into the growing complementary strand of the target nucleic acid by the catalytic activity of the nucleic acid polymerizing enzyme at each step in the sequence of base additions. A polymerase on the target nucleic acid molecule complex is provided in a position suitable to move along the target nucleic acid molecule and extend the oligonucleotide primer at an active site. A plurality of labelled types of nucleotide analogs are provided proximate to the active site, with each distinguishable type of nucleotide analog being complementary to a different nucleotide in the target nucleic acid sequence. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand at the active site, where the nucleotide analog being added is complementary to the nucleotide of the target nucleic acid at the active site. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The steps of providing labelled nucleotide analogs, polymerizing the growing nucleic acid strand, and identifying the added nucleotide analog are repeated so that the nucleic acid strand is further extended and the sequence of the target nucleic acid is determined.

  10. Method for sequencing nucleic acid molecules

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2006-06-06

    The present invention is directed to a method of sequencing a target nucleic acid molecule having a plurality of bases. In its principle, the temporal order of base additions during the polymerization reaction is measured on a molecule of nucleic acid, i.e. the activity of a nucleic acid polymerizing enzyme on the template nucleic acid molecule to be sequenced is followed in real time. The sequence is deduced by identifying which base is being incorporated into the growing complementary strand of the target nucleic acid by the catalytic activity of the nucleic acid polymerizing enzyme at each step in the sequence of base additions. A polymerase on the target nucleic acid molecule complex is provided in a position suitable to move along the target nucleic acid molecule and extend the oligonucleotide primer at an active site. A plurality of labelled types of nucleotide analogs are provided proximate to the active site, with each distinguishable type of nucleotide analog being complementary to a different nucleotide in the target nucleic acid sequence. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand at the active site, where the nucleotide analog being added is complementary to the nucleotide of the target nucleic acid at the active site. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The steps of providing labelled nucleotide analogs, polymerizing the growing nucleic acid strand, and identifying the added nucleotide analog are repeated so that the nucleic acid strand is further extended and the sequence of the target nucleic acid is determined.

  11. Phenolic acid esterases, coding sequences and methods

    DOEpatents

    Blum, David L.; Kataeva, Irina; Li, Xin-Liang; Ljungdahl, Lars G.

    2002-01-01

    Described herein are four phenolic acid esterases, three of which correspond to domains of previously unknown function within bacterial xylanases, from XynY and XynZ of Clostridium thermocellum and from a xylanase of Ruminococcus. The fourth specifically exemplified xylanase is a protein encoded within the genome of Orpinomyces PC-2. The amino acids of these polypeptides and nucleotide sequences encoding them are provided. Recombinant host cells, expression vectors and methods for the recombinant production of phenolic acid esterases are also provided.

  12. Amino-Acid Sequence of Porcine Pepsin

    PubMed Central

    Tang, J.; Sepulveda, P.; Marciniszyn, J.; Chen, K. C. S.; Huang, W-Y.; Tao, N.; Liu, D.; Lanier, J. P.

    1973-01-01

    As the culmination of several years of experiments, we propose a complete amino-acid sequence for porcine pepsin, an enzyme containing 327 amino-acid residues in a single polypeptide chain. In the sequence determination, the enzyme was treated with cyanogen bromide. Five resulting fragments were purified. The amino-acid sequence of four of the fragments accounted for 290 residues. Because the structure of a 37-residue carboxyl-terminal fragment was already known, it was not studied. The alignment of these fragments was determined from the sequence of methionyl-peptides we had previously reported. We also discovered the locations of activesite aspartyl residues, as well as the pairing of the three disulfide bridges. A minor component of commercial crystalline pepsin was found to contain two extra amino-acid residues, Ala-Leu-, at the amino-terminus of the molecule. This minor component was apparently derived from a different site of cleavage during the activation of porcine pepsinogen. PMID:4587252

  13. Thermal infrared anomaly indicating unformed strong earthquake sequences

    NASA Astrophysics Data System (ADS)

    Yao, Qinglin; Qiang, Zuji

    2015-01-01

    By processing and analyzing many satellite remote sensing images in infrared channels, we found visual evidence indicating earthquake sequences. In a sequence, large earthquakes could migrate thousands of kilometers along a linear route from one land mass to another, and infrared anomalies also cover these land masses. The sequence has at least two types: (1) quakes alternately occurring at two ends of a belt, or (2) quakes progressively migrating along a far-flung route. The former can be foreshowed in a single infrared belt with a legible directed structure and higher brightness temperature; the latter can be identified in advance from the shift, veer, and deformation of more areas with a higher brightness temperature. The thermal infrared anomalies with a shorter duration are used to predict the quake-sequence evolving in a longer period. Two sorts of sequences can meet and form a more complex structure that can also be shown in advance by an anomalous infrared area. These studies on the seismic sequences will provide a potent means for analyzing earthquake-pregnant fields and estimating the location of unformed earthquake sequences.

  14. Methods for analyzing nucleic acid sequences

    DOEpatents

    Korlach, Jonas; Webb, Watt W.; Levene, Michael; Turner, Stephen; Craighead, Harold G.; Foquet, Mathieu

    2011-05-17

    The present invention is directed to a method of sequencing a target nucleic acid. The method provides a complex comprising a polymerase enzyme, a target nucleic acid molecule, and a primer, wherein the complex is immobilized on a support Fluorescent label is attached to a terminal phosphate group of the nucleotide or nucleotide analog. The growing nucleic acid strand is extended by using the polymerase to add a nucleotide analog to the nucleic acid strand. The nucleotide analog added to the oligonucleotide primer as a result of the polymerizing step is identified. The time duration of the signal from labeled nucleotides or nucleotide analogs that become incorporated is distinguished from freely diffusing labels by a longer retention in the observation volume for the nucleotides or nucleotide analogs that become incorporated than for the freely diffusing labels.

  15. An Olfactory Indicator for Acid-Base Titrations.

    ERIC Educational Resources Information Center

    Flair, Mark N.; Setzer, William N.

    1990-01-01

    The use of an olfactory acid-base indicator in titrations for visually impaired students is discussed. Potential olfactory indicators include eugenol, thymol, vanillin, and thiophenol. Titrations performed with each indicator with eugenol proved to be successful. (KR)

  16. Amino acid sequence of the Amur tiger prion protein.

    PubMed

    Wu, Changde; Pang, Wanyong; Zhao, Deming

    2006-10-01

    Prion diseases are fatal neurodegenerative disorders in human and animal associated with conformational conversion of a cellular prion protein (PrP(C)) into the pathologic isoform (PrP(Sc)). Various data indicate that the polymorphisms within the open reading frame (ORF) of PrP are associated with the susceptibility and control the species barrier in prion diseases. In the present study, partial Prnp from 25 Amur tigers (tPrnp) were cloned and screened for polymorphisms. Four single nucleotide polymorphisms (T423C, A501G, C511A, A610G) were found; the C511A and A610G nucleotide substitutions resulted in the amino acid changes Lysine171Glutamine and Alanine204Threoine, respectively. The tPrnp amino acid sequence is similar to house cat (Felis catus ) and sheep, but differs significantly from other two cat Prnp sequences that were previously deposited in GenBank. PMID:16780982

  17. Hybridization and sequencing of nucleic acids using base pair mismatches

    DOEpatents

    Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

    2001-01-01

    Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.

  18. Identification of random nucleic acid sequence aberrations using dual capture probes which hybridize to different chromosome regions

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    1998-01-01

    A method is provided for detecting nucleic acid sequence aberrations using two immobilization steps. According to the method, a nucleic acid sequence aberration is detected by detecting nucleic acid sequences having both a first nucleic acid sequence type (e.g., from a first chromosome) and a second nucleic acid sequence type (e.g., from a second chromosome), the presence of the first and the second nucleic acid sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. In the method, immobilization of a first hybridization probe is used to isolate a first set of nucleic acids in the sample which contain the first nucleic acid sequence type. Immobilization of a second hybridization probe is then used to isolate a second set of nucleic acids from within the first set of nucleic acids which contain the second nucleic acid sequence type. The second set of nucleic acids are then detected, their presence indicating the presence of a nucleic acid sequence aberration.

  19. Identification of random nucleic acid sequence aberrations using dual capture probes which hybridize to different chromosome regions

    DOEpatents

    Lucas, J.N.; Straume, T.; Bogen, K.T.

    1998-03-24

    A method is provided for detecting nucleic acid sequence aberrations using two immobilization steps. According to the method, a nucleic acid sequence aberration is detected by detecting nucleic acid sequences having both a first nucleic acid sequence type (e.g., from a first chromosome) and a second nucleic acid sequence type (e.g., from a second chromosome), the presence of the first and the second nucleic acid sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. In the method, immobilization of a first hybridization probe is used to isolate a first set of nucleic acids in the sample which contain the first nucleic acid sequence type. Immobilization of a second hybridization probe is then used to isolate a second set of nucleic acids from within the first set of nucleic acids which contain the second nucleic acid sequence type. The second set of nucleic acids are then detected, their presence indicating the presence of a nucleic acid sequence aberration. 14 figs.

  20. Characterization and amino acid sequence of a fatty acid-binding protein from human heart.

    PubMed

    Offner, G D; Brecher, P; Sawlivich, W B; Costello, C E; Troxler, R F

    1988-05-15

    The complete amino acid sequence of a fatty acid-binding protein from human heart was determined by automated Edman degradation of CNBr, BNPS-skatole [3'-bromo-3-methyl-2-(2-nitrobenzenesulphenyl)indolenine], hydroxylamine, Staphylococcus aureus V8 proteinase, tryptic and chymotryptic peptides, and by digestion of the protein with carboxypeptidase A. The sequence of the blocked N-terminal tryptic peptide from citraconylated protein was determined by collisionally induced decomposition mass spectrometry. The protein contains 132 amino acid residues, is enriched with respect to threonine and lysine, lacks cysteine, has an acetylated valine residue at the N-terminus, and has an Mr of 14768 and an isoelectric point of 5.25. This protein contains two short internal repeated sequences from residues 48-54 and from residues 114-119 located within regions of predicted beta-structure and decreasing hydrophobicity. These short repeats are contained within two longer repeated regions from residues 48-60 and residues 114-125, which display 62% sequence similarity. These regions could accommodate the charged and uncharged moieties of long-chain fatty acids and may represent fatty acid-binding domains consistent with the finding that human heart fatty acid-binding protein binds 2 mol of oleate or palmitate/mol of protein. Detailed evidence for the amino acid sequences of the peptides has been deposited as Supplementary Publication SUP 50143 (23 pages) at the British Library Lending Division, Boston Spa, Yorkshire LS23 7BQ, U.K., from whom copies may be obtained as indicated in Biochem. J. (1988) 249, 5. PMID:3421901

  1. 77 FR 65537 - Requirements for Patent Applications Containing Nucleotide Sequence and/or Amino Acid Sequence...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-29

    ... Amino Acid Sequence Disclosures ACTION: Proposed collection; comment request. SUMMARY: The United States....'' SUPPLEMENTARY INFORMATION: I. Abstract Patent applications that contain nucleotide and/or amino acid sequence disclosures must include a copy of the sequence listing in accordance with the requirements in 37 CFR...

  2. Molecular cloning and amino acid sequence of human 5-lipoxygenase

    SciTech Connect

    Matsumoto, T.; Funk, C.D.; Radmark, O.; Hoeoeg, J.O.; Joernvall, H.; Samuelsson, B.

    1988-01-01

    5-Lipoxygenase (EC 1.13.11.34), a Ca/sup 2 +/- and ATP-requiring enzyme, catalyzes the first two steps in the biosynthesis of the peptidoleukotrienes and the chemotactic factor leukotriene B/sub 4/. A cDNA clone corresponding to 5-lipoxygenase was isolated from a human lung lambda gt11 expression library by immunoscreening with a polyclonal antibody. Additional clones from a human placenta lambda gt11 cDNA library were obtained by plaque hybridization with the /sup 32/P-labeled lung cDNA clone. Sequence data obtained from several overlapping clones indicate that the composite DNAs contain the complete coding region for the enzyme. From the deduced primary structure, 5-lipoxygenase encodes a 673 amino acid protein with a calculated molecular weight of 77,839. Direct analysis of the native protein and its proteolytic fragments confirmed the deduced composition, the amino-terminal amino acid sequence, and the structure of many internal segments. 5-Lipoxygenase has no apparent sequence homology with leukotriene A/sub 4/ hydrolase or Ca/sup 2 +/-binding proteins. RNA blot analysis indicated substantial amounts of an mRNA species of approx. = 2700 nucleotides in leukocytes, lung, and placenta.

  3. Predicting intrinsic disorder from amino acid sequence.

    PubMed

    Obradovic, Zoran; Peng, Kang; Vucetic, Slobodan; Radivojac, Predrag; Brown, Celeste J; Dunker, A Keith

    2003-01-01

    Blind predictions of intrinsic order and disorder were made on 42 proteins subsequently revealed to contain 9,044 ordered residues, 284 disordered residues in 26 segments of length 30 residues or less, and 281 disordered residues in 2 disordered segments of length greater than 30 residues. The accuracies of the six predictors used in this experiment ranged from 77% to 91% for the ordered regions and from 56% to 78% for the disordered segments. The average of the order and disorder predictions ranged from 73% to 77%. The prediction of disorder in the shorter segments was poor, from 25% to 66% correct, while the prediction of disorder in the longer segments was better, from 75% to 95% correct. Four of the predictors were composed of ensembles of neural networks. This enabled them to deal more efficiently with the large asymmetry in the training data through diversified sampling from the significantly larger ordered set and achieve better accuracy on ordered and long disordered regions. The exclusive use of long disordered regions for predictor training likely contributed to the disparity of the predictions on long versus short disordered regions, while averaging the output values over 61-residue windows to eliminate short predictions of order or disorder probably contributed to the even greater disparity for three of the predictors. This experiment supports the predictability of intrinsic disorder from amino acid sequence. PMID:14579347

  4. Amino acid sequence of the nonsecretory ribonuclease of human urine.

    PubMed

    Beintema, J J; Hofsteenge, J; Iwama, M; Morita, T; Ohgi, K; Irie, M; Sugiyama, R H; Schieven, G L; Dekker, C A; Glitz, D G

    1988-06-14

    The amino acid sequence of a nonsecretory ribonuclease isolated from human urine was determined except for the identity of the residue at position 7. Sequence information indicates that the ribonucleases of human liver and spleen and an eosinophil-derived neurotoxin are identical or very closely related gene products. The sequence is identical at about 30% of the amino acid positions with those of all of the secreted mammalian ribonucleases for which information is available. Identical residues include active-site residues histidine-12, histidine-119, and lysine-41, other residues known to be important for substrate binding and catalytic activity, and all eight half-cystine residues common to these enzymes. Major differences include a deletion of six residues in the (so-called) S-peptide loop, insertions of two, and nine residues, respectively, in three other external loops of the molecule, and an addition of three residues at the amino terminus. The sequence shows the human nonsecretory ribonuclease to belong to the same ribonuclease superfamily as the mammalian secretory ribonucleases, turtle pancreatic ribonuclease, and human angiogenin. Sequence data suggest that a gene duplication occurred in an ancient vertebrate ancestor; one branch led to the nonsecretory ribonuclease, while the other branch led to a second duplication, with one line leading to the secretory ribonucleases (in mammals) and the second line leading to pancreatic ribonuclease in turtle and an angiogenic factor in mammals (human angiogenin). The nonsecretory ribonuclease has five short carbohydrate chains attached via asparagine residues at the surface of the molecule; these chains may have been shortened by exoglycosidase action.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3166997

  5. Methods and compositions for efficient nucleic acid sequencing

    DOEpatents

    Drmanac, Radoje

    2002-01-01

    Disclosed are novel methods and compositions for rapid and highly efficient nucleic acid sequencing based upon hybridization with two sets of small oligonucleotide probes of known sequences. Extremely large nucleic acid molecules, including chromosomes and non-amplified RNA, may be sequenced without prior cloning or subcloning steps. The methods of the invention also solve various current problems associated with sequencing technology such as, for example, high noise to signal ratios and difficult discrimination, attaching many nucleic acid fragments to a surface, preparing many, longer or more complex probes and labelling more species.

  6. Methods and compositions for efficient nucleic acid sequencing

    DOEpatents

    Drmanac, Radoje

    2006-07-04

    Disclosed are novel methods and compositions for rapid and highly efficient nucleic acid sequencing based upon hybridization with two sets of small oligonucleotide probes of known sequences. Extremely large nucleic acid molecules, including chromosomes and non-amplified RNA, may be sequenced without prior cloning or subcloning steps. The methods of the invention also solve various current problems associated with sequencing technology such as, for example, high noise to signal ratios and difficult discrimination, attaching many nucleic acid fragments to a surface, preparing many, longer or more complex probes and labelling more species.

  7. Kit for detecting nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    2001-01-01

    A kit is provided for detecting a target nucleic acid sequence in a sample, the kit comprising: a first hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a first portion of the target sequence, the first hybridization probe including a first complexing agent for forming a binding pair with a second complexing agent; and a second hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a second portion of the target sequence to which the first hybridization probe does not selectively hybridize, the second hybridization probe including a detectable marker; a third hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a first portion of the target sequence, the third hybridization probe including the same detectable marker as the second hybridization probe; and a fourth hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a second portion of the target sequence to which the third hybridization probe does not selectively hybridize, the fourth hybridization probe including the first complexing agent for forming a binding pair with the second complexing agent; wherein the first and second hybridization probes are capable of simultaneously hybridizing to the target sequence and the third and fourth hybridization probes are capable of simultaneously hybridizing to the target sequence, the detectable marker is not present on the first or fourth hybridization probes and the first, second, third, and fourth hybridization probes each include a competitive nucleic acid sequence which is sufficiently complementary to a third portion of the target sequence that the competitive sequences of the first, second, third, and fourth hybridization probes compete with each other to hybridize to the third portion of the

  8. Dye Indicators for Acidic or Basic Surface Contamination

    NASA Technical Reports Server (NTRS)

    Lakin, A.; Schuler, F.

    1984-01-01

    Application of pH-sensitive dye solution serves as test for acidic or basic contamination of critical bonding surface. Aqueous solution of 0.1 percent Direct Red No. 28 capable of indicating acid activating solution down to 10 parts per million on hardware and tooling. Dye did not cause detectable contamination of surface.

  9. Unit: Indicating Acidity, Inspection Pack, National Trial Print.

    ERIC Educational Resources Information Center

    Australian Science Education Project, Toorak, Victoria.

    The introductory core activities in this trial unit, prepared for students in grades seven through nine of Australian schools, use indicators derived from flower pigments to provide a more convenient measure of acidity than taste. Students are offered choices among seven options after completion of the core: "How Acidic is That?"; "What Colour is…

  10. The amino acid sequence of chymopapain from Carica papaya.

    PubMed Central

    Watson, D C; Yaguchi, M; Lynn, K R

    1990-01-01

    Chymopapain is a polypeptide of 218 amino acid residues. It has considerable structural similarity with papain and papaya proteinase omega, including conservation of the catalytic site and of the disulphide bonding. Chymopapain is like papaya proteinase omega in carrying four extra residues between papain positions 168 and 169, but differs from both papaya proteinases in the composition of its S2 subsite, as well as in having a second thiol group, Cys-117. Some evidence for the amino acid sequence of chymopapain has been deposited as Supplementary Publication SUP 50153 (12 pages) at the British Library Document Supply Centre, Boston Spa., Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1990) 265, 5. The information comprises Supplement Tables 1-4, which contain, in order, amino acid compositions of peptides from tryptic, peptic, CNBr and mild acid cleavages, Supplement Fig. 1, showing re-fractionation of selected peaks from Fig. 2 of the main paper. Supplement Fig. 2, showing cation-exchange chromatography of the earliest-eluted peak of Fig. 3 of the main paper, Supplement Fig. 3, showing reverse-phase h.p.l.c. of the later-eluted peak from Fig. 3 of the main paper, and Supplement Fig. 4, showing the separation of peptides after mild acid hydrolysis of CNBr-cleavage fragment CB3. PMID:2106878

  11. The amino acid sequence of chymopapain from Carica papaya.

    PubMed

    Watson, D C; Yaguchi, M; Lynn, K R

    1990-02-15

    Chymopapain is a polypeptide of 218 amino acid residues. It has considerable structural similarity with papain and papaya proteinase omega, including conservation of the catalytic site and of the disulphide bonding. Chymopapain is like papaya proteinase omega in carrying four extra residues between papain positions 168 and 169, but differs from both papaya proteinases in the composition of its S2 subsite, as well as in having a second thiol group, Cys-117. Some evidence for the amino acid sequence of chymopapain has been deposited as Supplementary Publication SUP 50153 (12 pages) at the British Library Document Supply Centre, Boston Spa., Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1990) 265, 5. The information comprises Supplement Tables 1-4, which contain, in order, amino acid compositions of peptides from tryptic, peptic, CNBr and mild acid cleavages, Supplement Fig. 1, showing re-fractionation of selected peaks from Fig. 2 of the main paper. Supplement Fig. 2, showing cation-exchange chromatography of the earliest-eluted peak of Fig. 3 of the main paper, Supplement Fig. 3, showing reverse-phase h.p.l.c. of the later-eluted peak from Fig. 3 of the main paper, and Supplement Fig. 4, showing the separation of peptides after mild acid hydrolysis of CNBr-cleavage fragment CB3. PMID:2106878

  12. Solid phase sequencing of double-stranded nucleic acids

    DOEpatents

    Fu, Dong-Jing; Cantor, Charles R.; Koster, Hubert; Smith, Cassandra L.

    2002-01-01

    This invention relates to methods for detecting and sequencing of target double-stranded nucleic acid sequences, to nucleic acid probes and arrays of probes useful in these methods, and to kits and systems which contain these probes. Useful methods involve hybridizing the nucleic acids or nucleic acids which represent complementary or homologous sequences of the target to an array of nucleic acid probes. These probe comprise a single-stranded portion, an optional double-stranded portion and a variable sequence within the single-stranded portion. The molecular weights of the hybridized nucleic acids of the set can be determined by mass spectroscopy, and the sequence of the target determined from the molecular weights of the fragments. Nucleic acids whose sequences can be determined include nucleic acids in biological samples such as patient biopsies and environmental samples. Probes may be fixed to a solid support such as a hybridization chip to facilitate automated determination of molecular weights and identification of the target sequence.

  13. Analysis and Annotation of Nucleic Acid Sequence

    SciTech Connect

    States, David J.

    2004-07-28

    The aims of this project were to develop improved methods for computational genome annotation and to apply these methods to improve the annotation of genomic sequence data with a specific focus on human genome sequencing. The project resulted in a substantial body of published work. Notable contributions of this project were the identification of basecalling and lane tracking as error processes in genome sequencing and contributions to improved methods for these steps in genome sequencing. This technology improved the accuracy and throughput of genome sequence analysis. Probabilistic methods for physical map construction were developed. Improved methods for sequence alignment, alternative splicing analysis, promoter identification and NF kappa B response gene prediction were also developed.

  14. Analysis and Annotation of Nucleic Acid Sequence

    SciTech Connect

    David J. States

    1998-08-01

    The aims of this project were to develop improved methods for computational genome annotation and to apply these methods to improve the annotation of genomic sequence data with a specific focus on human genome sequencing. The project resulted in a substantial body of published work. Notable contributions of this project were the identification of basecalling and lane tracking as error processes in genome sequencing and contributions to improved methods for these steps in genome sequencing. This technology improved the accuracy and throughput of genome sequence analysis. Probabilistic methods for physical map construction were developed. Improved methods for sequence alignment, alternative splicing analysis, promoter identification and NF kappa B response gene prediction were also developed.

  15. Matrix genes of measles virus and canine distemper virus: cloning, nucleotide sequences, and deduced amino acid sequences.

    PubMed Central

    Bellini, W J; Englund, G; Richardson, C D; Rozenblatt, S; Lazzarini, R A

    1986-01-01

    The nucleotide sequences encoding the matrix (M) proteins of measles virus (MV) and canine distemper virus (CDV) were determined from cDNA clones containing these genes in their entirety. In both cases, single open reading frames specifying basic proteins of 335 amino acid residues were predicted from the nucleotide sequences. Both viral messages were composed of approximately 1,450 nucleotides and contained 400 nucleotides of presumptive noncoding sequences at their respective 3' ends. MV and CDV M-protein-coding regions were 67% homologous at the nucleotide level and 76% homologous at the amino acid level. Only chance homology was observed in the 400-nucleotide trailer sequences. Comparisons of the M protein sequences of MV and CDV with the sequence reported for Sendai virus (B. M. Blumberg, K. Rose, M. G. Simona, L. Roux, C. Giorgi, and D. Kolakofsky, J. Virol. 52:656-663; Y. Hidaka, T. Kanda, K. Iwasaki, A. Nomoto, T. Shioda, and H. Shibuta, Nucleic Acids Res. 12:7965-7973) indicated the greatest homology among these M proteins in the carboxyterminal third of the molecule. Secondary-structure analyses of this shared region indicated a structurally conserved, hydrophobic sequence which possibly interacted with the lipid bilayer. Images PMID:3754588

  16. From Artificial Amino Acids to Sequence-Defined Targeted Oligoaminoamides.

    PubMed

    Morys, Stephan; Wagner, Ernst; Lächelt, Ulrich

    2016-01-01

    Artificial oligoamino acids with appropriate protecting groups can be used for the sequential assembly of oligoaminoamides on solid-phase. With the help of these oligoamino acids multifunctional nucleic acid (NA) carriers can be designed and produced in highly defined topologies. Here we describe the synthesis of the artificial oligoamino acid Fmoc-Stp(Boc3)-OH, the subsequent assembly into sequence-defined oligomers and the formulation of tumor-targeted plasmid DNA (pDNA) polyplexes. PMID:27436323

  17. Detecting frame shifts by amino acid sequence comparison.

    PubMed

    Claverie, J M

    1993-12-20

    Various amino acid substitution scoring matrices are used in conjunction with local alignments programs to detect regions of similarity and infer potential common ancestry between proteins. The usual scoring schemes derive from the implicit hypothesis that related proteins evolve from a common ancestor by the accumulation of point mutations and that amino acids tend to be progressively substituted by others with similar properties. However, other frequent single mutation events, like nucleotide insertion or deletion and gene inversion, change the translation reading frame and cause previously encoded amino acid sequences to become unrecognizable at once. Here, I derive five new types of scoring matrix, each capable of detecting a specific frame shift (deletion, insertion and inversion in 3 frames) and use them with a regular local alignments program to detect amino acid sequences that may have derived from alternative reading frames of the same nucleotide sequence. Frame shifts are inferred from the sole comparison of the protein sequences. The five scoring matrices were used with the BLASTP program to compare all the protein sequences in the Swissprot database. Surprisingly, the searches revealed hundreds of highly significant frame shift matches, of which many are likely to represent sequencing errors. Others provide some evidence that frame shift mutations might be used in protein evolution as a way to create new amino acid sequences from pre-existing coding regions. PMID:7903399

  18. Segments of amino acid sequence similarity in beta-amylases.

    PubMed

    Friedberg, F; Rhodes, C

    1988-01-01

    In alpha-amylases from animals, plants and bacteria and in beta-amylases from plants and bacteria a number of segments exhibit amino acid sequence similarity specific to the alpha or to the beta type, respectively. In the case of the beta-amylases the similar sequence regions are extensive and they are disrupted only by short interspersed dissimilar regions. Close to the C terminus, however, no such sequence similarity exist. PMID:2464171

  19. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... acids are not intended to be embraced by this definition. Any amino acid sequence that contains post-translationally modified amino acids may be described as the amino acid sequence that is initially translated... sequence of four or more amino acids or an unbranched sequence of ten or more nucleotides....

  20. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... acids are not intended to be embraced by this definition. Any amino acid sequence that contains post-translationally modified amino acids may be described as the amino acid sequence that is initially translated... sequence of four or more amino acids or an unbranched sequence of ten or more nucleotides....

  1. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... acids are not intended to be embraced by this definition. Any amino acid sequence that contains post-translationally modified amino acids may be described as the amino acid sequence that is initially translated... sequence of four or more amino acids or an unbranched sequence of ten or more nucleotides....

  2. A method to find palindromes in nucleic acid sequences.

    PubMed

    Anjana, Ramnath; Shankar, Mani; Vaishnavi, Marthandan Kirti; Sekar, Kanagaraj

    2013-01-01

    Various types of sequences in the human genome are known to play important roles in different aspects of genomic functioning. Among these sequences, palindromic nucleic acid sequences are one such type that have been studied in detail and found to influence a wide variety of genomic characteristics. For a nucleotide sequence to be considered as a palindrome, its complementary strand must read the same in the opposite direction. For example, both the strands i.e the strand going from 5' to 3' and its complementary strand from 3' to 5' must be complementary. A typical nucleotide palindromic sequence would be TATA (5' to 3') and its complimentary sequence from 3' to 5' would be ATAT. Thus, a new method has been developed using dynamic programming to fetch the palindromic nucleic acid sequences. The new method uses less memory and thereby it increases the overall speed and efficiency. The proposed method has been tested using the bacterial (3891 KB bases) and human chromosomal sequences (Chr-18: 74366 kb and Chr-Y: 25554 kb) and the computation time for finding the palindromic sequences is in milli seconds. PMID:23515654

  3. N-terminal sequence of amino acids and some properties of an acid-stable alpha-amylase from citric acid-koji (Aspergillus usamii var.).

    PubMed

    Suganuma, T; Tahara, N; Kitahara, K; Nagahama, T; Inuzuka, K

    1996-01-01

    An acid-stable alpha-amylase (AA) was purified from an acidic extract of citric acid-koji (A. usamii var.). The N-terminal sequence of the first 20 amino acids of the enzyme was identical with that of AA from A. niger, but the two enzymes differed in molecular weight. HPLC analysis for identifying the anomers of products indicated that the AA hydrolyzed maltopentaose (G5) at the third glycoside bond predominantly, which differed from Taka-amylase A and the neutral alpha-amylase (NA) from the citric acid-koji. PMID:8824843

  4. Amino acid sequence repertoire of the bacterial proteome and the occurrence of untranslatable sequences.

    PubMed

    Navon, Sharon Penias; Kornberg, Guy; Chen, Jin; Schwartzman, Tali; Tsai, Albert; Puglisi, Elisabetta Viani; Puglisi, Joseph D; Adir, Noam

    2016-06-28

    Bioinformatic analysis of Escherichia coli proteomes revealed that all possible amino acid triplet sequences occur at their expected frequencies, with four exceptions. Two of the four underrepresented sequences (URSs) were shown to interfere with translation in vivo and in vitro. Enlarging the URS by a single amino acid resulted in increased translational inhibition. Single-molecule methods revealed stalling of translation at the entrance of the peptide exit tunnel of the ribosome, adjacent to ribosomal nucleotides A2062 and U2585. Interaction with these same ribosomal residues is involved in regulation of translation by longer, naturally occurring protein sequences. The E. coli exit tunnel has evidently evolved to minimize interaction with the exit tunnel and maximize the sequence diversity of the proteome, although allowing some interactions for regulatory purposes. Bioinformatic analysis of the human proteome revealed no underrepresented triplet sequences, possibly reflecting an absence of regulation by interaction with the exit tunnel. PMID:27307442

  5. On Quantum Algorithm for Multiple Alignment of Amino Acid Sequences

    NASA Astrophysics Data System (ADS)

    Iriyama, Satoshi; Ohya, Masanori

    2009-02-01

    The alignment of genome sequences or amino acid sequences is one of fundamental operations for the study of life. Usual computational complexity for the multiple alignment of N sequences with common length L by dynamic programming is O(LN). This alignment is considered as one of the NP problems, so that it is desirable to find a nice algorithm of the multiple alignment. Thus in this paper we propose the quantum algorithm for the multiple alignment based on the works12,1,2 in which the NP complete problem was shown to be the P problem by means of quantum algorithm and chaos information dynamics.

  6. Prebiotically plausible mechanisms increase compositional diversity of nucleic acid sequences

    PubMed Central

    Derr, Julien; Manapat, Michael L.; Rajamani, Sudha; Leu, Kevin; Xulvi-Brunet, Ramon; Joseph, Isaac; Nowak, Martin A.; Chen, Irene A.

    2012-01-01

    During the origin of life, the biological information of nucleic acid polymers must have increased to encode functional molecules (the RNA world). Ribozymes tend to be compositionally unbiased, as is the vast majority of possible sequence space. However, ribonucleotides vary greatly in synthetic yield, reactivity and degradation rate, and their non-enzymatic polymerization results in compositionally biased sequences. While natural selection could lead to complex sequences, molecules with some activity are required to begin this process. Was the emergence of compositionally diverse sequences a matter of chance, or could prebiotically plausible reactions counter chemical biases to increase the probability of finding a ribozyme? Our in silico simulations using a two-letter alphabet show that template-directed ligation and high concatenation rates counter compositional bias and shift the pool toward longer sequences, permitting greater exploration of sequence space and stable folding. We verified experimentally that unbiased DNA sequences are more efficient templates for ligation, thus increasing the compositional diversity of the pool. Our work suggests that prebiotically plausible chemical mechanisms of nucleic acid polymerization and ligation could predispose toward a diverse pool of longer, potentially structured molecules. Such mechanisms could have set the stage for the appearance of functional activity very early in the emergence of life. PMID:22319215

  7. The amino-acid sequence of kangaroo pancreatic ribonuclease.

    PubMed

    Gaastra, W; Welling, G W; Beintema, J J

    1978-05-01

    Red kangaroo (Macropus rufus) ribonuclease was isolated from pancreatic tissue by affinity chromatography. The amino acid sequence was determined by automatic sequencing of overlapping large fragments and by analysis of shorter peptides obtained by digestion with a number of proteolytic enzymes. The polypeptide chain consists of 122 amino acid residues. Compared to other ribonucleases, the N-terminal residue and residue 114 are deleted. In other pancreatic ribonucleases position 114 is occupied by a cis proline residue in an external loop at the surface of the molecule. Other remarkable substitutions are the presence of a tyrosine residue at position 123 instead of a serine which forms a hydrogen bond with the pyrimidine ring of a nucleotide substrate, and a number of hydrophobichydrophilic interchanges in the sequence 51-55, which forms part of an alpha-helix in bovine ribonuclease and exhibits few substitutions in the placental mammals. Kangaroo ribonuclease contains no carbohydrate, although the enzyme possesses a recognition site for carbohydrate attachment in the sequence Asn-Val-Thr (62-64). The enzyme differs at about 35-40% of the positions from all other mammalian pancreatic ribonucleases sequenced to date, which is in agreement with the early divergence between the marsupials and the placental mammals. From fragmentary data a tentative sequence of red-necked wallaby (Macropus rufogriseus) pancreatic ribonuclease has been derived. Eight differences with the kangaroo sequence were found. PMID:658039

  8. Amino acid sequence of Salmonella typhimurium branched-chain amino acid aminotransferase.

    PubMed

    Feild, M J; Nguyen, D C; Armstrong, F B

    1989-06-13

    The complete amino acid sequence of the subunit of branched-chain amino acid aminotransferase (transaminase B, EC 2.6.1.42) of Salmonella typhimurium was determined. An Escherichia coli recombinant containing the ilvGEDAY gene cluster of Salmonella was used as the source of the hexameric enzyme. The peptide fragments used for sequencing were generated by treatment with trypsin, Staphylococcus aureus V8 protease, endoproteinase Lys-C, and cyanogen bromide. The enzyme subunit contains 308 residues and has a molecular weight of 33,920. To determine the coenzyme-binding site, the pyridoxal 5-phosphate containing enzyme was treated with tritiated sodium borohydride prior to trypsin digestion. Peptide map comparisons with an apoenzyme tryptic digest and monitoring radioactivity incorporation allowed identification of the pyridoxylated peptide, which was then isolated and sequenced. The coenzyme-binding site is the lysyl residue at position 159. The amino acid sequence of Salmonella transaminase B is 97.4% identical with that of Escherichia coli, differing in only eight amino acid positions. Sequence comparisons of transaminase B to other known aminotransferase sequences revealed limited sequence similarity (24-33%) when conserved amino acid substitutions are allowed and alignments were forced to occur on the coenzyme-binding site. PMID:2669973

  9. Amino acid sequence of bovine heart coupling factor 6.

    PubMed Central

    Fang, J K; Jacobs, J W; Kanner, B I; Racker, E; Bradshaw, R A

    1984-01-01

    The amino acid sequence of bovine heart mitochondrial coupling factor 6 (F6) has been determined by automated Edman degradation of the whole protein and derived peptides. Preparations based on heat precipitation and ethanol extraction showed allotypic variation at three positions while material further purified by HPLC yielded only one sequence that also differed by a Phe-Thr replacement at residue 62. The mature protein contains 76 amino acids with a calculated molecular weight of 9006 and a pI of approximately equal to 5, in good agreement with experimentally measured values. The charged amino acids are mainly clustered at the termini and in one section in the middle; these three polar segments are separated by two segments relatively rich in nonpolar residues. Chou-Fasman analysis suggests three stretches of alpha-helix coinciding (or within) the high-charge-density sequences with a single beta-turn at the first polar-nonpolar junction. Comparison of the F6 sequence with those of other proteins did not reveal any homologous structures. PMID:6149548

  10. Sequences Of Amino Acids For Human Serum Albumin

    NASA Technical Reports Server (NTRS)

    Carter, Daniel C.

    1992-01-01

    Sequences of amino acids defined for use in making polypeptides one-third to one-sixth as large as parent human serum albumin molecule. Smaller, chemically stable peptides have diverse applications including service as artificial human serum and as active components of biosensors and chromatographic matrices. In applications involving production of artificial sera from new sequences, little or no concern about viral contaminants. Smaller genetically engineered polypeptides more easily expressed and produced in large quantities, making commercial isolation and production more feasible and profitable.

  11. Nanopores and nucleic acids: prospects for ultrarapid sequencing

    NASA Technical Reports Server (NTRS)

    Deamer, D. W.; Akeson, M.

    2000-01-01

    DNA and RNA molecules can be detected as they are driven through a nanopore by an applied electric field at rates ranging from several hundred microseconds to a few milliseconds per molecule. The nanopore can rapidly discriminate between pyrimidine and purine segments along a single-stranded nucleic acid molecule. Nanopore detection and characterization of single molecules represents a new method for directly reading information encoded in linear polymers. If single-nucleotide resolution can be achieved, it is possible that nucleic acid sequences can be determined at rates exceeding a thousand bases per second.

  12. Ab initio detection of fuzzy amino acid tandem repeats in protein sequences

    PubMed Central

    2012-01-01

    Background Tandem repetitions within protein amino acid sequences often correspond to regular secondary structures and form multi-repeat 3D assemblies of varied size and function. Developing internal repetitions is one of the evolutionary mechanisms that proteins employ to adapt their structure and function under evolutionary pressure. While there is keen interest in understanding such phenomena, detection of repeating structures based only on sequence analysis is considered an arduous task, since structure and function is often preserved even under considerable sequence divergence (fuzzy tandem repeats). Results In this paper we present PTRStalker, a new algorithm for ab-initio detection of fuzzy tandem repeats in protein amino acid sequences. In the reported results we show that by feeding PTRStalker with amino acid sequences from the UniProtKB/Swiss-Prot database we detect novel tandemly repeated structures not captured by other state-of-the-art tools. Experiments with membrane proteins indicate that PTRStalker can detect global symmetries in the primary structure which are then reflected in the tertiary structure. Conclusions PTRStalker is able to detect fuzzy tandem repeating structures in protein sequences, with performance beyond the current state-of-the art. Such a tool may be a valuable support to investigating protein structural properties when tertiary X-ray data is not available. PMID:22536906

  13. Quantum-Sequencing: Biophysics of quantum tunneling through nucleic acids

    NASA Astrophysics Data System (ADS)

    Casamada Ribot, Josep; Chatterjee, Anushree; Nagpal, Prashant

    2014-03-01

    Tunneling microscopy and spectroscopy has extensively been used in physical surface sciences to study quantum tunneling to measure electronic local density of states of nanomaterials and to characterize adsorbed species. Quantum-Sequencing (Q-Seq) is a new method based on tunneling microscopy for electronic sequencing of single molecule of nucleic acids. A major goal of third-generation sequencing technologies is to develop a fast, reliable, enzyme-free single-molecule sequencing method. Here, we present the unique ``electronic fingerprints'' for all nucleotides on DNA and RNA using Q-Seq along their intrinsic biophysical parameters. We have analyzed tunneling spectra for the nucleotides at different pH conditions and analyzed the HOMO, LUMO and energy gap for all of them. In addition we show a number of biophysical parameters to further characterize all nucleobases (electron and hole transition voltage and energy barriers). These results highlight the robustness of Q-Seq as a technique for next-generation sequencing.

  14. Correlation between fibroin amino acid sequence and physical silk properties.

    PubMed

    Fedic, Robert; Zurovec, Michal; Sehnal, Frantisek

    2003-09-12

    The fiber properties of lepidopteran silk depend on the amino acid repeats that interact during H-fibroin polymerization. The aim of our research was to relate repeat composition to insect biology and fiber strength. Representative regions of the H-fibroin genes were sequenced and analyzed in three pyralid species: wax moth (Galleria mellonella), European flour moth (Ephestia kuehniella), and Indian meal moth (Plodia interpunctella). The amino acid repeats are species-specific, evidently a diversification of an ancestral region of 43 residues, and include three types of regularly dispersed motifs: modifications of GSSAASAA sequence, stretches of tripeptides GXZ where X and Z represent bulky residues, and sequences similar to PVIVIEE. No concatenations of GX dipeptide or alanine, which are typical for Bombyx silkworms and Antheraea silk moths, respectively, were found. Despite different repeat structure, the silks of G. mellonella and E. kuehniella exhibit similar tensile strength as the Bombyx and Antheraea silks. We suggest that in these latter two species, variations in the repeat length obstruct repeat alignment, but sufficiently long stretches of iterated residues get superposed to interact. In the pyralid H-fibroins, interactions of the widely separated and diverse motifs depend on the precision of repeat matching; silk is strong in G. mellonella and E. kuehniella, with 2-3 types of long homogeneous repeats, and nearly 10 times weaker in P. interpunctella, with seven types of shorter erratic repeats. The high proportion of large amino acids in the H-fibroin of pyralids has probably evolved in connection with the spinning habit of caterpillars that live in protective silk tubes and spin continuously, enlarging the tubes on one end and partly devouring the other one. The silk serves as a depot of energetically rich and essential amino acids that may be scarce in the diet. PMID:12816957

  15. Nucleic acid sequence detection using multiplexed oligonucleotide PCR

    DOEpatents

    Nolan, John P.; White, P. Scott

    2006-12-26

    Methods for rapidly detecting single or multiple sequence alleles in a sample nucleic acid are described. Provided are all of the oligonucleotide pairs capable of annealing specifically to a target allele and discriminating among possible sequences thereof, and ligating to each other to form an oligonucleotide complex when a particular sequence feature is present (or, alternatively, absent) in the sample nucleic acid. The design of each oligonucleotide pair permits the subsequent high-level PCR amplification of a specific amplicon when the oligonucleotide complex is formed, but not when the oligonucleotide complex is not formed. The presence or absence of the specific amplicon is used to detect the allele. Detection of the specific amplicon may be achieved using a variety of methods well known in the art, including without limitation, oligonucleotide capture onto DNA chips or microarrays, oligonucleotide capture onto beads or microspheres, electrophoresis, and mass spectrometry. Various labels and address-capture tags may be employed in the amplicon detection step of multiplexed assays, as further described herein.

  16. The amino acid sequence of rabbit muscle triose phosphate isomerase.

    PubMed Central

    Corran, P H; Waley, S G

    1975-01-01

    The amino acid sequence of rabbit muscle triose phosphate isomerase was deduced by characterizing peptides that overlap the tryptic peptides. Thiol groups were modified by oxidation, carboxymethylation or aminoen. About 50 peptides that provided information about overlaps were isolated; the peptides were mostly characterized by their compositions and N-terminal residues. The peptide chains contain 248 amino acid residues, and no evidence for dissimilarity of the two subunits that comprise the native enzyme was found. The sequence of the rabbit muscle enzyme may be compared with that of the coelacanth enzyme (Kolb et al., 1974): 84% of the residues are in identical positions. Similarly, comparison of the sequence with that inferred for the chicken enzyme (Furth et al., 1974) shows that 87% of the residues are in identical positions. Limited though these comparisons are, they suggest that triose phosphate isomerase has one of the lowest rates of evolutionary change. An extended version of the present paper has been deposited as Supplementary Publication SUP 50040 (42 pages) at the British Library (Lending Division) (formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms given in Biochem. J. (1975) 145, 5. PMID:1171682

  17. The Complete Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis ssp. lactis IL1403

    PubMed Central

    Bolotin, Alexander; Wincker, Patrick; Mauger, Stéphane; Jaillon, Olivier; Malarme, Karine; Weissenbach, Jean; Ehrlich, S. Dusko; Sorokin, Alexei

    2001-01-01

    Lactococcus lactis is a nonpathogenic AT-rich gram-positive bacterium closely related to the genus Streptococcus and is the most commonly used cheese starter. It is also the best-characterized lactic acid bacterium. We sequenced the genome of the laboratory strain IL1403, using a novel two-step strategy that comprises diagnostic sequencing of the entire genome and a shotgun polishing step. The genome contains 2,365,589 base pairs and encodes 2310 proteins, including 293 protein-coding genes belonging to six prophages and 43 insertion sequence (IS) elements. Nonrandom distribution of IS elements indicates that the chromosome of the sequenced strain may be a product of recent recombination between two closely related genomes. A complete set of late competence genes is present, indicating the ability of L. lactis to undergo DNA transformation. Genomic sequence revealed new possibilities for fermentation pathways and for aerobic respiration. It also indicated a horizontal transfer of genetic information from Lactococcus to gram-negative enteric bacteria of Salmonella-Escherichia group. [The sequence data described in this paper has been submitted to the GenBank data library under accession no. AE005176.] PMID:11337471

  18. Antibody-specific model of amino acid substitution for immunological inferences from alignments of antibody sequences.

    PubMed

    Mirsky, Alexander; Kazandjian, Linda; Anisimova, Maria

    2015-03-01

    Antibodies are glycoproteins produced by the immune system as a dynamically adaptive line of defense against invading pathogens. Very elegant and specific mutational mechanisms allow B lymphocytes to produce a large and diversified repertoire of antibodies, which is modified and enhanced throughout all adulthood. One of these mechanisms is somatic hypermutation, which stochastically mutates nucleotides in the antibody genes, forming new sequences with different properties and, eventually, higher affinity and selectivity to the pathogenic target. As somatic hypermutation involves fast mutation of antibody sequences, this process can be described using a Markov substitution model of molecular evolution. Here, using large sets of antibody sequences from mice and humans, we infer an empirical amino acid substitution model AB, which is specific to antibody sequences. Compared with existing general amino acid models, we show that the AB model provides significantly better description for the somatic evolution of mice and human antibody sequences, as demonstrated on large next generation sequencing (NGS) antibody data. General amino acid models are reflective of conservation at the protein level due to functional constraints, with most frequent amino acids exchanges taking place between residues with the same or similar physicochemical properties. In contrast, within the variable part of antibody sequences we observed an elevated frequency of exchanges between amino acids with distinct physicochemical properties. This is indicative of a sui generis mutational mechanism, specific to antibody somatic hypermutation. We illustrate this property of antibody sequences by a comparative analysis of the network modularity implied by the AB model and general amino acid substitution models. We recommend using the new model for computational studies of antibody sequence maturation, including inference of alignments and phylogenetic trees describing antibody somatic hypermutation in

  19. Amino acid sequence prerequisites for the formation of cn ions.

    PubMed

    Downard, K M; Biemann, K

    1993-11-01

    Ammo acid sequence prerequisites are described for the formation of c, ions observed in high-energy collision-induced decomposition spectra of peptides. It is shown that the formation of cn ions is promoted by the nature of the amino acid C-terminal to the cleavage site. A propensity for cn cleavage preceding threonine, and to a lesser extent tryptophan, lysine, and serine, is demonstrated where fragmentation is directed N-terminally at these residues. In addition, the nature of the residue N-terminal to the cleavage site is shown to have little effect on cn ion formation. A mechanism for cn ion formation is proposed and its applicability to the results observed is discussed. PMID:24227531

  20. Ultrasensitive nucleic acid sequence detection by single-molecule electrophoresis

    SciTech Connect

    Castro, A; Shera, E.B.

    1996-09-01

    This is the final report of a one-year laboratory-directed research and development project at Los Alamos National Laboratory. There has been considerable interest in the development of very sensitive clinical diagnostic techniques over the last few years. Many pathogenic agents are often present in extremely small concentrations in clinical samples, especially at the initial stages of infection, making their detection very difficult. This project sought to develop a new technique for the detection and accurate quantification of specific bacterial and viral nucleic acid sequences in clinical samples. The scheme involved the use of novel hybridization probes for the detection of nucleic acids combined with our recently developed technique of single-molecule electrophoresis. This project is directly relevant to the DOE`s Defense Programs strategic directions in the area of biological warfare counter-proliferation.

  1. Characterization of the microbial acid mine drainage microbial community using culturing and direct sequencing techniques.

    PubMed

    Auld, Ryan R; Myre, Maxine; Mykytczuk, Nadia C S; Leduc, Leo G; Merritt, Thomas J S

    2013-05-01

    We characterized the bacterial community from an AMD tailings pond using both classical culturing and modern direct sequencing techniques and compared the two methods. Acid mine drainage (AMD) is produced by the environmental and microbial oxidation of minerals dissolved from mining waste. Surprisingly, we know little about the microbial communities associated with AMD, despite the fundamental ecological roles of these organisms and large-scale economic impact of these waste sites. AMD microbial communities have classically been characterized by laboratory culturing-based techniques and more recently by direct sequencing of marker gene sequences, primarily the 16S rRNA gene. In our comparison of the techniques, we find that their results are complementary, overall indicating very similar community structure with similar dominant species, but with each method identifying some species that were missed by the other. We were able to culture the majority of species that our direct sequencing results indicated were present, primarily species within the Acidithiobacillus and Acidiphilium genera, although estimates of relative species abundance were only obtained from direct sequencing. Interestingly, our culture-based methods recovered four species that had been overlooked from our sequencing results because of the rarity of the marker gene sequences, likely members of the rare biosphere. Further, direct sequencing indicated that a single genus, completely missed in our culture-based study, Legionella, was a dominant member of the microbial community. Our results suggest that while either method does a reasonable job of identifying the dominant members of the AMD microbial community, together the methods combine to give a more complete picture of the true diversity of this environment. PMID:23485423

  2. Carboxylic Acids as Indicators of Parent Body Conditions

    NASA Technical Reports Server (NTRS)

    Lerner N. R.; Chang, Sherwood (Technical Monitor)

    1995-01-01

    Alpha-hydroxy and alpha-amino carboxylic acids found on the Murchison meteorite are deuterium enriched. It is postulated that they arose from a common interstellar scurce: the reaction of carbonyl compounds in an aqueous mixture containing HCN and NH3. Carbonyl compounds react with HCN to form alpha-hydroxy nitriles, RR'CO + HCN right and left arrow RR'C(OH)CN. If ammonia is also present, the alpha-hydroxy nitriles will exist in equilibrium with the alpha-amino nitriles, RR'C(OH)CN + NH3 right and left arrow - RRCNH2CN + H2O. Both nitrites are hydrolyzed by water to form carboxylic acids: RR'C(OH)CN + H2O yields RR'C(OH)CO2H and RR'C(NH2)CN + H2O yields RR'C(NH2)CO2H.

  3. Nucleotide sequence of the phosphoglycerate kinase gene from the extreme thermophile Thermus thermophilus. Comparison of the deduced amino acid sequence with that of the mesophilic yeast phosphoglycerate kinase.

    PubMed Central

    Bowen, D; Littlechild, J A; Fothergill, J E; Watson, H C; Hall, L

    1988-01-01

    Using oligonucleotide probes derived from amino acid sequencing information, the structural gene for phosphoglycerate kinase from the extreme thermophile, Thermus thermophilus, was cloned in Escherichia coli and its complete nucleotide sequence determined. The gene consists of an open reading frame corresponding to a protein of 390 amino acid residues (calculated Mr 41,791) with an extreme bias for G or C (93.1%) in the codon third base position. Comparison of the deduced amino acid sequence with that of the corresponding mesophilic yeast enzyme indicated a number of significant differences. These are discussed in terms of the unusual codon bias and their possible role in enhanced protein thermal stability. Images Fig. 1. PMID:3052437

  4. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... in the sequence. (4) The enumeration of amino acids may start at the first amino acid of the first..., counting backwards starting with the amino acid next to number 1. Otherwise, the enumeration of amino acids... sequence every 5 amino acids. The enumeration method for amino acid sequences that is set forth......

  5. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... in the sequence. (4) The enumeration of amino acids may start at the first amino acid of the first..., counting backwards starting with the amino acid next to number 1. Otherwise, the enumeration of amino acids... sequence every 5 amino acids. The enumeration method for amino acid sequences that is set forth......

  6. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... in the sequence. (4) The enumeration of amino acids may start at the first amino acid of the first..., counting backwards starting with the amino acid next to number 1. Otherwise, the enumeration of amino acids... sequence every 5 amino acids. The enumeration method for amino acid sequences that is set forth......

  7. The amino acid sequence of ribonuclease U2 from Ustilago sphaerogena.

    PubMed Central

    Sato, S; Uchida, T

    1975-01-01

    1. RNAase (ribonuclease) U2, a purine-specific RNAase, was reduced, aminoethylated and hydrolysed with trypsin, chymotrypsin and thermolysin. On the basis of the analyses of the resulting peptides, the complete amino acid sequence of RNAase U2 was determined, 2. When the sequence was compared with the amino acid sequence of RNAase T1 (EC 3.1.4.8), the following regions were found to be similar in the two enzymes; Tyr-Pro-His-Gln-Tyr (38-42) in RNAase U2 and Tyr-Pro-His-Lys-Tyr (38-42) in RNAase T1, Glu-Phe-Pro-Leu-Val (61-65) in RNAase U2 and Glu-Trp-Pro-Ile-Leu (58-62) in RNAase T1, Asp-Arg-Val-Ile-Tyr-Gln (83-88) in RNAase U2 and Asp-Arg-Val-Phe-Asn (76-81) in RNAase T1 and Val-Thr-His-Thr-Gly-Ala (98-103) in RNAase U2 and Ile-Thr-His-Thr-Gly-Ala (90-95) in RNAase T1. All of the amino acid residues, histidine-40, glutamate-58, arginine-77 and histidine-92, which were found to play a crucial role in the biological activity of RNAase T1, were included in the regions cited here. 3. Detailed evidence for the amino acid sequence of the sequence of the proteins has been deposited as Supplementary Publication SUP 50041 (33 PAGES) AT THE British Library (Lending Division)(formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1975), 145, 5. PMID:1156364

  8. Predicting protein disorder by analyzing amino acid sequence

    PubMed Central

    Yang, Jack Y; Yang, Mary Qu

    2008-01-01

    Background Many protein regions and some entire proteins have no definite tertiary structure, presenting instead as dynamic, disorder ensembles under different physiochemical circumstances. These proteins and regions are known as Intrinsically Unstructured Proteins (IUP). IUP have been associated with a wide range of protein functions, along with roles in diseases characterized by protein misfolding and aggregation. Results Identifying IUP is important task in structural and functional genomics. We exact useful features from sequences and develop machine learning algorithms for the above task. We compare our IUP predictor with PONDRs (mainly neural-network-based predictors), disEMBL (also based on neural networks) and Globplot (based on disorder propensity). Conclusion We find that augmenting features derived from physiochemical properties of amino acids (such as hydrophobicity, complexity etc.) and using ensemble method proved beneficial. The IUP predictor is a viable alternative software tool for identifying IUP protein regions and proteins. PMID:18831799

  9. On human disease-causing amino acid variants: statistical study of sequence and structural patterns

    PubMed Central

    Alexov, Emil

    2015-01-01

    Statistical analysis was carried out on large set of naturally occurring human amino acid variations and it was demonstrated that there is a preference for some amino acid substitutions to be associated with diseases. At an amino acid sequence level, it was shown that the disease-causing variants frequently involve drastic changes of amino acid physico-chemical properties of proteins such as charge, hydrophobicity and geometry. Structural analysis of variants involved in diseases and being frequently observed in human population showed similar trends: disease-causing variants tend to cause more changes of hydrogen bond network and salt bridges as compared with harmless amino acid mutations. Analysis of thermodynamics data reported in literature, both experimental and computational, indicated that disease-causing variants tend to destabilize proteins and their interactions, which prompted us to investigate the effects of amino acid mutations on large databases of experimentally measured energy changes in unrelated proteins. Although the experimental datasets were linked neither to diseases nor exclusory to human proteins, the observed trends were the same: amino acid mutations tend to destabilize proteins and their interactions. Having in mind that structural and thermodynamics properties are interrelated, it is pointed out that any large change of any of them is anticipated to cause a disease. PMID:25689729

  10. Structural gene and complete amino acid sequence of Pseudomonas aeruginosa IFO 3455 elastase.

    PubMed Central

    Fukushima, J; Yamamoto, S; Morihara, K; Atsumi, Y; Takeuchi, H; Kawamoto, S; Okuda, K

    1989-01-01

    The DNA encoding the elastase of Pseudomonas aeruginosa IFO 3455 was cloned, and its complete nucleotide sequence was determined. When the cloned gene was ligated to pUC18, the Escherichia coli expression vector, bacteria carrying the gene exhibited high levels of both elastase activity and elastase antigens. The amino acid sequence, deduced from the nucleotide sequence, revealed that the mature elastase consisted of 301 amino acids with a relative molecular mass of 32,926 daltons. The amino acid composition predicted from the DNA sequence was quite similar to the chemically determined composition of purified elastase reported previously. We also observed nucleotide sequence encoding a signal peptide and "pro" sequence consisting of 197 amino acids upstream from the mature elastase protein gene. The amino acid sequence analysis revealed that both the N-terminal sequence of the purified elastase and the N-terminal side sequences of the C-terminal tryptic peptide as well as the internal lysyl peptide fragment were completely identical to the deduced amino acid sequences. The pattern of identity of amino acid sequences was quite evident in the regions that include structurally and functionally important residues of Bacillus subtilis thermolysin. PMID:2493453

  11. Human retroviruses and AIDS 1996. A compilation and analysis of nucleic acid and amino acid sequences

    SciTech Connect

    Myers, G.; Foley, B.; Korber, B.; Mellors, J.W.; Jeang, K.T.; Wain-Hobson, S.

    1997-04-01

    This compendium and the accompanying floppy diskettes are the result of an effort to compile and rapidly publish all relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses. The scope of the compendium and database is best summarized by the five parts that it comprises: (1) Nuclear Acid Alignments and Sequences; (2) Amino Acid Alignments; (3) Analysis; (4) Related Sequences; and (5) Database Communications. Information within all the parts is updated throughout the year on the Web site, http://hiv-web.lanl.gov. While this publication could take the form of a review or sequence monograph, it is not so conceived. Instead, the literature from which the database is derived has simply been summarized and some elementary computational analyses have been performed upon the data. Interpretation and commentary have been avoided insofar as possible so that the reader can form his or her own judgments concerning the complex information. In addition to the general descriptions of the parts of the compendium, the user should read the individual introductions for each part.

  12. Natural vs. random protein sequences: Discovering combinatorics properties on amino acid words.

    PubMed

    Santoni, Daniele; Felici, Giovanni; Vergni, Davide

    2016-02-21

    Casual mutations and natural selection have driven the evolution of protein amino acid sequences that we observe at present in nature. The question about which is the dominant force of proteins evolution is still lacking of an unambiguous answer. Casual mutations tend to randomize protein sequences while, in order to have the correct functionality, one expects that selection mechanisms impose rigid constraints on amino acid sequences. Moreover, one also has to consider that the space of all possible amino acid sequences is so astonishingly large that it could be reasonable to have a well tuned amino acid sequence indistinguishable from a random one. In order to study the possibility to discriminate between random and natural amino acid sequences, we introduce different measures of association between pairs of amino acids in a sequence, and apply them to a dataset of 1047 natural protein sequences and 10,470 random sequences, carefully generated in order to preserve the relative length and amino acid distribution of the natural proteins. We analyze the multidimensional measures with machine learning techniques and show that, to a reasonable extent, natural protein sequences can be differentiated from random ones. PMID:26656109

  13. Transcriptome Sequencing in Response to Salicylic Acid in Salvia miltiorrhiza

    PubMed Central

    Zhang, Xiaoru; Dong, Juane; Liu, Hailong; Wang, Jiao; Qi, Yuexin; Liang, Zongsuo

    2016-01-01

    Salvia miltiorrhiza is a traditional Chinese herbal medicine, whose quality and yield are often affected by diseases and environmental stresses during its growing season. Salicylic acid (SA) plays a significant role in plants responding to biotic and abiotic stresses, but the involved regulatory factors and their signaling mechanisms are largely unknown. In order to identify the genes involved in SA signaling, the RNA sequencing (RNA-seq) strategy was employed to evaluate the transcriptional profiles in S. miltiorrhiza cell cultures. A total of 50,778 unigenes were assembled, in which 5,316 unigenes were differentially expressed among 0-, 2-, and 8-h SA induction. The up-regulated genes were mainly involved in stimulus response and multi-organism process. A core set of candidate novel genes coding SA signaling component proteins was identified. Many transcription factors (e.g., WRKY, bHLH and GRAS) and genes involved in hormone signal transduction were differentially expressed in response to SA induction. Detailed analysis revealed that genes associated with defense signaling, such as antioxidant system genes, cytochrome P450s and ATP-binding cassette transporters, were significantly overexpressed, which can be used as genetic tools to investigate disease resistance. Our transcriptome analysis will help understand SA signaling and its mechanism of defense systems in S. miltiorrhiza. PMID:26808150

  14. Transcriptome Sequencing in Response to Salicylic Acid in Salvia miltiorrhiza.

    PubMed

    Zhang, Xiaoru; Dong, Juane; Liu, Hailong; Wang, Jiao; Qi, Yuexin; Liang, Zongsuo

    2016-01-01

    Salvia miltiorrhiza is a traditional Chinese herbal medicine, whose quality and yield are often affected by diseases and environmental stresses during its growing season. Salicylic acid (SA) plays a significant role in plants responding to biotic and abiotic stresses, but the involved regulatory factors and their signaling mechanisms are largely unknown. In order to identify the genes involved in SA signaling, the RNA sequencing (RNA-seq) strategy was employed to evaluate the transcriptional profiles in S. miltiorrhiza cell cultures. A total of 50,778 unigenes were assembled, in which 5,316 unigenes were differentially expressed among 0-, 2-, and 8-h SA induction. The up-regulated genes were mainly involved in stimulus response and multi-organism process. A core set of candidate novel genes coding SA signaling component proteins was identified. Many transcription factors (e.g., WRKY, bHLH and GRAS) and genes involved in hormone signal transduction were differentially expressed in response to SA induction. Detailed analysis revealed that genes associated with defense signaling, such as antioxidant system genes, cytochrome P450s and ATP-binding cassette transporters, were significantly overexpressed, which can be used as genetic tools to investigate disease resistance. Our transcriptome analysis will help understand SA signaling and its mechanism of defense systems in S. miltiorrhiza. PMID:26808150

  15. Control region sequences indicate that multiple externae represent multiple infections by Sacculina carcini (Cirripedia: Rhizocephala).

    PubMed

    Rees, David; Glenner, Henrik

    2014-08-01

    The rhizocephalan barnacle, Sacculina carcini, is a common parasite of the European shore crab, Carcinus maenas, in which it causes significant detrimental physical and behavioral modifications. In the vast majority of cases, the external portion of the parasite is present in the form of a single sac-like externa; in rare cases, double or even triple externae may occur on the same individual host. Here, we use a highly variable DNA marker, the mitochondrial control region (CR), to investigate whether multiple externae in S. carcini represent infection by multiple parasites or asexual cloning developed by a single parasite individual. Sequences for multiple externae from C. maenas hosts from the Danish inlet, Limfjorden, and from the mud flates at Roscoff, France, were compared. In almost all cases, double or triple externae from an individual host yielded different haplotypes. In the few cases where identical haplotypes were identified from externae on a multiple-infected host, this always represented the most commonly found haplotype in the population. This indicates that in Sacculina carcini, the presence of multiple externae on a single host reflects infection by different individual parasites. A haplotype network of CR sequences also suggests a degree of geographical partitioning, with no shared haplotypes between the Limfjorden and Roscoff. Our data represent the first complete CR sequences for a rhizocephalan, and a unique gene order was also revealed. Although the utility of CR sequences for population-level work must be investigated further, the CR has proved a simple to use and highly variable marker for studies of S. carcini and can easily be applied to a variety of studies in this important parasite. PMID:25473481

  16. Detection of Nucleic Acids with Graphene Nanopores: Ab Initio Characterization of a Novel Sequencing Device

    NASA Astrophysics Data System (ADS)

    Nelson, Tammie; Zhang, Bo; Prezhdo, Oleg

    2010-03-01

    We report an ab initio study of the interaction of two nucleobases, cytosine and adenine, with a novel graphene nanopore device for detecting the base sequence of a single-stranded nucleic acid (ssDNA or RNA). The nucleobases were inserted into a pore in a graphene nanoribbon, and the electrical current and conductance spectra were calculated as functions of voltage applied across the nanoribbon. The conductance spectra and charge densities were analyzed in the presence of each nucleobase in the graphene nanopore. The results indicate that, due to significant differences in the conductance spectra, the proposed device has adequate sensitivity to discriminate between different nucleotides. Moreover, we show that the nucleotide conductance spectra is not affected by its orientation inside the graphene nanopore. The proposed technique may be extremely useful for real applications in developing ultrafast, low cost DNA sequencing methods.

  17. Detection and isolation of nucleic acid sequences using a bifunctional hybridization probe

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    2000-01-01

    A method for detecting and isolating a target sequence in a sample of nucleic acids is provided using a bifunctional hybridization probe capable of hybridizing to the target sequence that includes a detectable marker and a first complexing agent capable of forming a binding pair with a second complexing agent. A kit is also provided for detecting a target sequence in a sample of nucleic acids using a bifunctional hybridization probe according to this method.

  18. 18S rDNA Sequences from Microeukaryotes Reveal Oil Indicators in Mangrove Sediment

    PubMed Central

    Santos, Henrique F.; Cury, Juliano C.; Carmo, Flavia L.; Rosado, Alexandre S.; Peixoto, Raquel S.

    2010-01-01

    Background Microeukaryotes are an effective indicator of the presence of environmental contaminants. However, the characterisation of these organisms by conventional tools is often inefficient, and recent molecular studies have revealed a great diversity of microeukaryotes. The full extent of this diversity is unknown, and therefore, the distribution, ecological role and responses to anthropogenic effects of microeukaryotes are rather obscure. The majority of oil from oceanic oil spills (e.g., the May 2010 accident in the Gulf of Mexico) converges on coastal ecosystems such as mangroves, which are threatened with worldwide disappearance, highlighting the need for efficient tools to indicate the presence of oil in these environments. However, no studies have used molecular methods to assess the effects of oil contamination in mangrove sediment on microeukaryotes as a group. Methodology/Principal Findings We evaluated the population dynamics and the prevailing 18S rDNA phylotypes of microeukaryotes in mangrove sediment microcosms with and without oil contamination, using PCR/DGGE and clone libraries. We found that microeukaryotes are useful for monitoring oil contamination in mangroves. Our clone library analysis revealed a decrease in both diversity and species richness after contamination. The phylogenetic group that showed the greatest sensitivity to oil was the Nematoda. After contamination, a large increase in the abundance of the groups Bacillariophyta (diatoms) and Biosoecida was detected. The oil-contaminated samples were almost entirely dominated by organisms related to Bacillariophyta sp. and Cafeteria minima, which indicates that these groups are possible targets for biomonitoring oil in mangroves. The DGGE fingerprints also indicated shifts in microeukaryote profiles; specific band sequencing indicated the appearance of Bacillariophyta sp. only in contaminated samples and Nematoda only in non-contaminated sediment. Conclusions/Significance We believe that

  19. Bacteria obtained from a sequencing batch reactor that are capable of growth on dehydroabietic acid.

    PubMed Central

    Mohn, W W

    1995-01-01

    Eleven isolates capable of growth on the resin acid dehydroabietic acid (DhA) were obtained from a sequencing batch reactor designed to treat a high-strength process stream from a paper mill. The isolates belonged to two groups, represented by strains DhA-33 and DhA-35, which were characterized. In the bioreactor, bacteria like DhA-35 were more abundant than those like DhA-33. The population in the bioreactor of organisms capable of growth on DhA was estimated to be 1.1 x 10(6) propagules per ml, based on a most-probable-number determination. Analysis of small-subunit rRNA partial sequences indicated that DhA-33 was most closely related to Sphingomonas yanoikuyae (Sab = 0.875) and that DhA-35 was most closely related to Zoogloea ramigera (Sab = 0.849). Both isolates additionally grew on other abietanes, i.e., abietic and palustric acids, but not on the pimaranes, pimaric and isopimaric acids. For DhA-33 and DhA-35 with DhA as the sole organic substrate, doubling times were 2.7 and 2.2 h, respectively, and growth yields were 0.30 and 0.25 g of protein per g of DhA, respectively. Glucose as a cosubstrate stimulated growth of DhA-33 on DhA and stimulated DhA degradation by the culture. Pyruvate as a cosubstrate did not stimulate growth of DhA-35 on DhA and reduced the specific rate of DhA degradation of the culture. DhA induced DhA and abietic acid degradation activities in both strains, and these activities were heat labile. Cell suspensions of both strains consumed DhA at a rate of 6 mumol mg of protein-1 h-1.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7793937

  20. Sequence-defined bioactive macrocycles via an acid-catalysed cascade reaction

    NASA Astrophysics Data System (ADS)

    Porel, Mintu; Thornlow, Dana N.; Phan, Ngoc N.; Alabi, Christopher A.

    2016-06-01

    Synthetic macrocycles derived from sequence-defined oligomers are a unique structural class whose ring size, sequence and structure can be tuned via precise organization of the primary sequence. Similar to peptides and other peptidomimetics, these well-defined synthetic macromolecules become pharmacologically relevant when bioactive side chains are incorporated into their primary sequence. In this article, we report the synthesis of oligothioetheramide (oligoTEA) macrocycles via a one-pot acid-catalysed cascade reaction. The versatility of the cyclization chemistry and modularity of the assembly process was demonstrated via the synthesis of >20 diverse oligoTEA macrocycles. Structural characterization via NMR spectroscopy revealed the presence of conformational isomers, which enabled the determination of local chain dynamics within the macromolecular structure. Finally, we demonstrate the biological activity of oligoTEA macrocycles designed to mimic facially amphiphilic antimicrobial peptides. The preliminary results indicate that macrocyclic oligoTEAs with just two-to-three cationic charge centres can elicit potent antibacterial activity against Gram-positive and Gram-negative bacteria.

  1. Analysis of the functional domains of biosynthetic threonine deaminase by comparison of the amino acid sequences of three wild-type alleles to the amino acid sequence of biodegradative threonine deaminase.

    PubMed

    Taillon, B E; Little, R; Lawther, R P

    1988-03-31

    The nucleotide sequence of the gene, ilvA, for biosynthetic threonine deaminase (Tda) from Salmonella typhimurium was determined. The deduced amino acid sequence was compared with the deduced amino acid sequences of the biosynthetic Tda from Escherichia coli K-12 (ilvA) and Saccharomyces cerevisiae (ILV1) and the biodegradative Tda from E. coli K-12 (tdc). The comparison indicated the presence of two types of blocks of homologous amino acids. The first type of homology is in the N-terminal portion of all four isozymes of Tda and probably indicates amino acids involved in catalysis. The second type of homology is found in the C-terminal portion of the three biosynthetic isozymes and presumably is involved in either (i) the binding or interaction of the allosteric effector isoleucine with the enzyme, or (ii) subunit interactions. The sites of amino acid changes of two E. coli K-12 ilvA alleles with altered response to isoleucine are consistent with the conclusion that the C-terminal portion of biosynthetic Tda is involved in allosteric regulation. PMID:3290055

  2. Evolution of a Genome-Encoded Bias in Amino Acid Biosynthetic Pathways Is a Potential Indicator of Amino Acid Dynamics in the Environment

    PubMed Central

    Fasani, Rick A.; Savageau, Michael A.

    2014-01-01

    Overcoming the stress of starvation is one of an organism’s most challenging phenotypic responses. Those organisms that frequently survive the challenge, by virtue of their fitness, will have evolved genomes that are shaped by their specific environments. Understanding this genotype–environment–phenotype relationship at a deep level will require quantitative predictive models of the complex molecular systems that link these aspects of an organism’s existence. Here, we treat one of the most fundamental molecular systems, protein synthesis, and the amino acid biosynthetic pathways involved in the stringent response to starvation. These systems face an inherent logical dilemma: Building an amino acid biosynthetic pathway to synthesize its product—the cognate amino acid of the pathway—may require that very amino acid when it is no longer available. To study this potential “catch-22,” we have created a generic model of amino acid biosynthesis in response to sudden starvation. Our mathematical analysis and computational results indicate that there are two distinctly different outcomes: Partial recovery to a new steady state, or full system failure. Moreover, the cell’s fate is dictated by the cognate bias, the number of cognate amino acids in the corresponding biosynthetic pathway relative to the average number of that amino acid in the proteome. We test these implications by analyzing the proteomes of over 1,800 sequenced microbes, which reveals statistically significant evidence of low cognate bias, a genetic trait that would avoid the biosynthetic quandary. Furthermore, these results suggest that the pattern of cognate bias, which is readily derived by genome sequencing, may provide evolutionary clues to an organism’s natural environment. PMID:25118252

  3. Partial amino acid sequence of human factor D:homology with serine proteases.

    PubMed Central

    Volanakis, J E; Bhown, A; Bennett, J C; Mole, J E

    1980-01-01

    Human factor D purified to homogeneity by a modified procedure was subjected to NH2-terminal amino acid sequence analysis by using a modified automated Beckman sequencer. We identified 48 of the first 57 NH2-terminal amino acids in a single sequencer run, using microgram quantities of factor D. The deduced amino acid sequence represents approximately 25% of the primary structure of factor D. This extended NH2-terminal amino acid sequence of factor D was compared to that of other trypsin-related serine proteases. By visual inspection, strong homologies (33--50% identity) were observed with all the serine proteases included in the comparison. Interestingly, factor D showed a higher degree of homology to serine proteases of pancreatic origin than to those of serum origin. Images PMID:6987665

  4. Amino acid sequence of Japanese quail (Coturnix japonica) and northern bobwhite (Colinus virginianus) myoglobin.

    PubMed

    Goodson, John; Beckstead, Robert B; Payne, Jason; Singh, Rakesh K; Mohan, Anand

    2015-08-15

    Myoglobin has an important physiological role in vertebrates, and as the primary sarcoplasmic pigment in meat, influences quality perception and consumer acceptability. In this study, the amino acid sequences of Japanese quail and northern bobwhite myoglobin were deduced by cDNA cloning of the coding sequence from mRNA. Japanese quail myoglobin was isolated from quail cardiac muscles, purified using ammonium sulphate precipitation and gel-filtration, and subjected to multiple enzymatic digestions. Mass spectrometry corroborated the deduced protein amino acid sequence at the protein level. Sequence analysis revealed both species' myoglobin structures consist of 153 amino acids, differing at only three positions. When compared with chicken myoglobin, Japanese quail showed 98% sequence identity, and northern bobwhite 97% sequence identity. The myoglobin in both quail species contained eight histidine residues instead of the nine present in chicken and turkey. PMID:25794748

  5. RNA:DNA Ratio and Other Nucleic Acid Derived Indices in Marine Ecology

    PubMed Central

    Chícharo, Maria Alexandra; Chícharo, Luis

    2008-01-01

    Some of most used indicators in marine ecology are nucleic acid-derived indices. They can be divided by target levels in three groups: 1) at the organism level as ecophysiologic indicators, indicators such as RNA:DNA ratios, DNA:dry weight and RNA:protein, 2) at the population level, indicators such as growth rate, starvation incidence or fisheries impact indicators, and 3) at the community level, indicators such as trophic interactions, exergy indices and prey identification. The nucleic acids derived indices, especially RNA:DNA ratio, have been applied with success as indicators of nutritional condition, well been and growth in marine organisms. They are also useful as indicators of natural or anthropogenic impacts in marine population and communities, such as upwelling or dredge fisheries, respectively. They can help in understanding important issues of marine ecology such as trophic interactions in marine environment, fish and invertebrate recruitment failure and biodiversity changes, without laborious work of counting, measuring and identification of small marine organisms. Besides the objective of integrate nucleic acid derived indices across levels of organization, the paper will also include a general characterization of most used nucleic acid derived indices in marine ecology and also advantages and limitations of them. We can conclude that using indicators, such RNA:DNA ratios and other nucleic acids derived indices concomitantly with organism and ecosystems measures of responses to climate change (distribution, abundance, activity, metabolic rate, survival) will allow for the development of more rigorous and realistic predictions of the effects of anthropogenic climate change on marine systems. PMID:19325815

  6. Negative Ion In-Source Decay Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry for Sequencing Acidic Peptides

    NASA Astrophysics Data System (ADS)

    McMillen, Chelsea L.; Wright, Patience M.; Cassady, Carolyn J.

    2016-05-01

    Matrix-assisted laser desorption/ionization (MALDI) in-source decay was studied in the negative ion mode on deprotonated peptides to determine its usefulness for obtaining extensive sequence information for acidic peptides. Eight biological acidic peptides, ranging in size from 11 to 33 residues, were studied by negative ion mode ISD (nISD). The matrices 2,5-dihydroxybenzoic acid, 2-aminobenzoic acid, 2-aminobenzamide, 1,5-diaminonaphthalene, 5-amino-1-naphthol, 3-aminoquinoline, and 9-aminoacridine were used with each peptide. Optimal fragmentation was produced with 1,5-diaminonphthalene (DAN), and extensive sequence informative fragmentation was observed for every peptide except hirudin(54-65). Cleavage at the N-Cα bond of the peptide backbone, producing c' and z' ions, was dominant for all peptides. Cleavage of the N-Cα bond N-terminal to proline residues was not observed. The formation of c and z ions is also found in electron transfer dissociation (ETD), electron capture dissociation (ECD), and positive ion mode ISD, which are considered to be radical-driven techniques. Oxidized insulin chain A, which has four highly acidic oxidized cysteine residues, had less extensive fragmentation. This peptide also exhibited the only charged localized fragmentation, with more pronounced product ion formation adjacent to the highly acidic residues. In addition, spectra were obtained by positive ion mode ISD for each protonated peptide; more sequence informative fragmentation was observed via nISD for all peptides. Three of the peptides studied had no product ion formation in ISD, but extensive sequence informative fragmentation was found in their nISD spectra. The results of this study indicate that nISD can be used to readily obtain sequence information for acidic peptides.

  7. tax and rex Sequences of bovine leukaemia virus from globally diverse isolates: rex amino acid sequence more variable than tax.

    PubMed

    McGirr, K M; Buehring, G C

    2005-02-01

    Bovine leukaemia virus (BLV) is an important agricultural problem with high costs to the dairy industry. Here, we examine the variation of the tax and rex genes of BLV. The tax and rex genes share 420 bases and have overlapping reading frames. The tax gene encodes a protein that functions as a transactivator of the BLV promoter, is required for viral replication, acts on cellular promoters, and is responsible for oncogenesis. The rex facilitates the export of viral mRNAs from the nucleus and regulates transcription. We have sequenced five new isolates of the tax/rex gene. We examined the five new and three previously published tax/rex DNA and predicted amino acid sequences of BLV isolates from cattle in representative regions worldwide. The highest variation among nucleic acid sequences for tax and rex was 7% and 5%, respectively; among predicted amino acid sequences for Tax and Rex, 9% and 11%, respectively. Significantly more nucleotide changes resulted in predicted amino acid changes in the rex gene than in the tax gene (P < or = 0.0006). This variability is higher than previously reported for any region of the viral genome. This research may also have implications for the development of Tax-based vaccines. PMID:15702995

  8. The amino acid sequence of protein CM-3 from Dendroaspis polylepis polylepis (black mamba) venom.

    PubMed

    Joubert, F J

    1985-01-01

    Protein CM-3 from Dendroaspis polylepis polylepis venom was purified by gel filtration and ion exchange chromatography. It comprises 65 amino acids including eight half-cystines. The complete amino acid sequence of protein CM-3 has been elucidated. The sequence (residues 1-50) resembles that of the N-terminal sequence of the subunits of a synergistic type protein and residues 51-65 that of the C-terminal sequence of an angusticeps type protein. Mixtures of protein CM-3 and angusticeps type proteins showed no apparent synergistic effect, in that their toxicity in combination was no greater than the sum of their individual toxicities. PMID:4029488

  9. Chemical Reaction between Boric Acid and Phosphine Indicates Boric Acid as an Antidote for Aluminium Phosphide Poisoning

    PubMed Central

    Soltani, Motahareh; Shetab-Boushehri, Seyed F.; Shetab-Boushehri, Seyed V.

    2016-01-01

    Objectives: Aluminium phosphide (AlP) is a fumigant pesticide which protects stored grains from insects and rodents. When it comes into contact with moisture, AlP releases phosphine (PH3), a highly toxic gas. No efficient antidote has been found for AlP poisoning so far and most people who are poisoned do not survive. Boric acid is a Lewis acid with an empty p orbital which accepts electrons. This study aimed to investigate the neutralisation of PH3 gas with boric acid. Methods: This study was carried out at the Baharlou Hospital, Tehran University of Medical Sciences, Tehran, Iran, between December 2013 and February 2014. The volume of released gas, rate of gas evolution and changes in pH were measured during reactions of AlP tablets with water, acidified water, saturated boric acid solution, acidified saturated boric acid solution, activated charcoal and acidified activated charcoal. Infrared spectroscopy was used to study the resulting probable adduct between PH3 and boric acid. Results: Activated charcoal significantly reduced the volume of released gas (P <0.01). Although boric acid did not significantly reduce the volume of released gas, it significantly reduced the rate of gas evolution (P <0.01). A gaseous adduct was formed in the reaction between pure AlP and boric acid. Conclusion: These findings indicate that boric acid may be an efficient and non-toxic antidote for PH3 poisoning. PMID:27606109

  10. Indications for /sup 99m/technetium dimercapto-succinic acid scan in children

    SciTech Connect

    Gordon, I.

    1987-03-01

    The /sup 99m/technetium dimercapto-succinic acid scan provides an image of functional renal parenchyma. This static scan has specific indications and cannot be used simply in place of a /sup 99m/technetium diethylenetriaminepentaacetic acid scan. The major clinical indications for this investigation are the detection and/or evaluation of a renal scar, the small or absent kidney, an occult duplex system, certain renal masses, systemic hypertension or suspected vasculitis. The physiology of the /sup 99m/technetium dimercapto-succinic acid scan is reviewed briefly.

  11. Molecular Characterization of Buffalo Haptoglobin: Sequence Based Structural Comparison Indicates Convergent Evolution Between Ruminants and Human.

    PubMed

    Niranjan, S K; Goyal, S; Dubey, P K; Vohra, V; Singh, S; Kathiravan, P; Kataria, R S

    2016-01-01

    Haptoglobin (Hp) protein has high affinity for hemoglobin (Hb) binding during intravascular hemolysis and scavenges the hemoglobin induced free radicals. Earlier reports indicate about uniqueness of Hp molecule in human and cattle, but in other animals, it is not much studied. In this paper, we characterized buffalo Hp molecule and determined its molecular structure, evolutionary importance, and tissue expression. Comparative analysis and predicted domain structure indicated that the buffalo Hp has an internal duplicated region in α-chain only similar to an alternate Hp2 allele in human. This duplicated part encoded for an extra complement control protein CCP domain. Phylogenetic analysis revealed that buffalo and other ruminants were found to group together separated from all other non-ruminants, including human. The key amino acid residues involved in Hp and Hb as well as Hp and macrophage scavenger receptor, CD163 interactions in buffalo, depicted a significant variation in comparison to other non-ruminant species. Constitutive expression of Hp was also confirmed across all the vital tissues of buffalo, for the first time. Results revealed that buffalo Hp is both structurally and functionally conserved, having internal duplication in α-chain similar to human Hp2 and other ruminant species, which might have evolved separately as a convergent evolutionary process. Furthermore, the presence of extra Hp CCP domain possibly in all ruminants may have an effect during dimerization of molecule in these species. PMID:26646629

  12. Stress degradation studies and stability-indicating TLC-densitometric method of glycyrrhetic acid

    PubMed Central

    2013-01-01

    Background Glycyrrhetic acid, a pentacyclic triterpenoid, possesses a broad range of pharmacological activities and serves as template to synthesize many bioactive drugs. This paper describes a simple, accurate, and sensitive stability-indicating TLC densitometric method for the determination of glycyrrhetic acid and its degradation product as per the ICH guidelines. Results Separation was carried out on TLC aluminium sheet pre-coated with silica gel 60F254 using chloroform, methanol and formic acid (9:0.9:0.1, v/v). Compact spot for glycyrrhetic acid was found at Rf value of 0.42 ± 0.03. Densitometric analysis was carried out in the absorbance mode at λmax 254 nm. Glycyrrhetic acid was found to be stable to the exposure of base, neutral, oxidation, dry heating treatment and wet heating treatment, but showed degradation under acidic and photochemical conditions. Moreover, fragmentation pattern of glycyrrhetic acid was developed by using a positive ion electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-QqTOF-MS/MS) hybrid instrument. A photo-degraded product was characterized through comparison of mass spectrometric studies with glycyrrhetic acid. Conclusion The developed stability-indicating TLC-densitometric method can be applied for routine analysis of glycyrrhetic acid in the presence of its degradation products. PMID:23327365

  13. The Chinese hamster Alu-equivalent sequence: a conserved highly repetitious, interspersed deoxyribonucleic acid sequence in mammals has a structure suggestive of a transposable element.

    PubMed Central

    Haynes, S R; Toomey, T P; Leinwand, L; Jelinek, W R

    1981-01-01

    A consensus sequence has been determined for a major interspersed deoxyribonucleic acid repeat in the genome of Chinese hamster ovary cells (CHO cells). This sequence is extensively homologous to (i) the human Alu sequence (P. L. Deininger et al., J. Mol. Biol., in press), (ii) the mouse B1 interspersed repetitious sequence (Krayev et al., Nucleic Acids Res. 8:1201-1215, 1980) (iii) an interspersed repetitious sequence from African green monkey deoxyribonucleic acid (Dhruva et al., Proc. Natl. Acad. Sci. U.S.A. 77:4514-4518, 1980) and (iv) the CHO and mouse 4.5S ribonucleic acid (this report; F. Harada and N. Kato, Nucleic Acids Res. 8:1273-1285, 1980). Because the CHO consensus sequence shows significant homology to the human Alu sequence it is termed the CHO Alu-equivalent sequence. A conserved structure surrounding CHO Alu-equivalent family members can be recognized. It is similar to that surrounding the human Alu and the mouse B1 sequences, and is represented as follows: direct repeat-CHO-Alu-A-rich sequence-direct repeat. A composite interspersed repetitious sequence has been identified. Its structure is represented as follows: direct repeat-residue 47 to 107 of CHO-Alu-non-Alu repetitious sequence-A-rich sequence-direct repeat. Because the Alu flanking sequences resemble those that flank known transposable elements, we think it likely that the Alu sequence dispersed throughout the mammalian genome by transposition. Images PMID:9279371

  14. Computer Simulation of the Determination of Amino Acid Sequences in Polypeptides

    ERIC Educational Resources Information Center

    Daubert, Stephen D.; Sontum, Stephen F.

    1977-01-01

    Describes a computer program that generates a random string of amino acids and guides the student in determining the correct sequence of a given protein by using experimental analytic data for that protein. (MLH)

  15. Accuracy of sequence alignment and fold assessment using reduced amino acid alphabets.

    PubMed

    Melo, Francisco; Marti-Renom, Marc A

    2006-06-01

    Reduced or simplified amino acid alphabets group the 20 naturally occurring amino acids into a smaller number of representative protein residues. To date, several reduced amino acid alphabets have been proposed, which have been derived and optimized by a variety of methods. The resulting reduced amino acid alphabets have been applied to pattern recognition, generation of consensus sequences from multiple alignments, protein folding, and protein structure prediction. In this work, amino acid substitution matrices and statistical potentials were derived based on several reduced amino acid alphabets and their performance assessed in a large benchmark for the tasks of sequence alignment and fold assessment of protein structure models, using as a reference frame the standard alphabet of 20 amino acids. The results showed that a large reduction in the total number of residue types does not necessarily translate into a significant loss of discriminative power for sequence alignment and fold assessment. Therefore, some definitions of a few residue types are able to encode most of the relevant sequence/structure information that is present in the 20 standard amino acids. Based on these results, we suggest that the use of reduced amino acid alphabets may allow to increasing the accuracy of current substitution matrices and statistical potentials for the prediction of protein structure of remote homologs. PMID:16506243

  16. Characterization of mouse cellular deoxyribonucleic acid homologous to Abelson murine leukemia virus-specific sequences.

    PubMed Central

    Dale, B; Ozanne, B

    1981-01-01

    The genome of Abelson murine leukemia virus (A-MuLV) consists of sequences derived from both BALB/c mouse deoxyribonucleic acid and the genome of Moloney murine leukemia virus. Using deoxyribonucleic acid linear intermediates as a source of retroviral deoxyribonucleic acid, we isolated a recombinant plasmid which contained 1.9 kilobases of the 3.5-kilobase mouse-derived sequences found in A-MuLV (A-MuLV-specific sequences). We used this clone, designated pSA-17, as a probe restriction enzyme and Southern blot analyses to examine the arrangement of homologous sequences in BALB/c deoxyribonucleic acid (endogenous Abelson sequences). The endogenous Abelson sequences within the mouse genome were interrupted by noncoding regions, suggesting that a rearrangement of the cell sequences was required to produce the sequence found in the virus. Endogenous Abelson sequences were arranged similarly in mice that were susceptible to A-MuLV tumors and in mice that were resistant to A-MuLV tumors. An examination of three BALB/c plasmacytomas and a BALB/c early B-cell tumor likewise revealed no alteration in the arrangement of the endogenous Abelson sequences. Homology to pSA-17 was also observed in deoxyribonucleic acids prepared from rat, hamster, chicken, and human cells. An isolate of A-MuLV which encoded a 160,000-dalton transforming protein (P160) contained 700 more base pairs of mouse sequences than the standard A-MuLV isolate, which encoded a 120,000-dalton transforming protein (P120). Images PMID:9279386

  17. The amino acid sequence of monal pheasant lysozyme and its activity.

    PubMed

    Araki, T; Matsumoto, T; Torikata, T

    1998-10-01

    The amino acid sequence of monal pheasant lysozyme and its activity were analyzed. Carboxymethylated lysozyme was digested with trypsin and the resulting peptides were sequenced. The established amino acid sequence had one amino acid substitution at position 102 (Arg to Gly) comparing with Indian peafowl lysozyme and four amino acid substitutions at positions 3 (Phe to Tyr), 15 (His to Leu), 41 (Gln to His), and 121 (Gln to His) with chicken lysozyme. Analysis of the time-courses of reaction using N-acetylglucosamine pentamer as a substrate showed a difference of binding free energy change (-0.4 kcal/mol) at subsites A between monal pheasant and Indian peafowl lysozyme. This was assumed to be caused by the amino acid substitution at subsite A with loss of a positive charge at position 102 (Arg102 to Gly). PMID:9836434

  18. Studies on monotreme proteins. VII. Amino acid sequence of myoglobin from the platypus, Ornithoryhynchus anatinus.

    PubMed

    Fisher, W K; Thompson, E O

    1976-03-01

    Myoglobin isolated from skeletal muscle of the platypus contains 153 amino acid residues. The complete amino acid sequence has been determined following cleavage with cyanogen bromide and further digestion of the four fragments with trypsin, chymotrypsin, pepsin and thermolysin. Sequences of the purified peptides were determined by the dansyl-Edman procedure. The amino acid sequence showed 25 differences from human myoglobin and 24 from kangaroo myoglobin. Amino acid sequences in myoglobins are more conserved than sequences in the alpha- and beta-globin chains, and platypus myoglobin shows a similar number of variations in sequence to kangaroo myoglobin when compared with myoglobin of other species. The date of divergence of the platypus from other mammals was estimated at 102 +/- 31 million years, based on the number of amino acid differences between species and allowing for mutations during the evolutionary period. This estimate differs widely from the estimate given by similar treatment of the alpha- and beta-chain sequences and a constant rate of mutation of globin chains is not supported. PMID:962722

  19. cDNA-derived amino acid sequences of myoglobins from nine species of whales and dolphins.

    PubMed

    Iwanami, Kentaro; Mita, Hajime; Yamamoto, Yasuhiko; Fujise, Yoshihiro; Yamada, Tadasu; Suzuki, Tomohiko

    2006-10-01

    We determined the myoglobin (Mb) cDNA sequences of nine cetaceans, of which six are the first reports of Mb sequences: sei whale (Balaenoptera borealis), Bryde's whale (Balaenoptera edeni), pygmy sperm whale (Kogia breviceps), Stejneger's beaked whale (Mesoplodon stejnegeri), Longman's beaked whale (Indopacetus pacificus), and melon-headed whale (Peponocephala electra), and three confirm the previously determined chemical amino acid sequences: sperm whale (Physeter macrocephalus), common minke whale (Balaenoptera acutorostrata) and pantropical spotted dolphin (Stenella attenuata). We found two types of Mb in the skeletal muscle of pantropical spotted dolphin: Mb I with the same amino acid sequence as that deposited in the protein database, and Mb II, which differs at two amino acid residues compared with Mb I. Using an alignment of the amino acid or cDNA sequences of cetacean Mb, we constructed a phylogenetic tree by the NJ method. Clustering of cetacean Mb amino acid and cDNA sequences essentially follows the classical taxonomy of cetaceans, suggesting that Mb sequence data is valid for classification of cetaceans at least to the family level. PMID:16962803

  20. Analysis of essential oils and fatty acids from Platycodi Radix using chemometric methods and retention indices.

    PubMed

    He, Min; Li, Yaping; Yan, Jun; Cao, Dongsheng; Liang, Yizeng

    2013-04-01

    The chemical composition of the essential oils and fatty acids among nine groups of Platycodi Radix in China was analyzed by gas chromatography-mass spectrometry. Complicated components were resolved using chemometric methods. Simultaneously, the various features among heuristic evolving latent projections, selective ion analysis and alternative moving window factor analysis were compared by using some experimental data. Temperature-programmed retention indices were applied in further identification of the chemical composition of the essential oils. The equivalent chain length, fraction chain length, and an established special retention indices library integrated with mass spectrometry were also applied to further identify the composition of fatty acids, including total fatty acids, esterified fatty acids and free fatty acids. A total of 121 different compounds accounting for 95.12-98.74% were identified among the essential oils. Chemical polymorphisms and variation existed in the essential oils of Platycodi Radix. Sixteen components were identified in fatty acids, and linoleic acid (18:2n-6c) and other unsaturated acid possess a characteristic majority. PMID:22964951

  1. Amino acid sequence of rabbit kidney neutral endopeptidase 24.11 (enkephalinase) deduced from a complementary DNA.

    PubMed Central

    Devault, A; Lazure, C; Nault, C; Le Moual, H; Seidah, N G; Chrétien, M; Kahn, P; Powell, J; Mallet, J; Beaumont, A

    1987-01-01

    Neutral endopeptidase (EC 3.4.24.11) is a major constituent of kidney brush border membranes. It is also present in the brain where it has been shown to be involved in the inactivation of opioid peptides, methionine- and leucine-enkephalins. For this reason this enzyme is often called 'enkephalinase'. In order to characterize the primary structure of the enzyme, oligonucleotide probes were designed from partial amino acid sequences and used to isolate clones from kidney cDNA libraries. Sequencing of the cDNA inserts revealed the complete primary structure of the enzyme. Neutral endopeptidase consists of 750 amino acids. It contains a short N-terminal cytoplasmic domain (27 amino acids), a single membrane-spanning segment (23 amino acids) and an extracellular domain that comprises most of the protein mass. The comparison of the primary structure of neutral endopeptidase with that of thermolysin, a bacterial Zn-metallopeptidase, indicates that most of the amino acid residues involved in Zn coordination and catalytic activity in thermolysin are found within highly honmologous sequences in neutral endopeptidase. Images Fig. 1. Fig. 3. PMID:2440677

  2. Draft Genome Sequences of Two Novel Acidimicrobiaceae Members from an Acid Mine Drainage Biofilm Metagenome.

    PubMed

    Pinto, Ameet J; Sharp, Jonathan O; Yoder, Michael J; Almstrand, Robert

    2016-01-01

    Bacteria belonging to the family Acidimicrobiaceae are frequently encountered in heavy metal-contaminated acidic environments. However, their phylogenetic and metabolic diversity is poorly resolved. We present draft genome sequences of two novel and phylogenetically distinct Acidimicrobiaceae members assembled from an acid mine drainage biofilm metagenome. PMID:26769942

  3. Draft Genome Sequences of Two Novel Acidimicrobiaceae Members from an Acid Mine Drainage Biofilm Metagenome

    PubMed Central

    Pinto, Ameet J.; Sharp, Jonathan O.; Yoder, Michael J.

    2016-01-01

    Bacteria belonging to the family Acidimicrobiaceae are frequently encountered in heavy metal-contaminated acidic environments. However, their phylogenetic and metabolic diversity is poorly resolved. We present draft genome sequences of two novel and phylogenetically distinct Acidimicrobiaceae members assembled from an acid mine drainage biofilm metagenome. PMID:26769942

  4. Event-related potential indices of congruency sequence effects without feature integration or contingency learning confounds.

    PubMed

    Larson, Michael J; Clayson, Peter E; Kirwan, C Brock; Weissman, Daniel H

    2016-06-01

    The congruency effect in Stroop-like tasks (i.e., increased response time and reduced accuracy in incongruent relative to congruent trials) is often smaller when the previous trial was incongruent as compared to congruent. This congruency sequence effect (CSE) is thought to reflect cognitive control processes that shift attention to the target and/or modulate the response engendered by the distracter differently after incongruent relative to congruent trials. The neural signatures of CSEs are therefore usually attributed to cognitive control processes that minimize distraction from irrelevant stimuli. However, CSEs in previous functional neuroimaging studies were ubiquitously confounded with feature integration and/or contingency learning processes. We therefore investigated whether a neural CSE can be observed without such confounds in a group of healthy young adults (n = 56). To this end, we combined a prime-probe task that lacks such confounds with high-density ERPs to identify, for the first time, the neural time course of confound-minimized CSEs. Replicating recent behavioral findings, we observed strong CSEs in this task for mean response time and mean accuracy. Critically, conceptually replicating prior ERP results from confounded tasks, we also observed a CSE in both the parietal conflict slow potential (conflict SP) and the frontomedial N450. These findings indicate for the first time that neural CSEs as indexed by ERPs can be observed without the typical confounds. More broadly, the present study provides a confound-minimized protocol that will help future researchers to better isolate the neural bases of control processes that minimize distraction from irrelevant stimuli. PMID:26854028

  5. Two distinct ferredoxins from Rhodobacter capsulatus: complete amino acid sequences and molecular evolution.

    PubMed

    Saeki, K; Suetsugu, Y; Yao, Y; Horio, T; Marrs, B L; Matsubara, H

    1990-09-01

    Two distinct ferredoxins were purified from Rhodobacter capsulatus SB1003. Their complete amino acid sequences were determined by a combination of protease digestion, BrCN cleavage and Edman degradation. Ferredoxins I and II were composed of 64 and 111 amino acids, respectively, with molecular weights of 6,728 and 12,549 excluding iron and sulfur atoms. Both contained two Cys clusters in their amino acid sequences. The first cluster of ferredoxin I and the second cluster of ferredoxin II had a sequence, CxxCxxCxxxCP, in common with the ferredoxins found in Clostridia. The second cluster of ferredoxin I had a sequence, CxxCxxxxxxxxCxxxCM, with extra amino acids between the second and third Cys, which has been reported for other photosynthetic bacterial ferredoxins and putative ferredoxins (nif-gene products) from nitrogen-fixing bacteria, and with a unique occurrence of Met. The first cluster of ferredoxin II had a CxxCxxxxCxxxCP sequence, with two additional amino acids between the second and third Cys, a characteristics feature of Azotobacter-[3Fe-4S] [4Fe-4S]-ferredoxin. Ferredoxin II was also similar to Azotobacter-type ferredoxins with an extended carboxyl (C-) terminal sequence compared to the common Clostridium-type. The evolutionary relationship of the two together with a putative one recently found to be encoded in nifENXQ region in this bacterium [Moreno-Vivian et al. (1989) J. Bacteriol. 171, 2591-2598] is discussed. PMID:2277040

  6. Amino Acid Sequence of Anionic Peroxidase from the Windmill Palm Tree Trachycarpus fortunei

    PubMed Central

    2015-01-01

    Palm peroxidases are extremely stable and have uncommon substrate specificity. This study was designed to fill in the knowledge gap about the structures of a peroxidase from the windmill palm tree Trachycarpus fortunei. The complete amino acid sequence and partial glycosylation were determined by MALDI-top-down sequencing of native windmill palm tree peroxidase (WPTP), MALDI-TOF/TOF MS/MS of WPTP tryptic peptides, and cDNA sequencing. The propeptide of WPTP contained N- and C-terminal signal sequences which contained 21 and 17 amino acid residues, respectively. Mature WPTP was 306 amino acids in length, and its carbohydrate content ranged from 21% to 29%. Comparison to closely related royal palm tree peroxidase revealed structural features that may explain differences in their substrate specificity. The results can be used to guide engineering of WPTP and its novel applications. PMID:25383699

  7. Comparisons of Reproducibility and Mean Values of Diffusion Tensor Imaging-Derived Indices between Unipolar and Bipolar Diffusion Pulse Sequences.

    PubMed

    Miao, Hsiao-Chien; Wu, Ming-Ting; Kao, E-Fong; Chiu, Yu-Hsien; Chou, Ming-Chung

    2015-01-01

    Eddy current distortion is an important issue that may influence the quantitative measurements of diffusion tensor imaging (DTI). The corrections of eddy current artifacts could be performed using bipolar diffusion gradients or unipolar gradients with affine registration. Whether the diffusion pulse sequence affects the quantification of DTI indices and the technique that produces more reliable DTI indices in terms of reproducibility both remain unclear. Therefore, the purpose of this study was to compare the reproducibility and mean values of DTI-derived indices between unipolar and bipolar diffusion pulse sequences based on actual human brain data. Five repeated datasets of unipolar and bipolar DTI were acquired from 10 healthy subjects at different echo times (TEs). The reproducibility and mean values of DTI indices were assessed by calculating the coefficient of variation and mean values of the 5 repeated measurements. The results revealed that the reproducibility and mean values of DTI indices were significantly affected by the pulse sequence. Unipolar DTI exhibited significantly higher reproducibility than bipolar DTI even at the same TE, and the mean values of DTI indices were significantly different between them. Therefore, we concluded that the reproducibility and mean values of DTI indices were significantly influenced by diffusion pulse sequences. PMID:25753738

  8. DNA sequences of Alu elements indicate a recent replacement of the human autosomal genetic complement

    SciTech Connect

    Knight, A.; Deininger, P.L.; Batzer, M.A.

    1996-04-30

    DNA sequences of neutral nuclear autosomal loci, compared across diverse human populations, provide a previously untapped perspective into the mode and tempo of the emergence of modern humans and a critical comparison with published clonally inherited mitochondrial DNA and Y chromosome measurements of human diversity. We obtained over 55 kilobases of sequence from three autosomal loci encompassing Alu repeats for representatives of diverse human populations as well as orthologous sequences for other hominoid species at one of these loci. Nucleotide diversity was exceedingly low. Most individuals and populations were identical. Only a single nucleotide difference distinguished presumed ancestral alleles from descendants. These results differ from those expected if alleles from divergent archaic populations were maintained through multiregional continuity. The observed virtual lack of sequence polymorphism is the signature of a recent single origin for modern humans, with general replacement of archaic populations. 47 refs., 2 figs., 1 tab.

  9. DNA sequences of Alu elements indicate a recent replacement of the human autosomal genetic complement.

    PubMed Central

    Knight, A; Batzer, M A; Stoneking, M; Tiwari, H K; Scheer, W D; Herrera, R J; Deininger, P L

    1996-01-01

    DNA sequences of neutral nuclear autosomal loci, compared across diverse human populations, provide a previously untapped perspective into the mode and tempo of the emergence of modern humans and a critical comparison with published clonally inherited mitochondrial DNA and Y chromosome measurements of human diversity. We obtained over 55 kilobases of sequence from three autosomal loci encompassing Alu repeats for representatives of diverse human populations as well as orthologous sequences for other hominoid species at one of these loci. Nucleotide diversity was exceedingly low. Most individuals and populations were identical. Only a single nucleotide difference distinguished presumed ancestral alleles from descendants. These results differ from those expected if alleles from divergent archaic populations were maintained through multiregional continuity. The observed virtual lack of sequence polymorphism is the signature of a recent single origin for modern humans, with general replacement of archaic populations. PMID:8633071

  10. Protein chemotaxonomy. XIII. Amino acid sequence of ferredoxin from Panax ginseng.

    PubMed

    Mino, Yoshiki

    2006-08-01

    The complete amino acid sequence of [2Fe-2S] ferredoxin from Panax ginseng (Araliaceae) has been determined by automated Edman degradation of the entire S-carboxymethylcysteinyl protein and of the peptides obtained by enzymatic digestion. This ferredoxin has a unique amino acid sequence, which includes an insertion of Tyr at the 3rd position from the amino-terminus and a deletion of two amino acid residues at the carboxyl terminus. This ferredoxin had 18 differences in its amino acid sequence compared to that of Petroselinum sativum (Umbelliferae). In contrast, 23-33 differences were observed compared to other dicotyledonous plants. This suggests that Panax ginseng is related taxonomically to umbelliferous plants. PMID:16880642

  11. Complete amino acid sequence and structure characterization of the taste-modifying protein, miraculin.

    PubMed

    Theerasilp, S; Hitotsuya, H; Nakajo, S; Nakaya, K; Nakamura, Y; Kurihara, Y

    1989-04-25

    The taste-modifying protein, miraculin, has the unusual property of modifying sour taste into sweet taste. The complete amino acid sequence of miraculin purified from miracle fruits by a newly developed method (Theerasilp, S., and Kurihara, Y. (1988) J. Biol. Chem. 263, 11536-11539) was determined by an automatic Edman degradation method. Miraculin was a single polypeptide with 191 amino acid residues. The calculated molecular weight based on the amino acid sequence and the carbohydrate content (13.9%) was 24,600. Asn-42 and Asn-186 were linked N-glycosidically to carbohydrate chains. High homology was found between the amino acid sequences of miraculin and soybean trypsin inhibitor. PMID:2708331

  12. Purification and partial amino acid sequence of the chloroplast cytochrome b-559.

    PubMed

    Widger, W R; Cramer, W A; Hermodson, M; Meyer, D; Gullifor, M

    1984-03-25

    The hydrophobic cytochrome b-559, purified from unstacked, ethanol-washed spinach thylakoid membranes, using extraction with 2% Triton X-100 in 4 M urea and three chromatographic steps in the presence of protease inhibitors, has a dominant band on sodium dodecyl sulfate-urea gels corresponding to Mr = 10,000. The yield of this preparation is 30-50% (5-10 mg) starting with 600 mg of chlorophyll. The heme content yields a calculated molecular weight of no more than 17,500/heme, and perhaps somewhat smaller after correction for impurities. The Mr = 10,000 band is stained by the tetramethylbenzidine-H2O2 heme reagent on lithium dodecyl sulfate gels run at 0 degrees C. The Mr = 10,000 protein, further separated by high performance liquid chromatography, contains a unique NH2 terminus that is not blocked, and the amino acid sequence for the first 27 residues is NH2-Ser-Gly-Ser-Thr-Gly-Glu-Arg-Ser-Phe-Ala-Asp-Ile-Ile-Thr-Ser-Ile-Arg-Tyr-Trp -Val-Ile-X-Ser-Ile-Thr-Ile-Pro. . . COOH. Approximately 55% of the amino acids are hydrophobic, based on amino acid analysis of the Mr = 10,000 peptide, which also indicated the presence of at least one histidine. Only one cytochrome b-559 component could be identified, whose yield indicated that it arises from a single b-559 protein in chloroplasts corresponding to the in situ high potential cytochrome of the chloroplast photosystem II. PMID:6706983

  13. Complete cDNA and derived amino acid sequence of human factor V

    SciTech Connect

    Jenny, R.J.; Pittman, D.D.; Toole, J.J.; Kriz, R.W.; Aldape, R.A.; Hewick, R.M.; Kaufman, R.J.; Mann, K.G.

    1987-07-01

    cDNA clones encoding human factor V have been isolated from an oligo(dT)-primed human fetal liver cDNA library prepared with vector Charon 21A. The cDNA sequence of factor V from three overlapping clones includes a 6672-base-pair (bp) coding region, a 90-bp 5' untranslated region, and a 163-bp 3' untranslated region within which is a poly(A)tail. The deduced amino acid sequence consists of 2224 amino acids inclusive of a 28-amino acid leader peptide. Direct comparison with human factor VIII reveals considerable homology between proteins in amino acid sequence and domain structure: a triplicated A domain and duplicated C domain show approx. 40% identity with the corresponding domains in factor VIII. As in factor VIII, the A domains of factor V share approx. 40% amino acid-sequence homology with the three highly conserved domains in ceruloplasmin. The B domain of factor V contains 35 tandem and approx. 9 additional semiconserved repeats of nine amino acids of the form Asp-Leu-Ser-Gln-Thr-Thr/Asn-Leu-Ser-Pro and 2 additional semiconserved repeats of 17 amino acids. Factor V contains 37 potential N-linked glycosylation sites, 25 of which are in the B domain, and a total of 19 cysteine residues.

  14. Urinary guanidinoacetic acid excretion as an indicator of gentamicin nephrotoxicity in rats.

    PubMed

    Kiyatake, Ikuo; Nakamura, Tsukasa; Koide, Hikaru

    2004-07-01

    The kidney is the main site of guanidinoacetic acid synthesis and excretion. The aim of this study was to examine whether urinary guanidinoacetic acid is a sensitive indicator for diagnosis of early-stage gentamicin nephrotoxicity. Early-stage renal injury was induced in rats by a single intravenous injection of 5, 10, or 30 mg/kg body weight gentamicin. Twenty-four hours after injection all rats were killed. Blood, urine and tissue guanidino compound concentrations were determined by high performance liquid chromatography. Glycine amidinotransferase activity in tissues was assayed according to the method of Pilsum. Urinary guanidinoacetic acid excretion was decreased in 5 mg/kg gentamicin-treated rats in comparison to that in control rats, whereas urine N-acetyl-beta-D-glucosaminidase activity and beta2-microglobulin were unchanged. Guanidinoacetic acid concentration and glycine amidinotransferase activity in the kidney were significantly decreased in 5, 10, and 30 mg/kg gentamicin-treated rats; the decreases were dose-dependent. These results suggest that the urine guanidinoacetic acid concentration is a more sensitive indicator of renal injury than conventional indicators such as urine N-acetyl-beta-D-glucosaminidase and beta2-microglobulin. PMID:15462098

  15. Aqueous extracts of Mozambican plants as alternative and environmentally safe acid-base indicators.

    PubMed

    Macuvele, Domingos Lusitaneo Pier; Sithole, Gerre Zebedias Samo; Cesca, Karina; Macuvele, Suzana Lília Pinare; Matsinhe, Jonas Valente

    2016-06-01

    Indicators are substances that change color as the pH of the medium. Many of these substances are dyes of synthetic origin. The mulala plant (Euclea natalensis), which roots are commonly used by rural communities for their oral hygiene, and roseira (Hibiscus rosa-sinensis), an ornamental plant, are abundant in Mozambique. Currently, synthetic acid-base indicators are most commonly used but have environmental implications and, on the other hand, are expensive products, so the demand for natural indicators started. This study investigated the applicability of aqueous extracts of H. rosa-sinensis and E. natalensis as acid-base indicators. Ground on this work, the extracts can be used as acid-base indicators. On the basis of the absorption spectroscopy in both the UV-Vis region and previous studies, it was possible to preliminarily pinpoint anthocyanins and naphthoquinones as responsible for the shifting of colors depending on the pH range of aqueous extracts of H. rosa-sinensis and E. natalensis. These natural indicators are easily accessible, inexpensive, easy to extract, environmentally safe, and locally available. PMID:26936478

  16. Purification, characterization, and amino acid sequencing of a. delta. /sup 5/-3-oxosteroid isomerase from Pseudomonas putida biotype B

    SciTech Connect

    Linden, K.G.

    1986-01-01

    Studies were performed on the ..delta../sup 5/-3-oxosteroid isomerase from Pseudomonas putida biotype B. The studies have involved three broad areas: improvement in the purification of the enzyme, further characterization of the purified enzyme, and completion of the amino acid sequence of the enzyme. For the purification of the enzyme, techniques for removing the isomerase from whole cells were studied, the effects of ionic strength on the binding of the isomerase to steroidal affinity resins was explored, and a new affinity resin was developed. Absorption spectra and the proton NMR spectra of the isomerase were obtained. Amino acid sequencing of the oxosteroid isomerase indicates that the enzyme is a dimeric protein consisting of two identical subunits each consisting of a polypeptide chain of 131 residues and a M/sub r/ = 14,536.

  17. Using Acid Number as a Leading Indicator of Refrigeration and Air Conditioning System Performance

    SciTech Connect

    Dennis Cartlidge; Hans Schellhase

    2003-07-31

    number (TAN), which includes both mineral acids and organic acids, is therefore a useful indicator which can be used to monitor the condition of the system in order to perform remedial maintenance, when required, to prevent system failure. The critical TAN value is the acid level at which remedial action should be taken to prevent the onset of rapid acid formation which can result in system failure. The level of 0.05 mg KOH/g of oil was established for CFC/mineral oil systems based on analysis of 700 used lubricants from operating systems and failed units. There is no consensus within the refrigeration industry as to the critical TAN value for HFC/POE systems, however, the value will be higher than the CFC/mineral oil systems critical TAN value because of the much weaker organic acids produced from POE. A similar study of used POE lubricants should be performed to establish a critical TAN limit for POE systems. Titrimetric analysis per ASTM procedures is the most commonly used method to determine TAN values in lubricants in the refrigeration industry and other industries dealing with lubricating oils. For field measurements, acid test kits are often used since they provide rapid, semi-quantitative TAN results.

  18. 76 FR 11891 - Temperature-Indicating Devices; Thermally Processed Low-Acid Foods Packaged in Hermetically...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-03

    ... March 3, 2011 Part III Department of Health and Human Services Food and Drug Administration 21 CFR Part 113 Temperature-Indicating Devices; Thermally Processed Low-Acid Foods Packaged in Hermetically Sealed... Regulations#0;#0; ] DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration 21 CFR Part...

  19. Analysis of Citric Acid in Beverages: Use of an Indicator Displacement Assay

    ERIC Educational Resources Information Center

    Umali, Alona P.; Anslyn, Eric V.; Wright, Aaron T.; Blieden, Clifford R.; Smith, Carolyne K.; Tian, Tian; Truong, Jennifer A.; Crumm, Caitlin E.; Garcia, Jorge E.; Lee, Soal; Mosier, Meredith; Nguyen, Chester P.

    2010-01-01

    The use of an indicator displacement assay permits the visualization of binding events between host and guest molecules. An undergraduate laboratory experiment is described to demonstrate the technique in the determination of citric acid content in commercially available beverages such as soda pop and fruit juices. Through the technique, students…

  20. USE OF FATTY ACID STABLE CARBON ISOTOPE RATIO TO INDICATE MICROBIAL CARBON SOURCE IN TROPICAL SOILS

    EPA Science Inventory


    We use measurements of the concentration and stable carbon isotope ratio of individual microbial phospholipid fatty acids (PLFAs) in soils as indicators of live microbial biomass levels, broad microbial community structure, and microbial carbon source. For studies of soil o...

  1. ACID-FAST BACTERIA AND YEASTS AS INDICATORS OF DISINFECTION EFFICIENCY

    EPA Science Inventory

    Since the coliform group of organisms is considered to be less resistant to chlorine than some bacterial and viral pathogens, the utility of both yeast and acid-fast oganisms as potntial indicators of disinfection efficiency was evaluated. In most laboratory studies these two gro...

  2. Analysis and Identification of Acid-Base Indicator Dyes by Thin-Layer Chromatography

    ERIC Educational Resources Information Center

    Clark, Daniel D.

    2007-01-01

    Thin-layer chromatography (TLC) is a very simple and effective technique that is used by chemists by different purposes, including the monitoring of the progress of a reaction. TLC can also be easily used for the analysis and identification of various acid-base indicator dyes.

  3. Urinary excretion of mevalonic acid as an indicator of cholesterol synthesis.

    PubMed

    Lindenthal, B; Simatupang, A; Dotti, M T; Federico, A; Lütjohann, D; von Bergmann, K

    1996-10-01

    Urinary excretion of mevalonic acid was investigated as an indicator of cholesterol synthesis. In normolipemic volunteers, excretion of mevalonic acid averaged 3.51 +/- 0.59 (SD) micrograms/kg x day1; (n = 24) and was not different from patients with hypercholesterolemia (3.30 +/- 0.92 micrograms/kg x day1; n = 24). In patients with cerebrotendineous xanthomatosis, the excretion was significantly higher (8.55 +/- 1.92 micrograms/kg x day1; n = 6, P < 0.001) but comparable to volunteers treated with cholestyramine (6.69 +/- 2.6 micrograms/kg x day1; n = 5). A significant correlation was found between 24-h excretion of mevalonic acid and cholesterol synthesis (r = 0.835; n = 35; P < 0.001). The coefficient of variation of excretion of mevalonic acid during 3 consecutive days was small (9.8%; n = 7). However, urinary output of mevalonic acid was significantly higher during the night (164 +/- 14 micrograms/12-h) than during the day (129 +/- 9 micrograms/12-h; n = 11; P < 0.05). In patients treated with simvastatin (40 mg/day) for 6 weeks, the ratio of mevalonic acid to creatinine in a morning urine sample decreased significantly compared to pretreatment values (110 +/- 25 micrograms/g vs. 66 +/- 25 micrograms/g; P < 0.001). Furthermore, the ratio of mevalonic acid to creatinine in a morning urine sample correlated with the ratio from the 24-h collection period (r = 0.714; n = 34; P < 0.001). The results indicate that the analysis of urinary mevalonic acid, either in 24-h collection or in a single morning sample, is an attractive method for evaluation of long and very short term changes of the rates of cholesterol synthesis. PMID:8906596

  4. Detection and isolation of nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    1997-01-01

    A method for detecting a target nucleic acid sequence in a sample is provided using hybridization probes which competitively hybridize to a target nucleic acid. According to the method, a target nucleic acid sequence is hybridized to first and second hybridization probes which are complementary to overlapping portions of the target nucleic acid sequence, the first hybridization probe including a first complexing agent capable of forming a binding pair with a second complexing agent and the second hybridization probe including a detectable marker. The first complexing agent attached to the first hybridization probe is contacted with a second complexing agent, the second complexing agent being attached to a solid support such that when the first and second complexing agents are attached, target nucleic acid sequences hybridized to the first hybridization probe become immobilized on to the solid support. The immobilized target nucleic acids are then separated and detected by detecting the detectable marker attached to the second hybridization probe. A kit for performing the method is also provided.

  5. Detection and isolation of nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, J.N.; Straume, T.; Bogen, K.T.

    1997-04-01

    A method for detecting a target nucleic acid sequence in a sample is provided using hybridization probes which competitively hybridize to a target nucleic acid. According to the method, a target nucleic acid sequence is hybridized to first and second hybridization probes which are complementary to overlapping portions of the target nucleic acid sequence, the first hybridization probe including a first complexing agent capable of forming a binding pair with a second complexing agent and the second hybridization probe including a detectable marker. The first complexing agent attached to the first hybridization probe is contacted with a second complexing agent, the second complexing agent being attached to a solid support such that when the first and second complexing agents are attached, target nucleic acid sequences hybridized to the first hybridization probe become immobilized on to the solid support. The immobilized target nucleic acids are then separated and detected by detecting the detectable marker attached to the second hybridization probe. A kit for performing the method is also provided. 7 figs.

  6. Sequence Comparison and Phylogeny of Nucleotide Sequence of Coat Protein and Nucleic Acid Binding Protein of a Distinct Isolate of Shallot virus X from India.

    PubMed

    Majumder, S; Baranwal, V K

    2011-06-01

    Shallot virus X (ShVX), a type species in the genus Allexivirus of the family Alfaflexiviridae has been associated with shallot plants in India and other shallot growing countries like Russia, Germany, Netherland, and New Zealand. Coat protein (CP) and nucleic acid binding protein (NB) region of the virus was obtained by reverse transcriptase polymerase chain reaction from scales leaves of shallot bulbs. The partial cDNA contained two open reading frames encoding proteins of molecular weights of 28.66 and 14.18 kDa belonging to Flexi_CP super-family and viral NB super-family, respectively. The percent identity and phylogenetic analysis of amino acid sequences of CP and NB region of the virus associated with shallot indicated that it was a distinct isolate of ShVX. PMID:23637504

  7. Massively parallel 454-sequencing of fungal communities in Quercus spp. ectomycorrhizas indicates seasonal dynamics in urban and rural sites.

    PubMed

    Jumpponen, Ari; Jones, Kenneth L; David Mattox, J; Yaege, Chulee

    2010-03-01

    We analysed two sites within and outside an urban development in a rural background to estimate the fungal richness, diversity and community composition in Quercus spp. ectomycorrhizas using massively parallel 454-sequencing in combination with DNA-tagging. Our analyses indicated that shallow sequencing ( approximately 150 sequences) of a large number of samples (192 in total) provided data that allowed identification of seasonal trends within the fungal communities: putative root-associated antagonists and saprobes that were abundant early in the growing season were replaced by common ectomycorrhizal fungi in the course of the growing season. Ordination analyses identified a number of factors that were correlated with the observed communities including host species as well as soil organic matter, nutrient and heavy metal enrichment. Overall, our application of the high throughput 454 sequencing provided an expedient means for characterization of fungal communities. PMID:20331769

  8. On How "n" and "i" Turned out to Become Indices in Mathematical Sequences and Formulae

    ERIC Educational Resources Information Center

    Schubring, Gert

    2011-01-01

    The evolution of indexing sequences and series has been little studied so far, although this topic presents difficulties for students. This symbolisation signified, however, an important means for operating more generally with finite and infinite series. This paper investigates the first steps during the eighteenth century and then the different…

  9. Sour sweets and acidic beverage consumption are risk indicators for dental erosion.

    PubMed

    Søvik, Jenny Bogstad; Skudutyte-Rysstad, Rasa; Tveit, Anne B; Sandvik, Leiv; Mulic, Aida

    2015-01-01

    This study aimed to investigate the association between dental erosive wear and potential background, behavioural and dietary risk indicators and to assess whether there is a dose-response relationship between the level of acidic beverage consumption and dental erosive wear among adolescents. Of 846 adolescents (aged 16-18 years) scheduled for dental recall examinations, 795 (94%) accepted to participate. All participants completed a self-administered questionnaire regarding their background (gender and age), tooth-brushing frequency and dietary habits (the amount and frequency of acidic food and beverage consumption as well as the chosen method and manner of consuming acidic drinks). The association between the presence of erosive lesions and the possible risk indicators was assessed by logistic regression analyses. Of all participants examined, 37% had ≥3 surfaces with dental erosions and were considered to be affected individuals. In the present study, multivariate logistic analyses revealed a significant association between the dental erosive wear and high consumption of sour sweets and sports drinks. The tooth-brushing frequency was not significantly associated with dental erosive wear. Additionally, to the best of our knowledge, the results are the first to indicate a dose-response relationship between the daily consumption of acidic drinks and dental erosive wear. PMID:25765077

  10. Conservation of Shannon's redundancy for proteins. [information theory applied to amino acid sequences

    NASA Technical Reports Server (NTRS)

    Gatlin, L. L.

    1974-01-01

    Concepts of information theory are applied to examine various proteins in terms of their redundancy in natural originators such as animals and plants. The Monte Carlo method is used to derive information parameters for random protein sequences. Real protein sequence parameters are compared with the standard parameters of protein sequences having a specific length. The tendency of a chain to contain some amino acids more frequently than others and the tendency of a chain to contain certain amino acid pairs more frequently than other pairs are used as randomness measures of individual protein sequences. Non-periodic proteins are generally found to have random Shannon redundancies except in cases of constraints due to short chain length and genetic codes. Redundant characteristics of highly periodic proteins are discussed. A degree of periodicity parameter is derived.

  11. High-affinity homologous peptide nucleic acid probes for targeting a quadruplex-forming sequence from a MYC promoter element.

    PubMed

    Roy, Subhadeep; Tanious, Farial A; Wilson, W David; Ly, Danith H; Armitage, Bruce A

    2007-09-18

    Guanine-rich DNA and RNA sequences are known to fold into secondary structures known as G-quadruplexes. Recent biochemical evidence along with the discovery of an increasing number of sequences in functionally important regions of the genome capable of forming G-quadruplexes strongly indicates important biological roles for these structures. Thus, molecular probes that can selectively target quadruplex-forming sequences (QFSs) are envisioned as tools to delineate biological functions of quadruplexes as well as potential therapeutic agents. Guanine-rich peptide nucleic acids have been previously shown to hybridize to homologous DNA or RNA sequences forming PNA-DNA (or RNA) quadruplexes. For this paper we studied the hybridization of an eight-mer G-rich PNA to a quadruplex-forming sequence derived from the promoter region of the MYC proto-oncogene. UV melting analysis, fluorescence assays, and surface plasmon resonance experiments reveal that this PNA binds to the MYC QFS in a 2:1 stoichiometry and with an average binding constant Ka = (2.0 +/- 0.2) x 10(8) M(-1) or Kd = 5.0 nM. In addition, experiments carried out with short DNA targets revealed a dependence of the affinity on the sequence of bases in the loop region of the DNA. A structural model for the hybrid quadruplex is proposed, and implications for gene targeting by G-rich PNAs are discussed. PMID:17718513

  12. Conversion of amino-acid sequence in proteins to classical music: search for auditory patterns

    PubMed Central

    2007-01-01

    We have converted genome-encoded protein sequences into musical notes to reveal auditory patterns without compromising musicality. We derived a reduced range of 13 base notes by pairing similar amino acids and distinguishing them using variations of three-note chords and codon distribution to dictate rhythm. The conversion will help make genomic coding sequences more approachable for the general public, young children, and vision-impaired scientists. PMID:17477882

  13. Protein location prediction using atomic composition and global features of the amino acid sequence

    SciTech Connect

    Cherian, Betsy Sheena; Nair, Achuthsankar S.

    2010-01-22

    Subcellular location of protein is constructive information in determining its function, screening for drug candidates, vaccine design, annotation of gene products and in selecting relevant proteins for further studies. Computational prediction of subcellular localization deals with predicting the location of a protein from its amino acid sequence. For a computational localization prediction method to be more accurate, it should exploit all possible relevant biological features that contribute to the subcellular localization. In this work, we extracted the biological features from the full length protein sequence to incorporate more biological information. A new biological feature, distribution of atomic composition is effectively used with, multiple physiochemical properties, amino acid composition, three part amino acid composition, and sequence similarity for predicting the subcellular location of the protein. Support Vector Machines are designed for four modules and prediction is made by a weighted voting system. Our system makes prediction with an accuracy of 100, 82.47, 88.81 for self-consistency test, jackknife test and independent data test respectively. Our results provide evidence that the prediction based on the biological features derived from the full length amino acid sequence gives better accuracy than those derived from N-terminal alone. Considering the features as a distribution within the entire sequence will bring out underlying property distribution to a greater detail to enhance the prediction accuracy.

  14. The thermo-hand method: evaluation of a new indicator pad for acid permeation of chemical protective gloves.

    PubMed

    Vo, Evanly; Nicholson, Jonathan; Gao, Pengfei; Zhuang, Zhenzhen; Berardinelli, Stephen P

    2003-01-01

    The thermo-hand method was developed to evaluate a new indicator pad for acid permeation through chemical protective gloves under in-use conditions (controlled conditions for the hand's skin temperature, hand movements, and relative humidity inside gloves). An indicator pad was used to detect both organic and inorganic acid permeation through glove materials. Breakthrough times for five types of gloves were determined and found to range from 5 to 308 min for propionic acid, from 4 to 293 min for acrylic acid, and from 15 min to >6 hours for HCl. Quantification was performed for propionic and acrylic acids following solvent desorption and gas chromatography. Both acids exhibited >99% adsorption (including the volume of acid, which reacted with an indicator to contribute the color change) on the pads at a spiking level of 1.8 micro L for each acid. Acid recovery for the system was calculated for each acid, with results ranging from 52-72% (RSD < or =4.0%) for both acids over the spiking range 0.2-1.8 micro L. The quantitative mass of the acids on the pads at the time of breakthrough detection ranged from 253-276 and 270-296 micro g/cm(2) for propionic acid and acrylic acid, respectively. The thermo-hand method and a new acid indicator pad together should be useful in detecting, collecting, and quantitatively analyzing acid permeation samples in the workplace. PMID:14674803

  15. Structural analysis of complementary DNA and amino acid sequences of human and rat androgen receptors

    SciTech Connect

    Chang, C.; Kokontis, J.; Liao, S. )

    1988-10-01

    Structural analysis of cDNAs for human and rat androgen receptors (ARs) indicates that the amino-terminal regions of ARs are rich in oligo- and poly(amino acid) motifs as in some homeotic genes. The human AR has a long stretch of repeated glycines, whereas rat AR has a long stretch of glutamines. There is a considerable sequence similarity among ARs and the receptors for glucocorticoids, progestins, and mineralocorticoids within the steroid-binding domains. The cysteine-rich DNA-binding domains are well conserved. Translation of mRNA transcribed from AR cDNAs yielded 94- and 76-kDa proteins and smaller forms that bind to DNA and have high affinity toward androgens. These rat or human ARs were recognized by human autoantibodies to natural Ars. Molecular hybridization studies, using AR cDNAs as probes, indicated that the ventral prostate and other male accessory organs are rich in AR mRNA and that the production of AR mRNA in the target organs may be autoregulated by androgens.

  16. [Repetitive DNA sequences as an indicator of the level of genetic isolation in fish].

    PubMed

    Shubina, E A; Ponomareva, E V; Klimov, A V; Klimova, A V; Kedrova, O S

    2015-01-01

    Although the functional role is still unknown for most types of nuclear noncoding repetitive sequences, some of them proved to provide adequate phylogenetic and taxonomic markers for studying the genetic relationships of organisms at the species and within-species levels. Several markers were used in this work. First, microsatellite markers were used to examine populations varying in the extent of genetic subdivision in marine and anadromous fish, including the Chilean jack mackerel Trachurus murphyi, anadromous brown trout Salmo trutta, and isolated and anadromous char populations. Locus polymorphism was proportional to the gene flow between populations in all cases. Second, satellite DNA was used to study the phylogenetic relationships within the genera Salmo, Oncorhynchus, Salvelinus, and Coregonus. Genetic distances agreed well with the taxonomic relationships based on morphological traits and various biochemical markers and correlated with the evolutionary ages estimated for the groups by other markers. Third, RAPD PCR with a set of 20-mer primers was performed to study the genus Coregonus and anadromous and isolated populations and species of the genus Salvelinus. The resulting phylogenetic trees may help to resolve some disputable taxonomic issues for the groups. A comparison showed that several RAPD-detected sequences contain conserved fragments of coding sequences and polymorphic repeats (minisatellites) from intergenic regions or introns. The finding point to a nonrandom nature of repetitive DNA divergence and may reflect the evolution of the fish groups examined. Heterochromatic satellite repeats were assumed to contribute to generating a reproductive barrier. PMID:26107893

  17. Multimodal phylogeny for taxonomy: integrating information from nucleotide and amino acid sequences.

    PubMed

    Bicego, Manuele; Dellaglio, Franco; Felis, Giovanna E

    2007-10-01

    The crucial role played by the analysis of microbial diversity in biotechnology-based innovations has increased the interest in the microbial taxonomy research area. Phylogenetic sequence analyses have contributed significantly to the advances in this field, also in the view of the large amount of sequence data collected in recent years. Phylogenetic analyses could be realized on the basis of protein-encoding nucleotide sequences or encoded amino acid molecules: these two mechanisms present different peculiarities, still starting from two alternative representations of the same information. This complementarity could be exploited to achieve a multimodal phylogenetic scheme that is able to integrate gene and protein information in order to realize a single final tree. This aspect has been poorly addressed in the literature. In this paper, we propose to integrate the two phylogenetic analyses using basic schemes derived from the multimodality fusion theory (or multiclassifier systems theory), a well-founded and rigorous branch for which its powerfulness has already been demonstrated in other pattern recognition contexts. The proposed approach could be applied to distance matrix-based phylogenetic techniques (like neighbor joining), resulting in a smart and fast method. The proposed methodology has been tested in a real case involving sequences of some species of lactic acid bacteria. With this dataset, both nucleotide sequence- and amino acid sequence-based phylogenetic analyses present some drawbacks, which are overcome with the multimodal analysis. PMID:17933011

  18. Phospholipid fatty acids as physiological indicators of Paracoccus denitrificans encapsulated in silica sol-gel hydrogels.

    PubMed

    Trögl, Josef; Jirková, Ivana; Kuráň, Pavel; Akhmetshina, Elmira; Brovdyová, Taťjána; Sirotkin, Alexander; Kirilina, Tatiana

    2015-01-01

    The phospholipid fatty acid (PLFA) content was determined in samples of Paracoccus denitrificans encapsulated in silica hydrogel films prepared from prepolymerized tetramethoxysilane (TMOS). Immediately after encapsulation the total PLFA concentration was linearly proportional to the optical density (600 nm) of the input microbial suspension (R2 = 0.99). After 7 days this relationship remained linear, but with significantly decreased slope, indicating a higher extinction of bacteria in suspensions of input concentration 108 cells/mL and higher. trans-Fatty acids, indicators of cytoplasmatic membrane disturbances, were below the detection limit. The cy/pre ratio (i.e., ratio of cyclopropylated fatty acids (cy17:0 + cy19:0) to their metabolic precursors (16:1ω7 + 18:1ω7)), an indicator of the transition of the culture to a stationary growth-phase, decreased depending on co-immobilization of nutrients in the order phosphate buffer > mineral medium > Luria Broth rich medium. The ratio, too, was logarithmically proportional to cell concentration. These results confirm the applicability of total PLFA as an indicator for the determination of living biomass and cy/pre ratio for determination of nutrient limitation of microorganisms encapsulated in sol-gel matrices. This may be of interest for monitoring of sol-gel encapsulated bacteria proposed as optical recognition elements in biosensor construction, as well as other biotechnological applications. PMID:25690547

  19. Phospholipid Fatty Acids as Physiological Indicators of Paracoccus denitrificans Encapsulated in Silica Sol-Gel Hydrogels

    PubMed Central

    Trögl, Josef; Jirková, Ivana; Kuráň, Pavel; Akhmetshina, Elmira; Brovdyová, Tat′jána; Sirotkin, Alexander; Kirilina, Tatiana

    2015-01-01

    The phospholipid fatty acid (PLFA) content was determined in samples of Paracoccus denitrificans encapsulated in silica hydrogel films prepared from prepolymerized tetramethoxysilane (TMOS). Immediately after encapsulation the total PLFA concentration was linearly proportional to the optical density (600 nm) of the input microbial suspension (R2 = 0.99). After 7 days this relationship remained linear, but with significantly decreased slope, indicating a higher extinction of bacteria in suspensions of input concentration 108 cells/mL and higher. trans-Fatty acids, indicators of cytoplasmatic membrane disturbances, were below the detection limit. The cy/pre ratio (i.e., ratio of cyclopropylated fatty acids (cy17:0 + cy19:0) to their metabolic precursors (16:1ω7 + 18:1ω7)), an indicator of the transition of the culture to a stationary growth-phase, decreased depending on co-immobilization of nutrients in the order phosphate buffer > mineral medium > Luria Broth rich medium. The ratio, too, was logarithmically proportional to cell concentration. These results confirm the applicability of total PLFA as an indicator for the determination of living biomass and cy/pre ratio for determination of nutrient limitation of microorganisms encapsulated in sol-gel matrices. This may be of interest for monitoring of sol-gel encapsulated bacteria proposed as optical recognition elements in biosensor construction, as well as other biotechnological applications. PMID:25690547

  20. The amino-acid sequence of leghemoglobin component a from Phaseolus vulgaris (kidney bean).

    PubMed

    Lehtovaara, P; Ellfolk, N

    1975-06-01

    1. Leghemoglobin component a from Phaseolus vulgaris (kidney bean) was digested with trypsin; 15 tryptic peptides and free lysine were purified and the amino acid sequences of the peptides determined. 2. The internal order of the tryptic peptides was determined by the bridge peptides obtained from the thermolytic digest and the dilute acid hydrolyzate of kidney bean leghemoglobin a; 12 thermolytic peptides and two acid hydrolysis peptides were purified and the sequences were partially or completely determined. 3. The complete amino acid sequence of kidney bean leghemoglobin a is compared to that of leghemoglobin a from soybean (Glycine max) and to some animal globins. As regards sequence, the kidney bean globin has 79% identity with the soybean globin and 21% identity with human hemoglobin gamma-chain. Seven of the 14 amino acid residues common to most globins are found in the kidney bean globin. Trp-15 and Tyr-145 are evolutionarily conserved in this globin, which confirms the concept of a common origin of animal and plant globins. PMID:809270

  1. Draft genome sequence of the docosahexaenoic acid producing thraustochytrid Aurantiochytrium sp. T66.

    PubMed

    Liu, Bin; Ertesvåg, Helga; Aasen, Inga Marie; Vadstein, Olav; Brautaset, Trygve; Heggeset, Tonje Marita Bjerkan

    2016-06-01

    Thraustochytrids are unicellular, marine protists, and there is a growing industrial interest in these organisms, particularly because some species, including strains belonging to the genus Aurantiochytrium, accumulate high levels of docosahexaenoic acid (DHA). Here, we report the draft genome sequence of Aurantiochytrium sp. T66 (ATCC PRA-276), with a size of 43 Mbp, and 11,683 predicted protein-coding sequences. The data has been deposited at DDBJ/EMBL/Genbank under the accession LNGJ00000000. The genome sequence will contribute new insight into DHA biosynthesis and regulation, providing a basis for metabolic engineering of thraustochytrids. PMID:27222814

  2. A classification of glycosyl hydrolases based on amino acid sequence similarities.

    PubMed Central

    Henrissat, B

    1991-01-01

    The amino acid sequences of 301 glycosyl hydrolases and related enzymes have been compared. A total of 291 sequences corresponding to 39 EC entries could be classified into 35 families. Only ten sequences (less than 5% of the sample) could not be assigned to any family. With the sequences available for this analysis, 18 families were found to be monospecific (containing only one EC number) and 17 were found to be polyspecific (containing at least two EC numbers). Implications on the folding characteristics and mechanism of action of these enzymes and on the evolution of carbohydrate metabolism are discussed. With the steady increase in sequence and structural data, it is suggested that the enzyme classification system should perhaps be revised. PMID:1747104

  3. New families in the classification of glycosyl hydrolases based on amino acid sequence similarities.

    PubMed Central

    Henrissat, B; Bairoch, A

    1993-01-01

    301 glycosyl hydrolases and related enzymes corresponding to 39 EC entries of the I.U.B. classification system have been classified into 35 families on the basis of amino-acid-sequence similarities [Henrissat (1991) Biochem. J. 280, 309-316]. Approximately half of the families were found to be monospecific (containing only one EC number), whereas the other half were found to be polyspecific (containing at least two EC numbers). A > 60% increase in sequence data for glycosyl hydrolases (181 additional enzymes or enzyme domains sequences have since become available) allowed us to update the classification not only by the addition of more members to already identified families, but also by the finding of ten new families. On the basis of a comparison of 482 sequences corresponding to 52 EC entries, 45 families, out of which 22 are polyspecific, can now be defined. This classification has been implemented in the SWISS-PROT protein sequence data bank. PMID:8352747

  4. Sequence-specific purification of nucleic acids by PNA-controlled hybrid selection.

    PubMed

    Orum, H; Nielsen, P E; Jørgensen, M; Larsson, C; Stanley, C; Koch, T

    1995-09-01

    Using an oligohistidine peptide nucleic acids (oligohistidine-PNA) chimera, we have developed a rapid hybrid selection method that allows efficient, sequence-specific purification of a target nucleic acid. The method exploits two fundamental features of PNA. First, that PNA binds with high affinity and specificity to its complementary nucleic acid. Second, that amino acids are easily attached to the PNA oligomer during synthesis. We show that a (His)6-PNA chimera exhibits strong binding to chelated Ni2+ ions without compromising its native PNA hybridization properties. We further show that these characteristics allow the (His)6-PNA/DNA complex to be purified by the well-established method of metal ion affinity chromatography using a Ni(2+)-NTA (nitrilotriactic acid) resin. Specificity and efficiency are the touchstones of any nucleic acid purification scheme. We show that the specificity of the (His)6-PNA selection approach is such that oligonucleotides differing by only a single nucleotide can be selectively purified. We also show that large RNAs (2224 nucleotides) can be captured with high efficiency by using multiple (His)6-PNA probes. PNA can hybridize to nucleic acids in low-salt concentrations that destabilize native nucleic acid structures. We demonstrate that this property of PNA can be utilized to purify an oligonucleotide in which the target sequence forms part of an intramolecular stem/loop structure. PMID:7495562

  5. In silico comparative analysis of DNA and amino acid sequences for prion protein gene.

    PubMed

    Kim, Y; Lee, J; Lee, C

    2008-01-01

    Genetic variability might contribute to species specificity of prion diseases in various organisms. In this study, structures of the prion protein gene (PRNP) and its amino acids were compared among species of which sequence data were available. Comparisons of PRNP DNA sequences among 12 species including human, chimpanzee, monkey, bovine, ovine, dog, mouse, rat, wallaby, opossum, chicken and zebrafish allowed us to identify candidate regulatory regions in intron 1 and 3'-untranslated region (UTR) in addition to the coding region. Highly conserved putative binding sites for transcription factors, such as heat shock factor 2 (HSF2) and myocite enhancer factor 2 (MEF2), were discovered in the intron 1. In 3'-UTR, the functional sequence (ATTAAA) for nucleus-specific polyadenylation was found in all the analysed species. The functional sequence (TTTTTAT) for maturation-specific polyadenylation was identically observed only in ovine, and one or two nucleotide mismatches in the other species. A comparison of the amino acid sequences in 53 species revealed a large sequence identity. Especially the octapeptide repeat region was observed in all the species but frog and zebrafish. Functional changes and susceptibility to prion diseases with various isoforms of prion protein could be caused by numeric variability and conformational changes discovered in the repeat sequences. PMID:18397498

  6. Amino acid sequence homology between Piv, an essential protein in site-specific DNA inversion in Moraxella lacunata, and transposases of an unusual family of insertion elements.

    PubMed Central

    Lenich, A G; Glasgow, A C

    1994-01-01

    Deletion analysis of the subcloned DNA inversion region of Moraxella lacunata indicates that Piv is the only M. lacunata-encoded factor required for site-specific inversion of the tfpQ/tfpI pilin segment. The predicted amino acid sequence of Piv shows significant homology solely with the transposases/integrases of a family of insertion sequence elements, suggesting that Piv is a novel site-specific recombinase. Images PMID:8021196

  7. Genetic structure of Florida green turtle rookeries as indicated by mitochondrial DNA control region sequences

    USGS Publications Warehouse

    Shamblin, Brian M.; Bagley, Dean A.; Ehrhart, Llewellyn M.; Desjardin, Nicole A.; Martin, R. Erik; Hart, Kristen M.; Naro-Maciel, Eugenia; Rusenko, Kirt; Stiner, John C.; Sobel, Debra; Johnson, Chris; Wilmers, Thomas; Wright, Laura J.; Nairn, Campbell J.

    2014-01-01

    Green turtle (Chelonia mydas) nesting has increased dramatically in Florida over the past two decades, ranking the Florida nesting aggregation among the largest in the Greater Caribbean region. Individual beaches that comprise several hundred kilometers of Florida’s east coast and Keys support tens to thousands of nests annually. These beaches encompass natural to highly developed habitats, and the degree of demographic partitioning among rookeries was previously unresolved. We characterized the genetic structure of ten Florida rookeries from Cape Canaveral to the Dry Tortugas through analysis of 817 base pair mitochondrial DNA (mtDNA) control region sequences from 485 nesting turtles. Two common haplotypes, CM-A1.1 and CM-A3.1, accounted for 87 % of samples, and the haplotype frequencies were strongly partitioned by latitude along Florida’s Atlantic coast. Most genetic structure occurred between rookeries on either side of an apparent genetic break in the vicinity of the St. Lucie Inlet that separates Hutchinson Island and Jupiter Island, representing the finest scale at which mtDNA structure has been documented in marine turtle rookeries. Florida and Caribbean scale analyses of population structure support recognition of at least two management units: central eastern Florida and southern Florida. More thorough sampling and deeper sequencing are necessary to better characterize connectivity among Florida green turtle rookeries as well as between the Florida nesting aggregation and others in the Greater Caribbean region.

  8. Sequence-independent and reversible photocontrol of transcription/expression systems using a photosensitive nucleic acid binder

    PubMed Central

    Estévez-Torres, André; Crozatier, Cécile; Diguet, Antoine; Hara, Tomoaki; Saito, Hirohide; Yoshikawa, Kenichi; Baigl, Damien

    2009-01-01

    To understand non-trivial biological functions, it is crucial to develop minimal synthetic models that capture their basic features. Here, we demonstrate a sequence-independent, reversible control of transcription and gene expression using a photosensitive nucleic acid binder (pNAB). By introducing a pNAB whose affinity for nucleic acids is tuned by light, in vitro RNA production, EGFP translation, and GFP expression (a set of reactions including both transcription and translation) were successfully inhibited in the dark and recovered after a short illumination at 365 nm. Our results indicate that the accessibility of the protein machinery to one or several nucleic acid binding sites can be efficiently regulated by changing the conformational/condensation state of the nucleic acid (DNA conformation or mRNA aggregation), thus regulating gene activity in an efficient, reversible, and sequence-independent manner. The possibility offered by our approach to use light to trigger various gene expression systems in a system-independent way opens interesting perspectives to study gene expression dynamics as well as to develop photocontrolled biotechnological procedures. PMID:19617550

  9. Amino acid sequence of a vitamin K-dependent Ca2+-binding peptide from bovine prothrombin.

    PubMed

    Howard, J B; Fausch, M D

    1975-08-10

    The amino acid sequence of a 31-residue peptide from bovine prothrombin has been determined. This peptide has been shown to contain the vitamin K-dependent modification required for Ca2+ binding (Nelsestuen, G. L., and Suttie, J. W. (1973) Proc. Natl. Acad. Sci. U. S. A. 70, 3366-3370) and the modified amino acid, gamma-carboxyglutamic acid (Nelsestuen, G. L., Zytkovicz, T., and Howard, J. B. (1974) J. Biol. Chem. 249, 6347-6350). The peptide was shown to correspond to residues 12 to 42 of prothrombin. PMID:807581

  10. Amino acid sequences around the cysteine residues of rabbit muscle triose phosphate isomerase

    PubMed Central

    Miller, Janet C.; Waley, S. G.

    1971-01-01

    1. The nature of the subunits in rabbit muscle triose phosphate isomerase has been investigated. 2. Amino acid analyses show that there are five cysteine residues and two methionine residues/subunit. 3. The amino acid sequences around the cysteine residues have been determined; these account for about 75 residues. 4. Cleavage at the methionine residues with cyanogen bromide gave three fragments. 5. These results show that the subunits correspond to polypeptide chains, containing about 230 amino acid residues. The chains in triose phosphate isomerase seem to be shorter than those of other glycolytic enzymes. PMID:5165707

  11. Complete amino acid sequence of the Mu heavy chain of a human IgM immunoglobulin.

    PubMed

    Putnam, F W; Florent, G; Paul, C; Shinoda, T; Shimizu, A

    1973-10-19

    The amino acid sequence of the micro, chain of a human IgM immunoglobulin, including the location of all disulfide bridges and oligosaccharides, has been determined. The homology of the constant regions of immunoglobulin micro, gamma, alpha, and epsilon heavy chains reveals evolutionary relationships and suggests that two genes code for each heavy chain. PMID:4742735

  12. Draft Genome Sequence of the Butyric Acid Producer Clostridium tyrobutyricum Strain CIP I-776 (IFP923)

    PubMed Central

    Clément, Benjamin; Lopes Ferreira, Nicolas

    2016-01-01

    Here, we report the draft genome sequence of Clostridium tyrobutyricum CIP I-776 (IFP923), an efficient producer of butyric acid. The genome consists of a single chromosome of 3.19 Mb and provides useful data concerning the metabolic capacities of the strain. PMID:26941139

  13. Draft Genome Sequence of Perfluorooctane Acid-Degrading Bacterium Pseudomonas parafulva YAB-1

    PubMed Central

    Tang, Chongjian; Peng, Qingjing; Peng, Qingzhong

    2015-01-01

    Pseudomonas parafulva YAB-1, isolated from perfluorinated compound-contaminated soil, has the ability to degrade perfluorooctane acid (PFOA) compound. Here, we report the draft genome sequence and annotation of the PFOA-degrading bacterium P. parafulva YAB-1. The data provide the basis to investigate the molecular mechanism of PFOA metabolism. PMID:26337877

  14. The developmental transcriptome landscape of bovine skeletal muscle defined by Ribo-Zero ribonucleic acid sequencing.

    PubMed

    Sun, X; Li, M; Sun, Y; Cai, H; Li, R; Wei, X; Lan, X; Huang, Y; Lei, C; Chen, H

    2015-12-01

    Ribonucleic acid sequencing (RNA-Seq) libraries are normally prepared with oligo(dT) selection of poly(A)+ mRNA, but it depends on intact total RNA samples. Recent studies have described Ribo-Zero technology, a novel method that can capture both poly(A)+ and poly(A)- transcripts from intact or fragmented RNA samples. We report here the first application of Ribo-Zero RNA-Seq for the analysis of the bovine embryonic, neonatal, and adult skeletal muscle whole transcriptome at an unprecedented depth. Overall, 19,893 genes were found to be expressed, with a high correlation of expression levels between the calf and the adult. Hundreds of genes were found to be highly expressed in the embryo and decreased at least 10-fold after birth, indicating their potential roles in embryonic muscle development. In addition, we present for the first time the analysis of global transcript isoform discovery in bovine skeletal muscle and identified 36,694 transcript isoforms. Transcriptomic data were also analyzed to unravel sequence variations; 185,036 putative SNP and 12,428 putative short insertions-deletions (InDel) were detected. Specifically, many stop-gain, stop-loss, and frameshift mutations were identified that probably change the relative protein production and sequentially affect the gene function. Notably, the numbers of stage-specific transcripts, alternative splicing events, SNP, and InDel were greater in the embryo than in the calf and the adult, suggesting that gene expression is most active in the embryo. The resulting view of the transcriptome at a single-base resolution greatly enhances the comprehensive transcript catalog and uncovers the global trends in gene expression during bovine skeletal muscle development. PMID:26641174

  15. The amino acid sequence of cytochrome c-555 from the methane-oxidizing bacterium Methylococcus capsulatus.

    PubMed Central

    Ambler, R P; Dalton, H; Meyer, T E; Bartsch, R G; Kamen, M D

    1986-01-01

    The amino acid sequence of the cytochrome c-555 from the obligate methanotroph Methylococcus capsulatus strain Bath (N.C.I.B. 11132) was determined. It is a single polypeptide chain of 96 residues, binding a haem group through the cysteine residues at positions 19 and 22, and the only methionine residue is a position 59. The sequence does not closely resemble that of any other cytochrome c that has yet been characterized. Detailed evidence for the amino acid sequence of the protein has been deposited as Supplementary Publication SUP 50131 (12 pages) at the British Library Lending Division, Boston Spa, West Yorkshire LS23 7BQ, U.K., from whom copies are available on prepayment. PMID:3006666

  16. Protein sequences and redox titrations indicate that the electron acceptors in reaction centers from heliobacteria are similar to Photosystem I

    NASA Technical Reports Server (NTRS)

    Trost, J. T.; Brune, D. C.; Blankenship, R. E.

    1992-01-01

    Photosynthetic reaction centers isolated from Heliobacillus mobilis exhibit a single major protein on SDS-PAGE of 47 000 Mr. Attempts to sequence the reaction center polypeptide indicated that the N-terminus is blocked. After enzymatic and chemical cleavage, four peptide fragments were sequenced from the Heliobacillus mobilis apoprotein. Only one of these sequences showed significant specific similarity to any of the protein and deduced protein sequences in the GenBank data base. This fragment is identical with 56% of the residues, including both cysteines, found in highly conserved region that is proposed to bind iron-sulfur center Fx in the Photosystem I reaction center peptide that is the psaB gene product. The similarity to the psaA gene product in this region is 48%. Redox titrations of laser-flash-induced photobleaching with millisecond decay kinetics on isolated reaction centers from Heliobacterium gestii indicate a midpoint potential of -414 mV with n = 2 titration behavior. In membranes, the behavior is intermediate between n = 1 and n = 2, and the apparent midpoint potential is -444 mV. This is compared to the behavior in Photosystem I, where the intermediate electron acceptor A1, thought to be a phylloquinone molecule, has been proposed to undergo a double reduction at low redox potentials in the presence of viologen redox mediators. These results strongly suggest that the acceptor side electron transfer system in reaction centers from heliobacteria is indeed analogous to that found in Photosystem I. The sequence similarities indicate that the divergence of the heliobacteria from the Photosystem I line occurred before the gene duplication and subsequent divergence that lead to the heterodimeric protein core of the Photosystem I reaction center.

  17. CHARACTERIZATION OF H CANYON CONDUCTIVITY METER INDICATIONS WITH ELEVATED URANIUM IN NITRIC ACID

    SciTech Connect

    Nash, C

    2007-10-31

    Solution conductivity data from the 1CU conductivity meter in H-Canyon shows that uranium concentration in the 0 to 30 gram per liter (g/L) range has no statistically significant effect on the calibration of free nitric acid measurement. Based on these results, no additional actions are needed on the 1CU Conductivity Meter prior to or during the processing of uranium solutions in the 0 to 30 g/L range. A model based only on free nitric acid concentration is shown to be appropriate for explaining the data. Data uncertainties for the free acid measurement of uranium-bearing solutions are 8.5% or less at 95% confidence. The analytical uncertainty for calibrating solutions is an order of magnitude smaller only when uranium is not present, allowing use of a more accurate analytical procedure. Literature work shows that at a free nitric acid level of 0.33 M, uranium concentration of 30 g/L and 25 C, solution conductivity is 96.4% of that of a uranium-free solution. The level of uncertainties in the literature data and its fitting equation do not justify calibration changes based on this small depression in solution conductivity. This work supports preparation of H-Canyon processing of Super Kukla fuel; however, the results will be applicable to the processing of any similar concentration uranium and nitric acid solution. Super Kukla fuel processing will increase the uranium concentration above the nominal zero to 10 g/L level, though not above 30 g/L. This work examined free nitric acid levels ranging from 0.18 to 0.52 molar. Temperature ranged from 27.9 to 28.3 C during conductivity testing. The data indicates that sequential order of measurement is not a significant factor. The conductivity meter was thus flushed effectively between measurements as desired.

  18. Allelic polymorphism in arabian camel ribonuclease and the amino acid sequence of bactrian camel ribonuclease.

    PubMed

    Welling, G W; Mulder, H; Beintema, J J

    1976-04-01

    Pancreatic ribonucleases from several species (whitetail deer, roe deer, guinea pig, and arabian camel) exhibit more than one amino acid at particular positions in their amino acid sequences. Since these enzymes were isolated from pooled pancreas, the origin of this heterogeneity is not clear. The pancreatic ribonucleases from 11 individual arabian camels (Camelus dromedarius) have been investigated with respect to the lysine-glutamine heterogeneity at position 103 (Welling et al., 1975). Six ribonucleases showed only one basic band and five showed two bands after polyacrylamide gel electrophoresis, suggesting a gene frequency of about 0.75 for the Lys gene and about 0.25 for the Gln gene. The amino acid sequence of bactrian camel (Camelus bactrianus) ribonuclease isolated from individual pancreatic tissue was determined and compared with that of arabian camel ribonuclease. The only difference was observed at position 103. In the ribonucleases from two unrelated bactrian camels, only glutamine was observed at that position. PMID:962846

  19. Characterization and cDNA sequence of Bothriechis schlegeliil-amino acid oxidase with antibacterial activity.

    PubMed

    Vargas Muñoz, Leidy Johana; Estrada-Gomez, Sebastian; Núñez, Vitelbina; Sanz, Libia; Calvete, Juan J

    2014-08-01

    Snake venoms are complex mixtures of proteins including l-amino acid oxidase (lAAO). A lAAO (named BslAAO) with a mass of 56kDa and a theoretical Ip of 5.79, was purified from Bothriechis schlegelii venom through size-exclusion, ion exchange and affinity chromatography. The entire protein sequence of 498 amino acids, was determined from cDNA using reverse-transcribed mRNA isolated from venom gland. The enzyme showed dose-dependent inhibition of bacterial growth. BslAAO showed inhibitory effect against S. aureus with a MIC of 4μg/mL and a MBC of 8μg/mL. Against Acinetobacter baumannii, showed a MIC of 2μg/mL and MBC of 4μg/mL, No effect was observed in Escherichia coli. This antibacterial activity was inhibited by catalase, indicating that antimicrobial activity was due to H2O2 production. BslAAO did not show any cytotoxic activity toward mouse myoblast cell line C2C12 or peripheral blood mononuclear cells. The enzyme oxidated l-Leu, with a Km of 16.37μM and a Vmax of 0.39μM/min. Snake venoms lAAOs, are potential frames of different therapeutics molecules since these enzymes exhibit low MICs and MBCs and show to be harmless to human cells due to microorganisms being generally several fold more sensitive to reactive oxygen species than human tissues. PMID:24875315

  20. Oxidized amino acids: culprits in human atherosclerosis and indicators of oxidative stress.

    PubMed

    Heinecke, Jay W

    2002-06-01

    Oxidized low-density lipoprotein (LDL) is implicated in atherogenesis, but the mechanisms that oxidize LDL in the human artery wall have proven difficult to identify. A powerful investigative approach is mass spectrometric quantification of the oxidized amino acids that are left in proteins by specific oxidation reactions. Comparison of these molecular fingerprints in biological samples with those produced in proteins by various in vitro oxidation systems can indicate which biochemical pathway has created damage in vivo. For example, the pattern of oxidized amino acids in proteins isolated from atherosclerotic lesions implicates reactive intermediates generated by myeloperoxidase, a major phagocyte enzyme. These intermediates include hypochlorous acid, tyrosyl radical, and reactive nitrogen species, each of which generates a different pattern of stable end products. Despite this strong evidence that myeloperoxidase promotes LDL oxidation in vivo, the antioxidant that has been tested most extensively in clinical trials, vitamin E, fails to inhibit myeloperoxidase pathways in vitro. Because the utility of an antioxidant depends critically on the nature of the pathway that inflicts tissue damage, interventions that specifically inhibit myeloperoxidase or other physiologically relevant pathways would be more logical candidates for the prevention of cardiovascular disease. Moreover, levels of oxidized amino acids in urine and plasma might reflect those in tissues and therefore identify individuals with high levels of oxidative stress. Trials with such subjects would seem more likely to uncover effective antioxidant therapies than trials involving the general population. PMID:12031894

  1. Use of a structural alphabet to find compatible folds for amino acid sequences

    PubMed Central

    Mahajan, Swapnil; de Brevern, Alexandre G; Sanejouand, Yves-Henri; Srinivasan, Narayanaswamy; Offmann, Bernard

    2015-01-01

    The structural annotation of proteins with no detectable homologs of known 3D structure identified using sequence-search methods is a major challenge today. We propose an original method that computes the conditional probabilities for the amino-acid sequence of a protein to fit to known protein 3D structures using a structural alphabet, known as “Protein Blocks” (PBs). PBs constitute a library of 16 local structural prototypes that approximate every part of protein backbone structures. It is used to encode 3D protein structures into 1D PB sequences and to capture sequence to structure relationships. Our method relies on amino acid occurrence matrices, one for each PB, to score global and local threading of query amino acid sequences to protein folds encoded into PB sequences. It does not use any information from residue contacts or sequence-search methods or explicit incorporation of hydrophobic effect. The performance of the method was assessed with independent test datasets derived from SCOP 1.75A. With a Z-score cutoff that achieved 95% specificity (i.e., less than 5% false positives), global and local threading showed sensitivity of 64.1% and 34.2%, respectively. We further tested its performance on 57 difficult CASP10 targets that had no known homologs in PDB: 38 compatible templates were identified by our approach and 66% of these hits yielded correctly predicted structures. This method scales-up well and offers promising perspectives for structural annotations at genomic level. It has been implemented in the form of a web-server that is freely available at http://www.bo-protscience.fr/forsa. PMID:25297700

  2. Use of a structural alphabet to find compatible folds for amino acid sequences.

    PubMed

    Mahajan, Swapnil; de Brevern, Alexandre G; Sanejouand, Yves-Henri; Srinivasan, Narayanaswamy; Offmann, Bernard

    2015-01-01

    The structural annotation of proteins with no detectable homologs of known 3D structure identified using sequence-search methods is a major challenge today. We propose an original method that computes the conditional probabilities for the amino-acid sequence of a protein to fit to known protein 3D structures using a structural alphabet, known as "Protein Blocks" (PBs). PBs constitute a library of 16 local structural prototypes that approximate every part of protein backbone structures. It is used to encode 3D protein structures into 1D PB sequences and to capture sequence to structure relationships. Our method relies on amino acid occurrence matrices, one for each PB, to score global and local threading of query amino acid sequences to protein folds encoded into PB sequences. It does not use any information from residue contacts or sequence-search methods or explicit incorporation of hydrophobic effect. The performance of the method was assessed with independent test datasets derived from SCOP 1.75A. With a Z-score cutoff that achieved 95% specificity (i.e., less than 5% false positives), global and local threading showed sensitivity of 64.1% and 34.2%, respectively. We further tested its performance on 57 difficult CASP10 targets that had no known homologs in PDB: 38 compatible templates were identified by our approach and 66% of these hits yielded correctly predicted structures. This method scales-up well and offers promising perspectives for structural annotations at genomic level. It has been implemented in the form of a web-server that is freely available at http://www.bo-protscience.fr/forsa. PMID:25297700

  3. Rapid and Sensitive Isothermal Detection of Nucleic-acid Sequence by Multiple Cross Displacement Amplification

    PubMed Central

    Wang, Yi; Wang, Yan; Ma, Ai-Jing; Li, Dong-Xun; Luo, Li-Juan; Liu, Dong-Xin; Jin, Dong; Liu, Kai; Ye, Chang-Yun

    2015-01-01

    We have devised a novel amplification strategy based on isothermal strand-displacement polymerization reaction, which was termed multiple cross displacement amplification (MCDA). The approach employed a set of ten specially designed primers spanning ten distinct regions of target sequence and was preceded at a constant temperature (61–65 °C). At the assay temperature, the double-stranded DNAs were at dynamic reaction environment of primer-template hybrid, thus the high concentration of primers annealed to the template strands without a denaturing step to initiate the synthesis. For the subsequent isothermal amplification step, a series of primer binding and extension events yielded several single-stranded DNAs and single-stranded single stem-loop DNA structures. Then, these DNA products enabled the strand-displacement reaction to enter into the exponential amplification. Three mainstream methods, including colorimetric indicators, agarose gel electrophoresis and real-time turbidity, were selected for monitoring the MCDA reaction. Moreover, the practical application of the MCDA assay was successfully evaluated by detecting the target pathogen nucleic acid in pork samples, which offered advantages on quick results, modest equipment requirements, easiness in operation, and high specificity and sensitivity. Here we expounded the basic MCDA mechanism and also provided details on an alternative (Single-MCDA assay, S-MCDA) to MCDA technique. PMID:26154567

  4. Shigella sonnei genome sequencing and phylogenetic analysis indicate recent global dissemination from Europe

    PubMed Central

    Holt, Kathryn E.; Baker, Stephen; Weill, François-Xavier; Holmes, Edward C.; Kitchen, Andrew; Yu, Jun; Sangal, Vartul; Brown, Derek J.; Coia, John E.; Kim, Dong Wook; Choi, Seon Young; Kim, Su Hee; da Silveira, Wanderley D.; Pickard, Derek J.; Farrar, Jeremy J.; Parkhill, Julian; Dougan, Gordon; Thomson, Nicholas R.

    2012-01-01

    Shigella are human-adapted Escherichia coli that have gained the ability to invade the human gut mucosa and cause dysentery1,2, spreading efficiently via low-dose fecal-oral transmission3,4. Historically, S. sonnei has been predominantly responsible for dysentery in developed countries, but is now emerging as a problem in the developing world, apparently replacing the more diverse S. flexneri in areas undergoing economic development and improvements in water quality4-6. Classical approaches have shown S. sonnei is genetically conserved and clonal7. We report here whole-genome sequencing of 132 globally-distributed isolates. Our phylogenetic analysis shows that the current S. sonnei population descends from a common ancestor that existed less than 500 years ago and has diversified into several distinct lineages with unique characteristics. Our analysis suggests the majority of this diversification occurred in Europe, followed by more recent establishment of local pathogen populations in other continents predominantly due to the pandemic spread of a single, rapidly-evolving, multidrug resistant lineage. PMID:22863732

  5. Software scripts for quality checking of high-throughput nucleic acid sequencers.

    PubMed

    Lazo, G R; Tong, J; Miller, R; Hsia, C; Rausch, C; Kang, Y; Anderson, O D

    2001-06-01

    We have developed a graphical interface to allow the researcher to view and assess the quality of sequencing results using a series of program scripts developed to process data generated by automated sequencers. The scripts are written in Perl programming language and are executable under the cgibin directory of a Web server environment. The scripts direct nucleic acid sequencing trace file data output from automated sequencers to be analyzed by the phred molecular biology program and are displayed as graphical hypertext mark-up language (HTML) pages. The scripts are mainly designed to handle 96-well microtiter dish samples, but the scripts are also able to read data from 384-well microtiter dishes 96 samples at a time. The scripts may be customized for different laboratory environments and computer configurations. Web links to the sources and discussion page are provided. PMID:11414222

  6. Effects of ammonium on elemental nutrition of red spruce and indicator plants grown in acid soil

    SciTech Connect

    Hoelldampf, B.; Barker, A.V. )

    1993-01-01

    Decline of high elevation red spruce forests in the northeastern United States has been related to acid rain, particularly with respect to the deposition of nitrogenous materials. Ca and Mg deficiencies may be induced by input of air-borne nitrogenous nutrients into the forest ecosystem. This research investigated the effects of N nutrition on mineral nutrition of red spruce and radish, as an indicator plant, grown in acid forest soil. Red spruce and radishes in the greenhouse were treated with complete nutrient solutions with 15 mM N supplied as 0, 3.75, 7.5, 11.25, or 15 mM NH[sub 4][sup +] with the remainder being supplied as NO[sub 3][sup [minus

  7. Identification of planarian homeobox sequences indicates the antiquity of most Hox/homeotic gene subclasses.

    PubMed Central

    Balavoine, G; Telford, M J

    1995-01-01

    The homeotic gene complex (HOM-C) is a cluster of genes involved in the anteroposterior axial patterning of animal embryos. It is composed of homeobox genes belonging to the Hox/HOM superclass. Originally discovered in Drosophila, Hox/HOM genes have been identified in organisms as distantly related as arthropods, vertebrates, nematodes, and cnidarians. Data obtained in parallel from the organization of the complex, the domains of gene expression during embryogenesis, and phylogenetic relationships allow the subdivision of the Hox/HOM superclass into five classes (lab, pb/Hox3, Dfd, Antp, and Abd-B) that appeared early during metazoan evolution. We describe a search for homologues of these genes in platyhelminths, triploblast metazoans emerging as an outgroup to the great coelomate ensemble. A degenerate PCR screening for Hox/HOM homeoboxes in three species of triclad planarians has revealed 10 types of Antennapedia-like genes. The homeobox-containing sequences of these PCR fragments allowed the amplification of the homeobox-coding exons for five of these genes in the species Polycelis nigra. A phylogenetic analysis shows that two genes are clear orthologues of Drosophila labial, four others are members of a Dfd/Antp superclass, and a seventh gene, although more difficult to classify with certainty, may be related to the pb/Hox3 class. Together with previously identified Hox/HOM genes in other flatworms, our analyses demonstrate the existence of an elaborate family of Hox/HOM genes in the ancestor of all triploblast animals. Images Fig. 4 PMID:7638172

  8. Nucleotide and predicted amino acid sequences of cloned human and mouse preprocathepsin B cDNAs.

    PubMed Central

    Chan, S J; San Segundo, B; McCormick, M B; Steiner, D F

    1986-01-01

    Cathepsin B is a lysosomal thiol proteinase that may have additional extralysosomal functions. To further our investigations on the structure, mode of biosynthesis, and intracellular sorting of this enzyme, we have determined the complete coding sequences for human and mouse preprocathepsin B by using cDNA clones isolated from human hepatoma and kidney phage libraries. The nucleotide sequences predict that the primary structure of preprocathepsin B contains 339 amino acids organized as follows: a 17-residue NH2-terminal prepeptide sequence followed by a 62-residue propeptide region, 254 residues in mature (single chain) cathepsin B, and a 6-residue extension at the COOH terminus. A comparison of procathepsin B sequences from three species (human, mouse, and rat) reveals that the homology between the propeptides is relatively conserved with a minimum of 68% sequence identity. In particular, two conserved sequences in the propeptide that may be functionally significant include a potential glycosylation site and the presence of a single cysteine at position 59. Comparative analysis of the three sequences also suggests that processing of procathepsin B is a multistep process, during which enzymatically active intermediate forms may be generated. The availability of the cDNA clones will facilitate the identification of possible active or inactive intermediate processive forms as well as studies on the transcriptional regulation of the cathepsin B gene. PMID:3463996

  9. Efficient Nucleic Acid Extraction and 16S rRNA Gene Sequencing for Bacterial Community Characterization.

    PubMed

    Anahtar, Melis N; Bowman, Brittany A; Kwon, Douglas S

    2016-01-01

    There is a growing appreciation for the role of microbial communities as critical modulators of human health and disease. High throughput sequencing technologies have allowed for the rapid and efficient characterization of bacterial communities using 16S rRNA gene sequencing from a variety of sources. Although readily available tools for 16S rRNA sequence analysis have standardized computational workflows, sample processing for DNA extraction remains a continued source of variability across studies. Here we describe an efficient, robust, and cost effective method for extracting nucleic acid from swabs. We also delineate downstream methods for 16S rRNA gene sequencing, including generation of sequencing libraries, data quality control, and sequence analysis. The workflow can accommodate multiple samples types, including stool and swabs collected from a variety of anatomical locations and host species. Additionally, recovered DNA and RNA can be separated and used for other applications, including whole genome sequencing or RNA-seq. The method described allows for a common processing approach for multiple sample types and accommodates downstream analysis of genomic, metagenomic and transcriptional information. PMID:27168460

  10. Efficient Nucleic Acid Extraction and 16S rRNA Gene Sequencing for Bacterial Community Characterization

    PubMed Central

    Anahtar, Melis N.; Bowman, Brittany A.; Kwon, Douglas S.

    2016-01-01

    There is a growing appreciation for the role of microbial communities as critical modulators of human health and disease. High throughput sequencing technologies have allowed for the rapid and efficient characterization of bacterial communities using 16S rRNA gene sequencing from a variety of sources. Although readily available tools for 16S rRNA sequence analysis have standardized computational workflows, sample processing for DNA extraction remains a continued source of variability across studies. Here we describe an efficient, robust, and cost effective method for extracting nucleic acid from swabs. We also delineate downstream methods for 16S rRNA gene sequencing, including generation of sequencing libraries, data quality control, and sequence analysis. The workflow can accommodate multiple samples types, including stool and swabs collected from a variety of anatomical locations and host species. Additionally, recovered DNA and RNA can be separated and used for other applications, including whole genome sequencing or RNA-seq. The method described allows for a common processing approach for multiple sample types and accommodates downstream analysis of genomic, metagenomic and transcriptional information. PMID:27168460

  11. Identification of the amino acid sequence that targets peroxiredoxin 6 to lysosome-like structures of lung epithelial cells.

    PubMed

    Sorokina, Elena M; Feinstein, Sheldon I; Milovanova, Tatyana N; Fisher, Aron B

    2009-11-01

    Peroxiredoxin 6 (Prdx6), an enzyme with glutathione peroxidase and PLA2 (aiPLA2) activities, is highly expressed in respiratory epithelium, where it participates in phospholipid turnover and antioxidant defense. Prdx6 has been localized by immunocytochemistry and subcellular fractionation to acidic organelles (lung lamellar bodies and lysosomes) and cytosol. On the basis of their pH optima, we have postulated that protein subcellular localization determines the balance between the two activities of Prdx6. Using green fluorescent protein-labeled protein expression in alveolar epithelial cell lines, we showed Prdx6 localization to organellar structures resembling lamellar bodies in mouse lung epithelial (MLE-12) cells and lysosomes in A549 cells. Localization within lamellar bodies/lysosomes was in the luminal compartment. Targeting to lysosome-like organelles was abolished by the deletion of amino acids 31-40 from the Prdx6 NH2-terminal region; deletion of the COOH-terminal region had no effect. A green fluorescent protein-labeled peptide containing only amino acids 31-40 showed lysosomal targeting that was abolished by mutation of S32 or G34 within the peptide. Studies with mutated protein indicated that lipid binding was not necessary for Prdx6 targeting. This peptide sequence has no homology to known organellar targeting motifs. These studies indicate that the localization of Prdx6 in acidic organelles and consequent PLA2 activity depend on a novel 10-aa peptide located at positions 31-40 of the protein. PMID:19700648

  12. Preparation of Nucleic Acid Libraries for Personalized Sequencing Systems Using an Integrated Microfluidic Hub Technology (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

    ScienceCinema

    Patel, Kamlesh D [Ken]; SNL,

    2013-01-25

    Kamlesh (Ken) Patel from Sandia National Laboratories (Livermore, California) presents "Preparation of Nucleic Acid Libraries for Personalized Sequencing Systems Using an Integrated Microfluidic Hub Technology " at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

  13. Cloning and nucleotide sequencing of genes for three small, acid-soluble proteins from Bacillus subtilis spores.

    PubMed Central

    Connors, M J; Mason, J M; Setlow, P

    1986-01-01

    Three Bacillus subtilis genes (termed sspA, sspB, and sspD) which code for small, acid-soluble spore proteins (SASPs) have been cloned, and their complete nucleotide sequence has been determined. The amino acid sequences of the SASPs coded for by these genes are similar to each other and to those of the SASP-1 of B. subtilis (coded for by the sspC gene) and the SASP-A/C family of B. megaterium. The sspA and sspB genes are expressed only in sporulation, in parallel with each other and with the sspC gene. Two regions upstream of the postulated transcription start sites for the sspA and B genes have significant homology with the analogous regions of the sspC gene and the SASP-A/C gene family. Purification of two of the three major B, subtilis SASPs (alpha and beta) and determination of their amino-terminal sequences indicated that the sspA gene codes for SASP-alpha and that the sspB gene codes for SASP-beta. This was confirmed by the introduction of deletion mutations into the cloned sspA and sspB genes and transfer of these deletions into the B. subtilis chromosome with concomitant loss of the wild-type gene. Images PMID:3009398

  14. Deduced amino acid sequence of human pulmonary surfactant proteolipid: SPL(pVal)

    SciTech Connect

    Whitsett, J.A.; Glasser, S.W.; Korfhagen, T.R.; Weaver, T.E.; Clark, J.; Pilot-Matias, T.; Meuth, J.; Fox, J.L.

    1987-05-01

    Hydrophobic, proteolipid-like protein of Mr 6500 was isolated from ether/ethanol extracts of human, canine and bovine pulmonary surfactant. Amino acid composition of the protein demonstrated a remarkable abundance of hydrophobic residues, particularly valine and leucine. The N-terminal amino acid sequence of the human protein was determined: N-Leu-Ile-Pro-Cys-Cys-Pro-Val-Asn-Leu-Lys-Arg-Leu-Leu-Ile-Val4... An oligonucleotide probe was used to screen an adult human lung cDNA library and resulted in detection of cDNA clones with predicted amino acid sequence with close identity to the N-terminal amino acid sequence of the human peptide. SPL(pVal) was found within the reading frame of a larger peptide. SPL(pVal) results from proteolytic processing of a larger preprotein. Northern blot analysis detected in a single 1.0 kilobase SPL(pVal) RNA which was less abundant in fetal than in adult lung. Mixtures of purified canine and bovine SPL(pVal) and synthetic phospholipids display properties of rapid adsorption and surface tension lowering activity characteristic of surfactant. Human SPL(pVal) is a pulmonary surfactant proteolipid which may therefore be useful in combination with phospholipids and/or other surfactant proteins for the treatment of surfactant deficiency such as hyaline membrane disease in newborn infants.

  15. Complete nucleic acid sequence of Penaeus stylirostris densovirus (PstDNV) from India.

    PubMed

    Rai, Praveen; Safeena, Muhammed P; Karunasagar, Iddya; Karunasagar, Indrani

    2011-06-01

    Infectious hypodermal and hematopoietic necrosis virus (IHHNV) of shrimp, recently been classified as Penaeus stylirostris densovirus (PstDNV). The complete nucleic acid sequence of PstDNV from India was obtained by cloning and sequencing of different DNA fragment of the virus. The genome organisation of PstDNV revealed that there were three major coding domains: a left ORF (NS1) of 2001 bp, a mid ORF (NS2) of 1092 bp and a right ORF (VP) of 990 bp. The complete genome and amino acid sequences of three proteins viz., NS1, NS2 and VP were compared with the genomes of the virus reported from Hawaii, China and Mexico and with partial sequence available from isolates from different regions. The phylogenetic analysis of shrimp, insect and vertebrate parvovirus sequences showed that the Indian PstDNV isolate is phylogenetically more closely related to one of the three isolates from Taiwan (AY355307), and two isolates (AY362547 and AY102034) from Thailand. PMID:21402111

  16. Human liver type pyruvate kinase: complete amino acid sequence and the expression in mammalian cells.

    PubMed Central

    Tani, K; Fujii, H; Nagata, S; Miwa, S

    1988-01-01

    Pyruvate kinase (PK) has four isozymes (L, R, M1, M2) that are encoded by two different genes. Among these isozymes, abnormalities of liver (L)-type PK is considered to be associated with hereditary nonspherocytic hemolytic anemia in humans. We isolated and determined the full-length sequence of human L-type PK cDNA. The cDNA contains 1629 base pairs encoding 543 amino acids, 68 base pairs of 5'-noncoding sequence, and 734 base pairs of 3'-noncoding sequence. The similarity between human and rat L-type PK was 86.9% at the nucleotide sequence level and 92.4% at the amino acid sequence level. The full-length L-type PK cDNA was placed under the promoter of simian virus 40 and introduced into monkey COS cells. Human L-type PK activity was detected in the extract of COS cells by the classical PK electrophoresis method. Images PMID:3126495

  17. Human liver type pyruvate kinase: Complete amino acid sequence and the expression in mammalian cells

    SciTech Connect

    Tani, Kenzaburo; Nagata, Shigekazu ); Fujii, Hisaichi ); Miwa, Shiro )

    1988-03-01

    Pyruvate kinase (PK) has four isozymes (L, R, M{sub 1}, M{sub 2}) that are encoded by two different genes. Among these isozymes, abnormalities of liver (L)-type PK is considered to be associated with hereditary nonspherocytic hemolytic anemia in humans. The authors isolated and determined the full-length sequence of human L-type PK cDNA. The cDNA contains 1,629 base pairs encoding 543 amino acids, 68 base pairs of 5{prime}-noncoding sequence, and 734 base pairs of 3{prime}-noncoding sequence. The similarity between human and rat L-type PK was 86.9% at the nucleotide sequence level and 92.4% at the amino acid sequence level. The full-length L-type PK cDNA was placed under the promoter of simian virus 40 and introduced into monkey COS cells. Human L-type PK activity was detected in the extract of COS cells by the classical PK electrophoresis method.

  18. Comparative sequence analyses indicate that Coffea (Asterids) and Vitis (Rosids) derive from the same paleo-hexaploid ancestral genome.

    PubMed

    Cenci, Alberto; Combes, Marie-Christine; Lashermes, Philippe

    2010-05-01

    The complete sequence of Vitis vinifera revealed that the rosid clade derives from a hexaploid ancestor. At present, no analysis of complete genome sequence is available for an asterid, the other large eudicot clade, which includes the economically important species potato, tomato and coffee. To elucidate the genomic history of asterids, we compared the sequence of an 800 kb region of diploid Coffea genome to the orthologous regions of V. vinifera, Populus trichocarpa and Arabidopsis thaliana. We found a very high level of collinearity between around 80 genes of the three rosid species and Coffea. Collinearity comparisons between orthologous and paralogous regions indicates that (1) the Coffea (and consequently all asterids) and rosids share the same hexaploid ancestor; (2) the diploidization process (loss of duplicated and redundant copies from the whole genome duplication) was very advanced in the most recent common ancestor of rosids and asterids. Finally, no additional polyploidization events were detected in the Coffea lineage. Differences in gene loss rates were detected among the three rosid species and linked to the divergence in protein sequences. PMID:20361338

  19. Molecular cytogenetics by polymerase catalyzed amplification or in situ labelling of specific nucleic acid sequences

    SciTech Connect

    Bolund, L.; Brandt, C.; Hindkjaer, J.; Koch, J.; Koelvraa, S.; Pedersen, S. )

    1993-01-01

    The Polymerase Chain Reaction (PCR) can be performed on isolated cells or chromosomes and the product can be analyzed by DNA technology or by FISH to test metaphases. The authors have good experiences analyzing aberrant chromosomes by FACS sorting, PCR with degenerated primers and painting of test metaphases with the PCR product. They also utilize polymerases for PRimed IN Situ labelling (PRINS) of specific nucleic acid sequences. In PRINS oligonucleotides are hybridized to their target sequences and labeled nucleotides are incorporated at the site of hybridization with the oligonucleotide as primer. PRINS may eventually allow the study of individual genes, gene expression and even somatic mutations (in mRNA) in single cells.

  20. DNA Cloning of Plasmodium falciparum Circumsporozoite Gene: Amino Acid Sequence of Repetitive Epitope

    NASA Astrophysics Data System (ADS)

    Enea, Vincenzo; Ellis, Joan; Zavala, Fidel; Arnot, David E.; Asavanich, Achara; Masuda, Aoi; Quakyi, Isabella; Nussenzweig, Ruth S.

    1984-08-01

    A clone of complementary DNA encoding the circumsporozoite (CS) protein of the human malaria parasite Plasmodium falciparum has been isolated by screening an Escherichia coli complementary DNA library with a monoclonal antibody to the CS protein. The DNA sequence of the complementary DNA insert encodes a four-amino acid sequence: proline-asparagine-alanine-asparagine, tandemly repeated 23 times. The CS β -lactamase fusion protein specifically binds monoclonal antibodies to the CS protein and inhibits the binding of these antibodies to native Plasmodium falciparum CS protein. These findings provide a basis for the development of a vaccine against Plasmodium falciparum malaria.

  1. Method for high-volume sequencing of nucleic acids: random and directed priming with libraries of oligonucleotides

    DOEpatents

    Studier, F.W.

    1995-04-18

    Random and directed priming methods for determining nucleotide sequences by enzymatic sequencing techniques, using libraries of primers of lengths 8, 9 or 10 bases, are disclosed. These methods permit direct sequencing of nucleic acids as large as 45,000 base pairs or larger without the necessity for subcloning. Individual primers are used repeatedly to prime sequence reactions in many different nucleic acid molecules. Libraries containing as few as 10,000 octamers, 14,200 nonamers, or 44,000 decamers would have the capacity to determine the sequence of almost any cosmid DNA. Random priming with a fixed set of primers from a smaller library can also be used to initiate the sequencing of individual nucleic acid molecules, with the sequence being completed by directed priming with primers from the library. In contrast to random cloning techniques, a combined random and directed priming strategy is far more efficient. 2 figs.

  2. Method for high-volume sequencing of nucleic acids: random and directed priming with libraries of oligonucleotides

    DOEpatents

    Studier, F. William

    1995-04-18

    Random and directed priming methods for determining nucleotide sequences by enzymatic sequencing techniques, using libraries of primers of lengths 8, 9 or 10 bases, are disclosed. These methods permit direct sequencing of nucleic acids as large as 45,000 base pairs or larger without the necessity for subcloning. Individual primers are used repeatedly to prime sequence reactions in many different nucleic acid molecules. Libraries containing as few as 10,000 octamers, 14,200 nonamers, or 44,000 decamers would have the capacity to determine the sequence of almost any cosmid DNA. Random priming with a fixed set of primers from a smaller library can also be used to initiate the sequencing of individual nucleic acid molecules, with the sequence being completed by directed priming with primers from the library. In contrast to random cloning techniques, a combined random and directed priming strategy is far more efficient.

  3. Partial amino acid sequence of apolipoprotein(a) shows that it is homologous to plasminogen

    SciTech Connect

    Eaton, D.L.; Fless, G.M.; Kohr, W.J.; McLean, J.W.; Xu, Q.T.; Miller, C.G.; Lawn, R.M.; Scanu, A.M.

    1987-05-01

    Apolipoprotein(a) (apo(a)) is a glycoprotein with M/sub r/ approx. 280,000 that is disulfide linked to apolipoprotein B in lipoprotein(a) particles. Elevated plasma levels of lipoprotein(a) are correlated with atherosclerosis. Partial amino acid sequence of apo(a) shows that it has striking homology to plasminogen. Plasminogen is a plasma serine protease zymogen that consists of five homologous and tandemly repeated domains called kringles and a trypsin-like protease domain. The amino-terminal sequence obtained for apo(a) is homologous to the beginning of kringle 4 but not the amino terminus of plasminogen. Apo(a) was subjected to limited proteolysis by trypsin or V8 protease, and fragments generated were isolated and sequenced. Sequences obtained from several of these fragments are highly (77-100%) homologous to plasminogen residues 391-421, which reside within kringle 4. Analysis of these internal apo(a) sequences revealed that apo(a) may contain at least two kringle 4-like domains. A sequence obtained from another tryptic fragment also shows homology to the end of kringle 4 and the beginning of kringle 5. Sequence data obtained from the two tryptic fragments shows homology with the protease domain of plasminogen. One of these sequences is homologous to the sequences surrounding the activation site of plasminogen. Plasminogen is activated by the cleavage of a specific arginine residue by urokinase and tissue plasminogen activator; however, the corresponding site in apo(a) is a serine that would not be cleaved by tissue plasminogen activator or urokinase. Using a plasmin-specific assay, no proteolytic activity could be demonstrated for lipoprotein(a) particles. These results suggest that apo(a) contains kringle-like domains and an inactive protease domain.

  4. The sequence diversity and expression among genes of the folic acid biosynthesis pathway in industrial Saccharomyces strains.

    PubMed

    Goncerzewicz, Anna; Misiewicz, Anna

    2015-01-01

    Folic acid is an important vitamin in human nutrition and its deficiency in pregnant women's diets results in neural tube defects and other neurological damage to the fetus. Additionally, DNA synthesis, cell division and intestinal absorption are inhibited in case of adults. Since this discovery, governments and health organizations worldwide have made recommendations concerning folic acid supplementation of food for women planning to become pregnant. In many countries this has led to the introduction of fortifications, where synthetic folic acid is added to flour. It is known that Saccharomyces strains (brewing and bakers' yeast) are one of the main producers of folic acid and they can be used as a natural source of this vitamin. Proper selection of the most efficient strains may enhance the folate content in bread, fermented vegetables, dairy products and beer by 100% and may be used in the food industry. The objective of this study was to select the optimal producing yeast strain by determining the differences in nucleotide sequences in the FOL2, FOL3 and DFR1 genes of folic acid biosynthesis pathway. The Multitemperature Single Strand Conformation Polymorphism (MSSCP) method and further nucleotide sequencing for selected strains were applied to indicate SNPs in selected gene fragments. The RT qPCR technique was also applied to examine relative expression of the FOL3 gene. Furthermore, this is the first time ever that industrial yeast strains were analysed regarding genes of the folic acid biosynthesis pathway. It was observed that a correlation exists between the folic acid amount produced by industrial yeast strains and changes in the nucleotide sequence of adequate genes. The most significant changes occur in the DFR1 gene, mostly in the first part, which causes major protein structure modifications in KKP 232, KKP 222 and KKP 277 strains. Our study shows that the large amount of SNP contributes to impairment of the selected enzymes and S. cerevisiae and S

  5. Characterising humic acid fouling of nanofiltration membranes using bisphenol A as a molecular indicator.

    PubMed

    Nghiem, Long D; Vogel, Dirk; Khan, Stuart

    2008-09-01

    Fouling of nanofiltration (NF) membranes by humic acids was investigated using bisphenol A (BPA) as an indicator chemical to differentiate between various mechanisms that may lead to a change in solute rejection. Three commercially available NF membranes were investigated and an accelerated fouling condition was achieved with a foulant mixture containing humic acids in an electrolyte matrix. The effects of membrane fouling on the rejection of BPA were interpreted with respect to the membrane pore sizes and the fouling characteristics. Results reported here indicate that calcium concentration in the feed solution could be a major factor governing the humic acid fouling process. Moreover, a critical concentration of calcium in the feed solution was observed, at which membrane fouling was most severe. Membrane fouling characteristics were observed by their influence on BPA rejection. Such influence could result in either an increase or decrease in rejection of BPA by the three different membranes depending on the rejection mechanisms involved. It is hypothesized that these mechanisms could occur simultaneously and that the effects of each might not be easily distinguished. However, it was observed that their relative contribution was largely dependent upon membrane pore size. Pore blocking, which resulted in a considerable improvement in rejection, was prominent for the more open pore size TFC-SR2 membrane. In contrast, the cake-enhanced concentration polarisation effect was more severe for the tighter NF-270 and NF-90 membranes. For hydrophobic solutes such as BPA, the formation of the fouling layer could also interfere with the solute-membrane interaction, and therefore, exert considerable influence on the separation process. PMID:18678386

  6. Acidic deposition, cation mobilization, and biochemical indicators of stress in healthy red spruce

    USGS Publications Warehouse

    Shortle, W.C.; Smith, K.T.; Minocha, R.; Lawrence, G.B.; David, M.B.

    1997-01-01

    Dendrochemical and biochemical markers link stress in apparently healthy red spruce trees (Picea rubens) to acidic deposition. Acidic deposition to spruce forests of the northeastern USA increased sharply during the 1960s. Previous reports related visible damage of trees at high elevations to root and soil processes. In this report, dendrochemical and foliar biochemical markers indicate perturbations in biological processes in healthy red spruce trees across the northeastern USA. Previous research on the dendrochemistry of red spruce stemwood indicated that under uniform environmental conditions, stemwood concentrations of Ca and Mg decreased with increasing radial distance from the pith. For nine forest locations, frequency analysis shows that 28 and 52% of samples of red spruce stemwood formed in the 1960s are enriched in Ca and Mg, respectively, relative to wood formed prior to and after the 1960s. This enrichment in trees throughout the northeastern USA may be interpretable as a signal of increased availability of essential cations in forest soils. Such a temporary increase in the availability of Ca and Mg could be caused by cation mobilization, a consequence of increased acidic deposition. During cation mobilization, essential Ca and Mg as well as potentially harmful Al become more available for interaction with binding sites in the soil and absorbing roots. As conditions which favor cation mobilization continue, Ca and Mg can be leached or displaced from the soil. A measure of the interaction between Ca and Al is the Al/Ca binding ratio (molar charge ratio of exchangeable Al to exchangeable Ca). As the Al/Ca binding ratio in the root zone increased from 0.3 to 1.9, the foliar concentration of the biochemical stress marker putrescine also increased from 45 to 145 nm g-1. The correlation of the putrescine concentration to the Al/Ca binding ratio (adj. r2 = 0.68, P < 0.027) suggests that foliar stress may be linked to soil chemistry.

  7. Amino acids labelled with 11C as indicator of the effect of dietary treatment of hyperammonaemia.

    PubMed

    Hardell, L I; Stålnacke, C G; Lundqvist, H; Malmborg, P; Långström, B

    1984-01-01

    Short-lived radioactive carbon, 11C, (T 1/2 = 20 min) was incorporated into an essential amino acid [11C-methyl] -L-methionine, to form a true biological amino acid tracer with external detectability. This was tested in a study of the physiological tracer dynamics in a hyperammonaemic patient before and after a change in the dietary treatment. The protein intake was unchanged between the two investigations but the energy intake was increased from 53 to 63 kcal/kg BW/day. The tracer radioactivity was given per os. In the second investigation a relative decrease of radioactivity in the low molecular weight fraction of blood plasma was seen. Also the external measurements indicated a higher hepatic retention of radioactivity in the second investigation but no increased excretion of tracer. This may reflect an increased ability of the liver to utilize the incoming methionine from the vena porta. The hyperammonaemia remained over the second investigation but seven months later the ammonia content in the blood was almost normalized and the patient had also gained 3 kg in weight. The correlation between changes in tracer dynamics and changes in therapeutical effect of the diet is not further verified in this experiment but the investigation indicates the value of further studies in this topic using 11C-labelled amino acids also including the use of the newly introduced positron tomographic technique. It may be possible to develop this type of nuclide technique further to achieve a clinically useful method of optimizing therapeutic regiments in this type of metabolic disease. PMID:6393522

  8. Using Chou's pseudo amino acid composition to predict protein quaternary structure: a sequence-segmented PseAAC approach.

    PubMed

    Zhang, Shao-Wu; Chen, Wei; Yang, Feng; Pan, Quan

    2008-10-01

    In the protein universe, many proteins are composed of two or more polypeptide chains, generally referred to as subunits, which associate through noncovalent interactions and, occasionally, disulfide bonds to form protein quaternary structures. It has long been known that the functions of proteins are closely related to their quaternary structures; some examples include enzymes, hemoglobin, DNA polymerase, and ion channels. However, it is extremely labor-expensive and even impossible to quickly determine the structures of hundreds of thousands of protein sequences solely from experiments. Since the number of protein sequences entering databanks is increasing rapidly, it is highly desirable to develop computational methods for classifying the quaternary structures of proteins from their primary sequences. Since the concept of Chou's pseudo amino acid composition (PseAAC) was introduced, a variety of approaches, such as residue conservation scores, von Neumann entropy, multiscale energy, autocorrelation function, moment descriptors, and cellular automata, have been utilized to formulate the PseAAC for predicting different attributes of proteins. Here, in a different approach, a sequence-segmented PseAAC is introduced to represent protein samples. Meanwhile, multiclass SVM classifier modules were adopted to classify protein quaternary structures. As a demonstration, the dataset constructed by Chou and Cai [(2003) Proteins 53:282-289] was adopted as a benchmark dataset. The overall jackknife success rates thus obtained were 88.2-89.1%, indicating that the new approach is quite promising for predicting protein quaternary structure. PMID:18427713

  9. Self-sequencing of amino acids and origins of polyfunctional protocells.

    PubMed

    Fox, S W

    1984-01-01

    The primal role of the origins of proteins in molecular evolution is discussed. On the basis of this premise, the significance of the experimentally established self-sequencing of amino acids under simulated geological conditions is explained as due to the fact that the products are highly nonrandom and accordingly contain many kinds of information. When such thermal proteins are aggregated into laboratory protocells, an action that occurs readily, the resultant protocells also contain many kinds of information. Residue-by-residue order, enzymic activities, and lipid quality accordingly occur within each preparation of proteinoid (thermal protein). In this paper are reviewed briefly the phenomenon of self-sequencing of amino acids, its relationship to evolutionary processes, other significance of such self-ordering, and the experimental evidence for original polyfunctional protocells. PMID:6462684

  10. Self-Sequencing of Amino Acids and Origins of Polyfunctional Protocells

    NASA Astrophysics Data System (ADS)

    Fox, Sidney W.

    1984-12-01

    The primal role of the origins of proteins in molecular evolution is discussed. On the basis of this premise, the significance of the experimentally established self-sequencing of amino acids under simulated geological conditions is explained as due to the fact that the products are highly nonrandom and accordingly contain many kinds of information. When such thermal proteins are aggregated into laboratory protocells, an action that occurs readily, the resultant protocells also contain many kinds of information. Residue-by-residue order, enzymic activities, and lipid quality accordingly occur within each preparation of proteinoid (thermal protein). In this paper are reviewed briefly the phenomenon of self-sequencing of amino acids, its relationship to evolutionary processes, other significance of such self-ordering, and the experimental evidence for original polyfunctional protocells.

  11. Sequence of morphological transitions in two-dimensional pattern growth from aqueous ascorbic Acid solutions.

    PubMed

    Paranjpe, A S

    2002-08-12

    A sequence of morphological transitions in two-dimensional dehydration patterns of aqueous solutions of ascorbic acid is observed with humidity as a control parameter. Change in morphology occurs due to humidity induced variation in the concentration of the metastable supersaturated solution phase formed after initial solvent evaporation. As percent humidity is varied from 40 to 80, patterns change from compact circular --> radial --> density modulated radial (a new morphology) --> density modulated circular --> density modulated dendritic (a new morphology) --> dense branching. PMID:12190528

  12. Self-sequencing of amino acids and origins of polyfunctional protocells

    NASA Technical Reports Server (NTRS)

    Fox, S. W.

    1984-01-01

    The role of proteins in the origin of living things is discussed. It has been experimentally established that amino acids can sequence themselves under simulated geological conditions with highly nonrandom products which accordingly contain diverse information. Multiple copies of each type of macromolecule are formed, resulting in greater power for any protoenzymic molecule than would accrue from a single copy of each type. Thermal proteins are readily incorporated into laboratory protocells. The experimental evidence for original polyfunctional protocells is discussed.

  13. Snake venom. The amino acid sequence of protein A from Dendroaspis polylepis polylepis (black mamba) venom.

    PubMed

    Joubert, F J; Strydom, D J

    1980-12-01

    Protein A from Dendroaspis polylepis polylepis venom comprises 81 amino acids, including ten half-cystine residues. The complete primary structures of protein A and its variant A' were elucidated. The sequences of proteins A and A', which differ in a single position, show no homology with various neurotoxins and non-neurotoxic proteins and represent a new type of elapid venom protein. PMID:7461607

  14. Pedogenesis and paleoenvironmental records in the tephra-paleosol sequences of Central Mexico: micromorphometric indicators.

    NASA Astrophysics Data System (ADS)

    Diaz, Jaime; Solleiro, Elizabeth; Sedov, Sergey

    2014-05-01

    tepetates (hardened volcanic soils) in Mexico have been studied from an agronomic and mechanisms viewpoint, however due to their different origins have not been established mechanisms that link them to environmental studies. This paper uses morphometric tools to determine the origin of the Tepetates in Mexico, using them as indicators of environmental changes and periods of instability of the landscape. The Glacis de Buenavista is located mainly in the northwestern portion of the state of Morelos, between the parallels 18° 20'and 19° 08' north latitude and meridians 98° 37' and 99° 30' west, forming a piedmont relief caused by erosion-sedimentation phases of volcanic materials from the Sierra de Zempoala mainly, and that is a big fan of Pleistocene age. Covers an area of 202.7 km 2. The maximum altitude is in the north, 2600 m, and minimun in the south to the 970 m, with an average slope of 6 . The Glacis de Buenavista has soils cover which consists of Luvisols, Vertisols, and layers of Tepetates, Phaeozems and Luvisols, in particular, represent soils polycyclic polygenetic high degree of development that have been formed in the Holocene. The next question is what mechanisms control the presence of a floor or other unit that occurs in the area? Since the lateral boundaries between soil types are abrupt, which speaks of discontinuities in the processes that have given rise. The morphometric study was carried out in thins sections of the blocks of Tepetates undisturbed. The blocks were vacuum impregnated with polyester resin. The thins sections were scanned with an Epson scanner 7100 with high resolution 9600 dpi and 24 bit color depth and analyzed using Image Pro Plus 5.1 program. The stratigraphic control was performed by 14C dating of organic matter. The pedofeatures measured were roundness of soil fragments, matrix percentage, percentage of papules and porosity. Was also carried out a microscopic control using a microscope BX51 Olimpus pedofeatures observing

  15. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... approved by the Director of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 51... base or modified or unusual amino acid may be presented in a given sequence as the corresponding unmodified base or amino acid if the modified base or modified or unusual amino acid is one of those...

  16. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... approved by the Director of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 51... base or modified or unusual amino acid may be presented in a given sequence as the corresponding unmodified base or amino acid if the modified base or modified or unusual amino acid is one of those...

  17. Nanopore Analysis of Nucleic Acids: Single-Molecule Studies of Molecular Dynamics, Structure, and Base Sequence

    NASA Astrophysics Data System (ADS)

    Olasagasti, Felix; Deamer, David W.

    Nucleic acids are linear polynucleotides in which each base is covalently linked to a pentose sugar and a phosphate group carrying a negative charge. If a pore having roughly the crosssectional diameter of a single-stranded nucleic acid is embedded in a thin membrane and a voltage of 100 mV or more is applied, individual nucleic acids in solution can be captured by the electrical field in the pore and translocated through by single-molecule electrophoresis. The dimensions of the pore cannot accommodate anything larger than a single strand, so each base in the molecule passes through the pore in strict linear sequence. The nucleic acid strand occupies a large fraction of the pore's volume during translocation and therefore produces a transient blockade of the ionic current created by the applied voltage. If it could be demonstrated that each nucleotide in the polymer produced a characteristic modulation of the ionic current during its passage through the nanopore, the sequence of current modulations would reflect the sequence of bases in the polymer. According to this basic concept, nanopores are analogous to a Coulter counter that detects nanoscopic molecules rather than microscopic [1,2]. However, the advantage of nanopores is that individual macromolecules can be characterized because different chemical and physical properties affect their passage through the pore. Because macromolecules can be captured in the pore as well as translocated, the nanopore can be used to detect individual functional complexes that form between a nucleic acid and an enzyme. No other technique has this capability.

  18. Purification, amino acid sequence and mode of action of bifidocin B produced by Bifidobacterium bifidum NCFB 1454.

    PubMed

    Yildirim, Z; Winters, D K; Johnson, M G

    1999-01-01

    Bifidocin B produced by Bifidobacterium bifidum NCFB 1454 was purified to homogeneity by a rapid and simple three step purification procedure which included freeze drying, Micro-Cel adsorption/desorption and cation exchange chromatography. The purification resulted in 18% recovery and an approximately 1900-fold increase in the specific activity and purity of bifidocin B. Treatment with bifidocin B caused sensitive cells to lose high amounts of intracellular K+ ions and u.v.-absorbing materials, and to become more permeable to ONPG. Bifidocin B adsorbed to the Gram-positive bacteria but not the Gram-negative bacteria tested. Its adsorption was pH-dependent but not time-dependent. For sensitive cells, the adsorption and lethal action of bifidocin B was very rapid. In 5 min, 95% of bifidocin B adsorbed onto sensitive cells. Several salts inhibited the binding of bifidocin B, which could be overcome by increasing the amount of bifidocin B added. Pre-treatment of sensitive cells and cell walls with detergents, organic solvents or enzymes did not cause a reduction in subsequent cellular binding of bifidocin B, but cell wall preparations treated with methanol:chloroform and hot 20% (w/v) TCA lost the ability to adsorb bifidocin B. Also, the addition of purified heterologous lipoteichoic acid to sensitive cells completely blocked the adsorption of bifidocin B. The amino acid sequence indicated that the bacteriocin contained 36 residues. N-terminal amino acid sequence analysis yielded a sequence of KYYGNGVTCGLHDCRVDRGKATCGIINNGGMWGDIG. Curing experiments with 20 micrograms ml-1 acriflavine yielded cell derivatives that no longer produced bifidocin B but retained immunity to bifidocin B. Production of bifidocin B, but not immunity to bifidocin B, was associated with a plasmid of about 8 kb in this strain. PMID:10030011

  19. Complete amino acid sequence of a histidine-rich proteolytic fragment of human ceruloplasmin.

    PubMed

    Kingston, I B; Kingston, B L; Putnam, F W

    1979-04-01

    The complete amino acid sequence has been determined for a fragment of human ceruloplasmin [ferroxidase; iron(II):oxygen oxidoreductase, EC 1.16.3.1]. The fragment (designated Cp F5) contains 159 amino acid residues and has a molecular weight of 18,650; it lacks carbohydrate, is rich in histidine, and contains one free cysteine that may be part of a copper-binding site. This fragment is present in most commercial preparations of ceruloplasmin, probably owing to proteolytic degradation, but can also be obtained by limited cleavage of single-chain ceruloplasmin with plasmin. Cp F5 probably is an intact domain attached to the COOH-terminal end of single-chain ceruloplasmin via a labile interdomain peptide bond. A model of the secondary structure predicted by empirical methods suggests that almost one-third of the amino acid residues are distributed in alpha helices, about a third in beta-sheet structure, and the remainder in beta turns and unidentified structures. Computer analysis of the amino acid sequence has not demonstrated a statistically significant relationship between this ceruloplasmin fragment and any other protein, but there is some evidence for an internal duplication. PMID:287005

  20. The amino acid sequence of Lady Amherst's pheasant (Chrysolophus amherstiae) and golden pheasant (Chrysolophus pictus) egg-white lysozymes.

    PubMed

    Araki, T; Kuramoto, M; Torikata, T

    1990-09-01

    The amino acids of Lady Amherst's pheasant and golden pheasant egg-white lysozymes have been sequenced. The carboxymethylated lysozymes were digested with trypsin followed by sequencing of the tryptic peptides. Lady Amherst's pheasant lysozyme proved to consist of 129 amino acid residues, and a relative molecular mass of 14,423 Da was calculated. This lysozyme had 6 amino acids substitutions when compared with hen egg-white lysozyme: Phe3 to Tyr, His15 to Leu, Gln41 to His, Asn77 to His, Gln 121 to Asn, and a newly found substitution of Ile124 to Thr. The amino acid sequence of golden pheasant lysozyme was identical to that of Lady Amherst's phesant lysozyme. The phylogenetic tree constructured by the comparison of amino acid sequences of phasianoid birds lysozymes revealed a minimum genetic distance between these pheasants and the turkey-peafowl group. PMID:1368578

  1. Complete Genome Sequence of a thermotolerant sporogenic lactic acid bacterium, Bacillus coagulans strain 36D1

    PubMed Central

    Rhee, Mun Su; Moritz, Brélan E.; Xie, Gary; Glavina del Rio, T.; Dalin, E.; Tice, H.; Bruce, D.; Goodwin, L.; Chertkov, O.; Brettin, T.; Han, C.; Detter, C.; Pitluck, S.; Land, Miriam L.; Patel, Milind; Ou, Mark; Harbrucker, Roberta; Ingram, Lonnie O.; Shanmugam, K. T.

    2011-01-01

    Bacillus coagulans is a ubiquitous soil bacterium that grows at 50-55 °C and pH 5.0 and ferments various sugars that constitute plant biomass to L (+)-lactic acid. The ability of this sporogenic lactic acid bacterium to grow at 50-55 °C and pH 5.0 makes this organism an attractive microbial biocatalyst for production of optically pure lactic acid at industrial scale not only from glucose derived from cellulose but also from xylose, a major constituent of hemicellulose. This bacterium is also considered as a potential probiotic. Complete genome sequence of a representative strain, B. coagulans strain 36D1, is presented and discussed. PMID:22675583

  2. Complete amino acid sequence of globin chains and biological activity of fragmented crocodile hemoglobin (Crocodylus siamensis).

    PubMed

    Srihongthong, Saowaluck; Pakdeesuwan, Anawat; Daduang, Sakda; Araki, Tomohiro; Dhiravisit, Apisak; Thammasirirak, Sompong

    2012-08-01

    Hemoglobin, α-chain, β-chain and fragmented hemoglobin of Crocodylus siamensis demonstrated both antibacterial and antioxidant activities. Antibacterial and antioxidant properties of the hemoglobin did not depend on the heme structure but could result from the compositions of amino acid residues and structures present in their primary structure. Furthermore, thirteen purified active peptides were obtained by RP-HPLC analyses, corresponding to fragments in the α-globin chain and the β-globin chain which are mostly located at the N-terminal and C-terminal parts. These active peptides operate on the bacterial cell membrane. The globin chains of Crocodylus siamensis showed similar amino acids to the sequences of Crocodylus niloticus. The novel amino acid substitutions of α-chain and β-chain are not associated with the heme binding site or the bicarbonate ion binding site, but could be important through their interactions with membranes of bacteria. PMID:22648692

  3. [Partial sequence homology of FtsZ in phylogenetics analysis of lactic acid bacteria].

    PubMed

    Zhang, Bin; Dong, Xiu-zhu

    2005-10-01

    FtsZ is a structurally conserved protein, which is universal among the prokaryotes. It plays a key role in prokaryote cell division. A partial fragment of the ftsZ gene about 800bp in length was amplified and sequenced and a partial FtsZ protein phylogenetic tree for the lactic acid bacteria was constructed. By comparing the FtsZ phylogenetic tree with the 16S rDNA tree, it was shown that the two trees were similar in topology. Both trees revealed that Pediococcus spp. were closely related with L. casei group of Lactobacillus spp. , but less related with other lactic acid cocci such as Enterococcus and Streptococcus. The results also showed that the discriminative power of FtsZ was higher than that of 16S rDNA for either inter-species or inter-genus and could be a very useful tool in species identification of lactic acid bacteria. PMID:16342751

  4. Early indications of soil recovery from acidic deposition in U.S. red spruce forests

    USGS Publications Warehouse

    Lawrence, Gregory B.; Shortle, Walter C.; David, Mark B.; Smith, Kevin T.; Warby, Richard A.F.; Lapenis, Andrei G.

    2012-01-01

    Forty to fifty percent decreases in acidic deposition through the 1980s and 1990s led to partial recovery of acidified surface waters in the northeastern United States; however, the limited number of studies that have assessed soil change found increased soil acidification during this period. From existing data, it's not clear whether soils continued to worsen in the 1990s or if recovery had begun. To evaluate possible changes in soils through the 1990s, soils in six red spruce (Picea rubens Sarg.) stands in New York, Vermont, New Hampshire, and Maine, first sampled in 1992 to 1993, were resampled in 2003 to 2004. The Oa-horizon pH increased (P 42−, which decreased the mobility of Al throughout the upper soil profile. Results indicate a nascent recovery driven largely by vegetation processes.

  5. Identification of genes and pathways involved in the synthesis of Mead acid (20:3n-9), an indicator of essential fatty acid deficiency.

    PubMed

    Ichi, Ikuyo; Kono, Nozomu; Arita, Yuka; Haga, Shizuka; Arisawa, Kotoko; Yamano, Misato; Nagase, Mana; Fujiwara, Yoko; Arai, Hiroyuki

    2014-01-01

    In mammals, 5,8,11-eicosatrienoic acid (Mead acid, 20:3n-9) is synthesized from oleic acid during a state of essential fatty acid deficiency (EFAD). Mead acid is thought to be produced by the same enzymes that synthesize arachidonic acid and eicosapentaenoic acid, but the genes and the pathways involved in the conversion of oleic acid to Mead acid have not been fully elucidated. The levels of polyunsaturated fatty acids in cultured cells are generally very low compared to those in mammalian tissues. In this study, we found that cultured cells, such as NIH3T3 and Hepa1-6 cells, have significant levels of Mead acid, indicating that cells in culture are in an EFAD state under normal culture conditions. We then examined the effect of siRNA-mediated knockdown of fatty acid desaturases and elongases on the level of Mead acid, and found that knockdown of Elovl5, Fads1, or Fads2 decreased the level of Mead acid. This and the measured levels of possible intermediate products for the synthesis of Mead acid such as 18:2n-9, 20:1n-9 and 20:2n-9 in the knocked down cells indicate two pathways for the synthesis of Mead acid: pathway 1) 18:1n-9→(Fads2)→18:2n-9→(Elovl5)→20:2n-9→(Fads1)→20:3n-9 and pathway 2) 18:1n-9→(Elovl5)→20:1n-9→(Fads2)→20:2n-9→(Fads1)→20:3n-9. PMID:24184513

  6. Acidic deposition, cation mobilization, and biochemical indicators of stress in healthy red spruce

    SciTech Connect

    Shortle, W.C.; Smith, K.T.; Minocha, R.

    1997-05-01

    Dendrochemical and biochemical markers link stress in apparently healthy red spruce trees (Picea rubens) to acidic deposition. Previous reports related visible damage of trees at high elevations to root and soil processes. In this report, dendrochemical and foliar biochemical markers indicate perturbations in biological processes in healthy red spruce trees across the northeastern USA. Previous research on the dendrochemistry of red spruce stemwood indicated that under uniform environmental conditions, stemwood concentrations of Ca and Mg decreased with increasing radial distance from the pith. For nine forest locations, frequency analysis shows that 28 and 52% of samples of red spruce stemwood formed in the 1960s are enriched in Ca and Mg, respectively, relative to wood formed prior to and after the 1960s. This enrichment in trees throughout the northeastern USA may be interpretable as a signal of increased availability of essential cations in forest soils. Such a temporary increase in the availability of Ca and Mg could be caused by cation mobilization, a consequence of increased acidic deposition. During cation mobilization, essential and Ca and Mg as well as potentially harmful Al become more available for interaction with binding sites in the soil and absorbing roots. As conditions which favor cation mobilization continue, Ca and Mg can be leached or displaced from the soil. A measure of the interaction between Ca and Al is the Al/Ca binding ratio (molar charge ratio of exchangeable Al to exchangeable Ca). As the Al/Ca binding ratio in the root zone increased from 0.3 to 1.9, the foliar concentration of the biochemical stress marker putrescine also increased form 45 to 145 nm g{sup {minus}1}. The correlation of the putrescine concentration to the Al/Ca binding ratio (adj. r{sup 2} = 0.68, P <0.027) suggests that foliar stress may be linked to soil chemistry. 32 refs., 2 figs., 1 tab.

  7. Comparative characterization of random-sequence proteins consisting of 5, 12, and 20 kinds of amino acids.

    PubMed

    Tanaka, Junko; Doi, Nobuhide; Takashima, Hideaki; Yanagawa, Hiroshi

    2010-04-01

    Screening of functional proteins from a random-sequence library has been used to evolve novel proteins in the field of evolutionary protein engineering. However, random-sequence proteins consisting of the 20 natural amino acids tend to aggregate, and the occurrence rate of functional proteins in a random-sequence library is low. From the viewpoint of the origin of life, it has been proposed that primordial proteins consisted of a limited set of amino acids that could have been abundantly formed early during chemical evolution. We have previously found that members of a random-sequence protein library constructed with five primitive amino acids show high solubility (Doi et al., Protein Eng Des Sel 2005;18:279-284). Although such a library is expected to be appropriate for finding functional proteins, the functionality may be limited, because they have no positively charged amino acid. Here, we constructed three libraries of 120-amino acid, random-sequence proteins using alphabets of 5, 12, and 20 amino acids by preselection using mRNA display (to eliminate sequences containing stop codons and frameshifts) and characterized and compared the structural properties of random-sequence proteins arbitrarily chosen from these libraries. We found that random-sequence proteins constructed with the 12-member alphabet (including five primitive amino acids and positively charged amino acids) have higher solubility than those constructed with the 20-member alphabet, though other biophysical properties are very similar in the two libraries. Thus, a library of moderate complexity constructed from 12 amino acids may be a more appropriate resource for functional screening than one constructed from 20 amino acids. PMID:20162614

  8. N-Terminal Amino Acid Sequence Determination of Proteins by N-Terminal Dimethyl Labeling: Pitfalls and Advantages When Compared with Edman Degradation Sequence Analysis.

    PubMed

    Chang, Elizabeth; Pourmal, Sergei; Zhou, Chun; Kumar, Rupesh; Teplova, Marianna; Pavletich, Nikola P; Marians, Kenneth J; Erdjument-Bromage, Hediye

    2016-07-01

    In recent history, alternative approaches to Edman sequencing have been investigated, and to this end, the Association of Biomolecular Resource Facilities (ABRF) Protein Sequencing Research Group (PSRG) initiated studies in 2014 and 2015, looking into bottom-up and top-down N-terminal (Nt) dimethyl derivatization of standard quantities of intact proteins with the aim to determine Nt sequence information. We have expanded this initiative and used low picomole amounts of myoglobin to determine the efficiency of Nt-dimethylation. Application of this approach on protein domains, generated by limited proteolysis of overexpressed proteins, confirms that it is a universal labeling technique and is very sensitive when compared with Edman sequencing. Finally, we compared Edman sequencing and Nt-dimethylation of the same polypeptide fragments; results confirm that there is agreement in the identity of the Nt amino acid sequence between these 2 methods. PMID:27006647

  9. N-Terminal Amino Acid Sequence Determination of Proteins by N-Terminal Dimethyl Labeling: Pitfalls and Advantages When Compared with Edman Degradation Sequence Analysis

    PubMed Central

    Chang, Elizabeth; Pourmal, Sergei; Zhou, Chun; Kumar, Rupesh; Teplova, Marianna; Pavletich, Nikola P.; Marians, Kenneth J.

    2016-01-01

    In recent history, alternative approaches to Edman sequencing have been investigated, and to this end, the Association of Biomolecular Resource Facilities (ABRF) Protein Sequencing Research Group (PSRG) initiated studies in 2014 and 2015, looking into bottom-up and top-down N-terminal (Nt) dimethyl derivatization of standard quantities of intact proteins with the aim to determine Nt sequence information. We have expanded this initiative and used low picomole amounts of myoglobin to determine the efficiency of Nt-dimethylation. Application of this approach on protein domains, generated by limited proteolysis of overexpressed proteins, confirms that it is a universal labeling technique and is very sensitive when compared with Edman sequencing. Finally, we compared Edman sequencing and Nt-dimethylation of the same polypeptide fragments; results confirm that there is agreement in the identity of the Nt amino acid sequence between these 2 methods. PMID:27006647

  10. Partial amino acid sequence of fructose-1,6-bisphosphatase from the blue-green algae Synechococcus leopoliensis.

    PubMed

    Marcus, F; Latshaw, S P; Steup, M; Gerbling, K P

    1989-08-01

    Purified fructose-1,6-bisphosphatase from the cyanobacterium Synechococcus leopoliensis was S-carboxymethylated and cleaved with trypsin. The resulting peptides were purified by reversed-phase high performance liquid chromatography and the amino acid sequence of six of the purified peptides was determined by gas-phase microsequencing. The results revealed sequence homology with other fructose-1,6-bisphosphatases. The obtained sequence data provides information required for the design of oligonucleotide hybridization probes to screen existing libraries of cyanobacterial DNA. The determination of the amino acid sequence of cyanobacterial proteins may yield important information with respect to the endosymbiotic theory of evolution. PMID:2550924

  11. Protein sequence analysis by incorporating modified chaos game and physicochemical properties into Chou's general pseudo amino acid composition.

    PubMed

    Xu, Chunrui; Sun, Dandan; Liu, Shenghui; Zhang, Yusen

    2016-10-01

    In this contribution we introduced a novel graphical method to compare protein sequences. By mapping a protein sequence into 3D space based on codons and physicochemical properties of 20 amino acids, we are able to get a unique P-vector from the 3D curve. This approach is consistent with wobble theory of amino acids. We compute the distance between sequences by their P-vectors to measure similarities/dissimilarities among protein sequences. Finally, we use our method to analyze four datasets and get better results compared with previous approaches. PMID:27375218

  12. Amino acid sequences of peptides from a tryptic digest of a urea-soluble protein fraction (U.S.3) from oxidized wool

    PubMed Central

    Corfield, M. C.; Fletcher, J. C.; Robson, A.

    1967-01-01

    1. A tryptic digest of the protein fraction U.S.3 from oxidized wool has been separated into 32 peptide fractions by cation-exchange resin chromatography. 2. Most of these fractions have been resolved into their component peptides by a combination of the techniques of cation-exchange resin chromatography, paper chromatography and paper electrophoresis. 3. The amino acid compositions of 58 of the peptides in the digest present in the largest amounts have been determined. 4. The amino acid sequences of 38 of these have been completely elucidated and those of six others partially derived. 5. These findings indicate that the parent protein in wool from which the protein fraction U.S.3 is derived has a minimum molecular weight of 74000. 6. The structures of wool proteins are discussed in the light of the peptide sequences determined, and, in particular, of those sequences in fraction U.S.3 that could not be elucidated. PMID:16742497

  13. The complete amino acid sequence of the major Kunitz trypsin inhibitor from the seeds of Prosopsis juliflora.

    PubMed

    Negreiros, A N; Carvalho, M M; Xavier Filho, J; Blanco-Labra, A; Shewry, P R; Richardson, M

    1991-01-01

    The major inhibitor of trypsin in seeds of Prosopsis juliflora was purified by precipitation with ammonium sulphate, ion-exchange column chromatography on DEAE- and CM-Sepharose and preparative reverse phase HPLC on a Vydac C-18 column. The protein inhibited trypsin in the stoichiometric ratio of 1:1, but had only weak activity against chymotrypsin and did not inhibit human salivary or porcine pancreatic alpha-amylases. SDS-PAGE indicated that the inhibitor has a Mr of ca 20,000, and IEF-PAGE showed that the pI is 8.8. The complete amino acid sequence was determined by automatic degradation, and by DABITC/PITC microsequence analysis of peptides obtained from enzyme digestions of the reduced and S-carboxymethylated protein with trypsin, chymotrypsin, elastase, the Glu-specific protease from S. aureus and the Lys-specific protease from Lysobacter enzymogenes. The inhibitor consisted of two polypeptide chains, of 137 residues (alpha chain) and 38 residues (beta chain) linked together by a single disulphide bond. The amino acid sequence of the protein exhibited homology with a number of Kunitz proteinase inhibitors from other legume seeds, the bifunctional subtilisin/alpha-amylase inhibitors from cereals and the taste-modifying protein miraculin. PMID:1367792

  14. Perfluoroalkyl Acids in Maternal Serum and Indices of Fetal Growth: The Aarhus Birth Cohort

    PubMed Central

    Bach, Cathrine Carlsen; Bech, Bodil Hammer; Nohr, Ellen Aagaard; Olsen, Jørn; Matthiesen, Niels Bjerregård; Bonefeld-Jørgensen, Eva Cecilie; Bossi, Rossana; Henriksen, Tine Brink

    2015-01-01

    Background: Previous studies indicated an association between intrauterine exposure to perfluorooctane sulfonate (PFOS) or perfluorooctanoate (PFOA) and lower birth weight. However, these perfluoroalkyl acids (PFAAs) have to some extent been substituted by other compounds on which little is known. Objectives: We investigated the association between specific PFAAs and birth weight, birth length, and head circumference at birth. Methods: We studied 1,507 mothers and their children from the Aarhus Birth Cohort (2008–2013). Nulliparous women were included during pregnancy, and serum levels of 16 PFAAs were measured between 9 and 20 completed gestational weeks (96% within 13 weeks). For compounds with quantifiable values in > 50% of samples (7 compounds), we report the associations with birth weight, birth length, and head circumference at birth determined by multivariable linear regression. Results: Estimated mean birth weights were lower among women with serum perfluorohexane sulfonate, perfluoroheptane sulfonate, and PFOS concentrations above the lowest exposure quartile, but we found no consistent monotonic dose–response patterns. These associations were stronger when the population was restricted to term births (n = 1,426). For PFOS, the birth weight estimates for the highest versus lowest quartile were –50 g (95% CI: –123, 23 g) in all births and –62 g (95% CI: –126, 3 g) in term births. For the other PFAAs, the direction of the associations was inconsistent, and no overall association with birth weight was apparent. No PFAAs were associated with birth length or head circumference at birth. Conclusions: Overall, we did not find strong or consistent associations between PFAAs and birth weight or other indices of fetal growth, though estimated mean birth weights were lower among those with exposures above the lowest quartile for some compounds. Citation: Bach CC, Bech BH, Nohr EA, Olsen J, Matthiesen NB, Bonefeld-Jørgensen EC, Bossi R, Henriksen TB

  15. Whole genome sequencing indicates Corynebacterium jeikeium comprises 4 separate genomospecies and identifies a dominant genomospecies among clinical isolates

    PubMed Central

    Salipante, Stephen J.; Sengupta, Dhruba J.; Cummings, Lisa A.; Robinson, Aaron; Kurosawa, Kyoko; Hoogestraat, Daniel R.; Cookson, Brad T.

    2014-01-01

    Corynebacterium jeikeium is an opportunistic pathogen which has been noted for significant genomic diversity. The population structure within this species remains poorly understood. Here we explore the relationships among fifteen clinical isolates of C. jeikeium (reference strains K411 and ATCC 43734, and 13 primary isolates collected over a period of 7 years) through genetic, genomic, and phenotypic studies. We report a high degree of divergence among strains based on 16S ribosomal RNA (rRNA) gene and rpoB gene sequence analysis, supporting the presence of genetically distinct subgroups. Whole genome sequencing indicates genomic-level dissimilarity among subgroups, which qualify as 4 separate and distinct Corynebacterium species based on an Average Nucleotide Identity (ANIb) threshold of < 95%. Functional distinctions in antibiotic susceptibilities and metabolic profiles characterize two of these genomospecies, allowing their differentiation from others through routine laboratory testing. The remaining genomospecies can be classified through a biphasic approach integrating phenotypic testing and rpoB gene sequencing. The genomospecies predominantly recovered from patient specimens does not include either of the existing C. jeikeium reference strains, implying that studies of this pathogen would benefit from examination of representatives from the primary disease-causing group. The clinically dominant genomospecies also has the smallest genome size and gene repertoire, suggesting the possibility of increased virulence relative to the other genomospecies. The ability to classify isolates to one of the four C. jeikeium genomospecies in a clinical context provides diagnostic information for tailoring antimicrobial therapy and may aid in identification of species-specific disease associations. PMID:25116839

  16. Nucleic and amino acid sequences relating to a novel transketolase, and methods for the expression thereof

    DOEpatents

    Croteau, Rodney Bruce; Wildung, Mark Raymond; Lange, Bernd Markus; McCaskill, David G.

    2001-01-01

    cDNAs encoding 1-deoxyxylulose-5-phosphate synthase from peppermint (Mentha piperita) have been isolated and sequenced, and the corresponding amino acid sequences have been determined. Accordingly, isolated DNA sequences (SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7) are provided which code for the expression of 1-deoxyxylulose-5-phosphate synthase from plants. In another aspect the present invention provides for isolated, recombinant DXPS proteins, such as the proteins having the sequences set forth in SEQ ID NO:4, SEQ ID NO:6 and SEQ ID NO:8. In other aspects, replicable recombinant cloning vehicles are provided which code for plant 1-deoxyxylulose-5-phosphate synthases, or for a base sequence sufficiently complementary to at least a portion of 1-deoxyxylulose-5-phosphate synthase DNA or RNA to enable hybridization therewith. In yet other aspects, modified host cells are provided that have been transformed, transfected, infected and/or injected with a recombinant cloning vehicle and/or DNA sequence encoding a plant 1-deoxyxylulose-5-phosphate synthase. Thus, systems and methods are provided for the recombinant expression of the aforementioned recombinant 1-deoxyxylulose-5-phosphate synthase that may be used to facilitate its production, isolation and purification in significant amounts. Recombinant 1-deoxyxylulose-5-phosphate synthase may be used to obtain expression or enhanced expression of 1-deoxyxylulose-5-phosphate synthase in plants in order to enhance the production of 1-deoxyxylulose-5-phosphate, or its derivatives such as isopentenyl diphosphate (BP), or may be otherwise employed for the regulation or expression of 1-deoxyxylulose-5-phosphate synthase, or the production of its products.

  17. Novel method for PIK3CA mutation analysis: locked nucleic acid--PCR sequencing.

    PubMed

    Ang, Daphne; O'Gara, Rebecca; Schilling, Amy; Beadling, Carol; Warrick, Andrea; Troxell, Megan L; Corless, Christopher L

    2013-05-01

    Somatic mutations in PIK3CA are commonly seen in invasive breast cancer and several other carcinomas, occurring in three hotspots: codons 542 and 545 of exon 9 and in codon 1047 of exon 20. We designed a locked nucleic acid (LNA)-PCR sequencing assay to detect low levels of mutant PIK3CA DNA with attention to avoiding amplification of a pseudogene on chromosome 22 that has >95% homology to exon 9 of PIK3CA. We tested 60 FFPE breast DNA samples with known PIK3CA mutation status (48 cases had one or more PIK3CA mutations, and 12 were wild type) as identified by PCR-mass spectrometry. PIK3CA exons 9 and 20 were amplified in the presence or absence of LNA-oligonucleotides designed to bind to the wild-type sequences for codons 542, 545, and 1047, and partially suppress their amplification. LNA-PCR sequencing confirmed all 51 PIK3CA mutations; however, the mutation detection rate by standard Sanger sequencing was only 69% (35 of 51). Of the 12 PIK3CA wild-type cases, LNA-PCR sequencing detected three additional H1047R mutations in "normal" breast tissue and one E545K in usual ductal hyperplasia. Histopathological review of these three normal breast specimens showed columnar cell change in two (both with known H1047R mutations) and apocrine metaplasia in one. The novel LNA-PCR shows higher sensitivity than standard Sanger sequencing and did not amplify the known pseudogene. PMID:23541593

  18. Genome Sequence Analysis of the Naphthenic Acid Degrading and Metal Resistant Bacterium Cupriavidus gilardii CR3.

    PubMed

    Wang, Xiaoyu; Chen, Meili; Xiao, Jingfa; Hao, Lirui; Crowley, David E; Zhang, Zhewen; Yu, Jun; Huang, Ning; Huo, Mingxin; Wu, Jiayan

    2015-01-01

    Cupriavidus sp. are generally heavy metal tolerant bacteria with the ability to degrade a variety of aromatic hydrocarbon compounds, although the degradation pathways and substrate versatilities remain largely unknown. Here we studied the bacterium Cupriavidus gilardii strain CR3, which was isolated from a natural asphalt deposit, and which was shown to utilize naphthenic acids as a sole carbon source. Genome sequencing of C. gilardii CR3 was carried out to elucidate possible mechanisms for the naphthenic acid biodegradation. The genome of C. gilardii CR3 was composed of two circular chromosomes chr1 and chr2 of respectively 3,539,530 bp and 2,039,213 bp in size. The genome for strain CR3 encoded 4,502 putative protein-coding genes, 59 tRNA genes, and many other non-coding genes. Many genes were associated with xenobiotic biodegradation and metal resistance functions. Pathway prediction for degradation of cyclohexanecarboxylic acid, a representative naphthenic acid, suggested that naphthenic acid undergoes initial ring-cleavage, after which the ring fission products can be degraded via several plausible degradation pathways including a mechanism similar to that used for fatty acid oxidation. The final metabolic products of these pathways are unstable or volatile compounds that were not toxic to CR3. Strain CR3 was also shown to have tolerance to at least 10 heavy metals, which was mainly achieved by self-detoxification through ion efflux, metal-complexation and metal-reduction, and a powerful DNA self-repair mechanism. Our genomic analysis suggests that CR3 is well adapted to survive the harsh environment in natural asphalts containing naphthenic acids and high concentrations of heavy metals. PMID:26301592

  19. Genome Sequence Analysis of the Naphthenic Acid Degrading and Metal Resistant Bacterium Cupriavidus gilardii CR3

    PubMed Central

    Xiao, Jingfa; Hao, Lirui; Crowley, David E.; Zhang, Zhewen; Yu, Jun; Huang, Ning; Huo, Mingxin; Wu, Jiayan

    2015-01-01

    Cupriavidus sp. are generally heavy metal tolerant bacteria with the ability to degrade a variety of aromatic hydrocarbon compounds, although the degradation pathways and substrate versatilities remain largely unknown. Here we studied the bacterium Cupriavidus gilardii strain CR3, which was isolated from a natural asphalt deposit, and which was shown to utilize naphthenic acids as a sole carbon source. Genome sequencing of C. gilardii CR3 was carried out to elucidate possible mechanisms for the naphthenic acid biodegradation. The genome of C. gilardii CR3 was composed of two circular chromosomes chr1 and chr2 of respectively 3,539,530 bp and 2,039,213 bp in size. The genome for strain CR3 encoded 4,502 putative protein-coding genes, 59 tRNA genes, and many other non-coding genes. Many genes were associated with xenobiotic biodegradation and metal resistance functions. Pathway prediction for degradation of cyclohexanecarboxylic acid, a representative naphthenic acid, suggested that naphthenic acid undergoes initial ring-cleavage, after which the ring fission products can be degraded via several plausible degradation pathways including a mechanism similar to that used for fatty acid oxidation. The final metabolic products of these pathways are unstable or volatile compounds that were not toxic to CR3. Strain CR3 was also shown to have tolerance to at least 10 heavy metals, which was mainly achieved by self-detoxification through ion efflux, metal-complexation and metal-reduction, and a powerful DNA self-repair mechanism. Our genomic analysis suggests that CR3 is well adapted to survive the harsh environment in natural asphalts containing naphthenic acids and high concentrations of heavy metals. PMID:26301592

  20. Bile acid sulfotransferase I from rat liver sulfates bile acids and 3-hydroxy steroids: purification, N-terminal amino acid sequence, and kinetic properties.

    PubMed

    Barnes, S; Buchina, E S; King, R J; McBurnett, T; Taylor, K B

    1989-04-01

    A bile acid:3'phosphoadenosine-5'phosphosulfate:sulfotransferase (BAST I) from adult female rat liver cytosol has been purified 157-fold by a two-step isolation procedure. The N-terminal amino acid sequence of the 30,000 subunit has been determined for the first 35 residues. The Vmax of purified BAST I is 18.7 nmol/min per mg protein with N-(3-hydroxy-5 beta-cholanoyl)glycine (glycolithocholic acid) as substrate, comparable to that of the corresponding purified human BAST (Chen, L-J., and I. H. Segel, 1985. Arch. Biochem. Biophys. 241: 371-379). BAST I activity has a broad pH optimum from 5.5-7.5. Although maximum activity occurs with 5 mM MgCl2, Mg2+ is not essential for BAST I activity. The greatest sulfotransferase activity and the highest substrate affinity is observed with bile acids or steroids that have a steroid nucleus containing a 3 beta-hydroxy group and a 5-6 double bond or a trans A-B ring junction. These substrates have normal hyperbolic initial velocity curves with substrate inhibition occurring above 5 microM. Of the saturated 5 beta-bile acids, those with a single 3-hydroxy group are the most active. The addition of a second hydroxy group at the 6- or 7-position eliminates more than 99% of the activity. In contrast, 3 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oic acid (deoxycholic acid) is an excellent substrate. The initial velocity curves for glycolithocholic and deoxycholic acid conjugates are sigmoidal rather than hyperbolic, suggestive of an allosteric effect. Maximum activity is observed at 80 microM for glycolithocholic acid. All substrates, bile acids and steroids, are inhibited by the 5 beta-bile acid, 3-keto-5 beta-cholanoic acid. The data suggest that BAST I is the same protein as hydrosteroid sulfotransferase 2 (Marcus, C. J., et al. 1980. Anal. Biochem. 107: 296-304). PMID:2754334

  1. Extremely Acidophilic Protists from Acid Mine Drainage Host Rickettsiales-Lineage Endosymbionts That Have Intervening Sequences in Their 16S rRNA Genes

    PubMed Central

    Baker, Brett J.; Hugenholtz, Philip; Dawson, Scott C.; Banfield, Jillian F.

    2003-01-01

    During a molecular phylogenetic survey of extremely acidic (pH < 1), metal-rich acid mine drainage habitats in the Richmond Mine at Iron Mountain, Calif., we detected 16S rRNA gene sequences of a novel bacterial group belonging to the order Rickettsiales in the Alphaproteobacteria. The closest known relatives of this group (92% 16S rRNA gene sequence identity) are endosymbionts of the protist Acanthamoeba. Oligonucleotide 16S rRNA probes were designed and used to observe members of this group within acidophilic protists. To improve visualization of eukaryotic populations in the acid mine drainage samples, broad-specificity probes for eukaryotes were redesigned and combined to highlight this component of the acid mine drainage community. Approximately 4% of protists in the acid mine drainage samples contained endosymbionts. Measurements of internal pH of the protists showed that their cytosol is close to neutral, indicating that the endosymbionts may be neutrophilic. The endosymbionts had a conserved 273-nucleotide intervening sequence (IVS) in variable region V1 of their 16S rRNA genes. The IVS does not match any sequence in current databases, but the predicted secondary structure forms well-defined stem loops. IVSs are uncommon in rRNA genes and appear to be confined to bacteria living in close association with eukaryotes. Based on the phylogenetic novelty of the endosymbiont sequences and initial culture-independent characterization, we propose the name “Candidatus Captivus acidiprotistae.” To our knowledge, this is the first report of an endosymbiotic relationship in an extremely acidic habitat. PMID:12957940

  2. Unconventional amino acid sequence of the sun anemone (Stoichactis helianthus) polypeptide neurotoxin

    SciTech Connect

    Kem, W.; Dunn, B.; Parten, B.; Pennington, M.; Price, D.

    1986-05-01

    A 5000 dalton polypeptide neurotoxin (Sh-NI) purified by G50 Sephadex, P-cellulose, and SP-Sephadex chromatography was homogeneous by isoelectric focusing. Sh-NI was highly toxic to crayfish (LD/sub 50/ 0.6 ..mu..g/kg) but without effect upon mice at 15,000 ..mu..g/kg (i.p. injection). The reduced, /sup 3/H-carboxymethylated toxin and its fragments were subjected to automatic Edman degradation and the resulting PTH-amino acids were identified by HPLC, back hydrolysis, and scintillation counting. Peptides resulting from proteolytic (clostripain, staphylococcal protease) and chemical (tryptophan) cleavage were sequenced. The sequence is: AACKCDDEGPDIRTAPLTGTVDLGSCNAGWEKCASYYTIIADCCRKKK. This sequence differs considerably from the homologous Anemonia and Anthopleura toxins; many of the identical residues (6 half-cystines, G9, P10, R13, G19, G29, W30) are probably critical for folding rather than receptor recognition. However, the Sh-NI sequence closely resembles Radioanthus macrodactylus neurotoxin III and r. paumotensis II. The authors propose that Sh-NI and related Radioanthus toxins act upon a different site on the sodium channel.

  3. Repeat sequence chromosome specific nucleic acid probes and methods of preparing and using

    DOEpatents

    Weier, H.U.G.; Gray, J.W.

    1995-06-27

    A primer directed DNA amplification method to isolate efficiently chromosome-specific repeated DNA wherein degenerate oligonucleotide primers are used is disclosed. The probes produced are a heterogeneous mixture that can be used with blocking DNA as a chromosome-specific staining reagent, and/or the elements of the mixture can be screened for high specificity, size and/or high degree of repetition among other parameters. The degenerate primers are sets of primers that vary in sequence but are substantially complementary to highly repeated nucleic acid sequences, preferably clustered within the template DNA, for example, pericentromeric alpha satellite repeat sequences. The template DNA is preferably chromosome-specific. Exemplary primers and probes are disclosed. The probes of this invention can be used to determine the number of chromosomes of a specific type in metaphase spreads, in germ line and/or somatic cell interphase nuclei, micronuclei and/or in tissue sections. Also provided is a method to select arbitrarily repeat sequence probes that can be screened for chromosome-specificity. 18 figs.

  4. Repeat sequence chromosome specific nucleic acid probes and methods of preparing and using

    DOEpatents

    Weier, Heinz-Ulrich G.; Gray, Joe W.

    1995-01-01

    A primer directed DNA amplification method to isolate efficiently chromosome-specific repeated DNA wherein degenerate oligonucleotide primers are used is disclosed. The probes produced are a heterogeneous mixture that can be used with blocking DNA as a chromosome-specific staining reagent, and/or the elements of the mixture can be screened for high specificity, size and/or high degree of repetition among other parameters. The degenerate primers are sets of primers that vary in sequence but are substantially complementary to highly repeated nucleic acid sequences, preferably clustered within the template DNA, for example, pericentromeric alpha satellite repeat sequences. The template DNA is preferably chromosome-specific. Exemplary primers ard probes are disclosed. The probes of this invention can be used to determine the number of chromosomes of a specific type in metaphase spreads, in germ line and/or somatic cell interphase nuclei, micronuclei and/or in tissue sections. Also provided is a method to select arbitrarily repeat sequence probes that can be screened for chromosome-specificity.

  5. Real refractive indices of infrared-characterized nitric-acid/ice films: Implications for optical measurements of polar stratospheric clouds

    NASA Technical Reports Server (NTRS)

    Middlebrook, Ann M.; Berland, Brian S.; George, Steven M.; Tolbert, Margaret A.; Toon, Owen B.

    1994-01-01

    The infrared spectra of nitric-acid/ice films representative of polar stratospheric clouds (PSCs) were collected with simultaneous optical interference measurements to determine the real refractive indices at lambda = 632 nm. Ice and amphorous nitric-acid/ice films were prepared by condensation of water and nitric acid vapors onto a wedged Al2O3 substrate. The real refractive indices of these films were determined from the optical interference of a reflected helium-neon laser during film growth. The indices of the amphorous films varied smoothly from n = 1.30 for ice to n = 1.49 for nitric acid, similar to observations in previous work. We were unable to obtain the refractive index of crystlline films during adsorption because of optical scattering caused by surface roughness. Therefore crystlline nitric acid hydrate films were prepared by annealing amphorous nitric-acid/ice films. Further heating caused desorption of the crystalline hydrate films. During desorption, the refractive indices for ice, NAM (nitric acid monohydrate), alpha- and beta-NAT (nitric acid trihydrate) films were measured using the optical interference technique. In agreement with earlier data, the real refractive indices for ice and NAM determined in desorption were n = 1.30 +/- 0.01 and n = 1.53 +/- 0.03, respectively. The real refractive indices for alpha- and beta-NAT were found to be n = 1.51 +/- 0.01 and n greater than or equal to 1.46, respectively. Our measurements also suggest that the shape of crystalline nitric acid particles may depend on whether they nucleate from the liquid or by vapor deposition. If confirmed by future studies, this observation may provide a means of distinguishing the nucleation mechanism of crystalline PSCs.

  6. Morphological tranformation of calcite crystal growth by prismatic "acidic" polypeptide sequences.

    SciTech Connect

    Kim, I; Giocondi, J L; Orme, C A; Collino, J; Evans, J S

    2007-02-13

    Many of the interesting mechanical and materials properties of the mollusk shell are thought to stem from the prismatic calcite crystal assemblies within this composite structure. It is now evident that proteins play a major role in the formation of these assemblies. Recently, a superfamily of 7 conserved prismatic layer-specific mollusk shell proteins, Asprich, were sequenced, and the 42 AA C-terminal sequence region of this protein superfamily was found to introduce surface voids or porosities on calcite crystals in vitro. Using AFM imaging techniques, we further investigate the effect that this 42 AA domain (Fragment-2) and its constituent subdomains, DEAD-17 and Acidic-2, have on the morphology and growth kinetics of calcite dislocation hillocks. We find that Fragment-2 adsorbs on terrace surfaces and pins acute steps, accelerates then decelerates the growth of obtuse steps, forms clusters and voids on terrace surfaces, and transforms calcite hillock morphology from a rhombohedral form to a rounded one. These results mirror yet are distinct from some of the earlier findings obtained for nacreous polypeptides. The subdomains Acidic-2 and DEAD-17 were found to accelerate then decelerate obtuse steps and induce oval rather than rounded hillock morphologies. Unlike DEAD-17, Acidic-2 does form clusters on terrace surfaces and exhibits stronger obtuse velocity inhibition effects than either DEAD-17 or Fragment-2. Interestingly, a 1:1 mixture of both subdomains induces an irregular polygonal morphology to hillocks, and exhibits the highest degree of acute step pinning and obtuse step velocity inhibition. This suggests that there is some interplay between subdomains within an intra (Fragment-2) or intermolecular (1:1 mixture) context, and sequence interplay phenomena may be employed by biomineralization proteins to exert net effects on crystal growth and morphology.

  7. Fast computational methods for predicting protein structure from primary amino acid sequence

    DOEpatents

    Agarwal, Pratul Kumar

    2011-07-19

    The present invention provides a method utilizing primary amino acid sequence of a protein, energy minimization, molecular dynamics and protein vibrational modes to predict three-dimensional structure of a protein. The present invention also determines possible intermediates in the protein folding pathway. The present invention has important applications to the design of novel drugs as well as protein engineering. The present invention predicts the three-dimensional structure of a protein independent of size of the protein, overcoming a significant limitation in the prior art.

  8. Amino-terminal amino acid sequence of the major structural polypeptides of avian retroviruses: sequence homology between reticuloendotheliosis virus p30 and p30s of mammalian retroviruses.

    PubMed Central

    Hunter, E; Bhown, A S; Bennett, J C

    1978-01-01

    The major structural polypeptides, p30 of reticuloendotheliosis virus (REV) (strain T) and p27 of avian sarcoma virus B77, have been compared with regard to amino acid composition. NH2-terminal amino acid sequence, and immunological crossreactions. The amino acid composition of the two polypeptides is distinct, and a comparison of the first 30 NH2-terminal amino acids of REV p30 with that for the first 25 of B77 p27 yields only three homologous residues. In competition radioimmunoassays the polypeptides show no crossreactivity. A comparison of the amino acid composition and NH2-terminal amino acid sequence of REV p30 with those reported for several mammalian retrovirus p30s shows remarkable similarities. Both REV and mammalian p30s contain a large number of polar residues in their amino acid composition and show approximately 40% homology in the first 30 NH2-terminal amino acids. No crossreactivity could be observed, however, in competition radioimmunoassays between Rauscher murine leukemia virus p30 and that of REV. The observations reported here suggest a close evolutionary relationship between REV and the mammalian retroviruses. Images PMID:208072

  9. Purification and amino acid sequence of aminopeptidase P from pig kidney.

    PubMed

    Vergas Romero, C; Neudorfer, I; Mann, K; Schäfer, W

    1995-04-01

    Aminopeptidase P from kidney cortex was purified in high yield (recovery greater than or equal to 20%) by a series of column chromatographic steps after solubilization of the membrane-bound glycoprotein with n-butanol. A coupled enzymic assay, using Gly-Pro-Pro-NH-Nap as substrate and dipeptidyl-peptidase IV as auxilliary enzyme, was used to monitor the purification. The purification procedure yielded two forms of aminopeptidase P differing in their carbohydrate composition (glycoforms). Both enzyme preparations were homogeneous as assessed by SDS/PAGE silver staining, and isoelectric focusing. Both forms possessed the same substrate specificity, catalysed the same reaction, and consisted of identical protein chains. The amino acid sequence determined by Edman degradation and mass spectrometry consisted of 623 amino acids. Six N-glycosylation sites, all contained in the N-terminal half of the protein, were characterized. PMID:7744038

  10. Draft Genome Sequence of Cupriavidus sp. Strain SK-3, a 4-Chlorobiphenyl- and 4-Clorobenzoic Acid-Degrading Bacterium

    PubMed Central

    Vilo, Claudia; Benedik, Michael J.; Ilori, Matthew

    2014-01-01

    We report the draft genome sequence of Cupriavidus sp. strain SK-3, which can use 4-chlorobiphenyl and 4-clorobenzoic acid as the sole carbon source for growth. The draft genome sequence allowed the study of the polychlorinated biphenyl degradation mechanism and the recharacterization of the strain SK-3 as a Cupriavidus species. PMID:24994805

  11. Draft Genome Sequence of Bacillus subtilis subsp. natto Strain CGMCC 2108, a High Producer of Poly-γ-Glutamic Acid

    PubMed Central

    Tan, Siyuan; Su, Anping; Zhang, Chen; Ren, Yuanyuan

    2016-01-01

    Here, we report the 4.1-Mb draft genome sequence of Bacillus subtilis subsp. natto strain CGMCC 2108, a high producer of poly-γ-glutamic acid (γ-PGA). This sequence will provide further help for the biosynthesis of γ-PGA and will greatly facilitate research efforts in metabolic engineering of B. subtilis subsp. natto strain CGMCC 2108. PMID:27231363

  12. New monoclonal antibodies to the Ebola virus glycoprotein: Identification and analysis of the amino acid sequence of the variable domains.

    PubMed

    Panina, A A; Aliev, T K; Shemchukova, O B; Dement'yeva, I G; Varlamov, N E; Pozdnyakova, L P; Bokov, M N; Dolgikh, D A; Sveshnikov, P G; Kirpichnikov, M P

    2016-03-01

    We determined the nucleotide and amino acid sequences of variable domains of three new monoclonal antibodies to the glycoprotein of Ebola virus capsid. The framework and hypervariable regions of immunoglobulin heavy and light chains were identified. The primary structures were confirmed using massspectrometry analysis. Immunoglobulin database search showed the uniqueness of the sequences obtained. PMID:27193713

  13. Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis subsp. lactis TOMSC161, Isolated from a Nonscalded Curd Pressed Cheese

    PubMed Central

    Velly, H.; Abraham, A.-L.; Loux, V.; Delacroix-Buchet, A.; Fonseca, F.; Bouix, M.

    2014-01-01

    Lactococcus lactis is a lactic acid bacterium used in the production of many fermented foods, such as dairy products. Here, we report the genome sequence of L. lactis subsp. lactis TOMSC161, isolated from nonscalded curd pressed cheese. This genome sequence provides information in relation to dairy environment adaptation. PMID:25377704

  14. Draft Genome Sequence of Bacillus subtilis subsp. natto Strain CGMCC 2108, a High Producer of Poly-γ-Glutamic Acid.

    PubMed

    Tan, Siyuan; Meng, Yonghong; Su, Anping; Zhang, Chen; Ren, Yuanyuan

    2016-01-01

    Here, we report the 4.1-Mb draft genome sequence of Bacillus subtilis subsp. natto strain CGMCC 2108, a high producer of poly-γ-glutamic acid (γ-PGA). This sequence will provide further help for the biosynthesis of γ-PGA and will greatly facilitate research efforts in metabolic engineering of B. subtilis subsp. natto strain CGMCC 2108. PMID:27231363

  15. Aminolevulinic acid-dehydratase activity in green sunfish: An indicator of lead bioavailability in suspended sediments

    SciTech Connect

    Caldwell, C.; Steingraeber, M.

    1995-12-31

    Green sunfish (Lepomis cyanellus, 6-32 g) were exposed for 28 days to suspended sediments and sediments not in suspension (bedded). Blood was collected for aminolevulinic acid-dehydratase activity (ALA-D), hemoglobin and blood lead concentrations, and whole body lead concentrations. Results of the metal analysis of filtered and unfiltered water from both suspended and bedded sediment tests revealed that the majority of the metal was associated with particulate matter in suspension. Fish subjected to suspended sediments (0.2, 1.6, and 21.9 ug/g lead, dry weight) had significantly reduced ALA-D and hemoglobin concentrations than fish in bedded sediments. However, there was no significant correlation between lead concentrations in suspended sediments and ALA-D activity. In addition, there were no significant correlations between ALA-D activity and hemoglobin concentrations. Whole body analysis and blood lead concentrations were not good indicators of lead exposure in sediment tests having relatively low concentrations of lead.

  16. ANTICALIgN: visualizing, editing and analyzing combined nucleotide and amino acid sequence alignments for combinatorial protein engineering.

    PubMed

    Jarasch, Alexander; Kopp, Melanie; Eggenstein, Evelyn; Richter, Antonia; Gebauer, Michaela; Skerra, Arne

    2016-07-01

    ANTIC ALIGN: is an interactive software developed to simultaneously visualize, analyze and modify alignments of DNA and/or protein sequences that arise during combinatorial protein engineering, design and selection. ANTIC ALIGN: combines powerful functions known from currently available sequence analysis tools with unique features for protein engineering, in particular the possibility to display and manipulate nucleotide sequences and their translated amino acid sequences at the same time. ANTIC ALIGN: offers both template-based multiple sequence alignment (MSA), using the unmutated protein as reference, and conventional global alignment, to compare sequences that share an evolutionary relationship. The application of similarity-based clustering algorithms facilitates the identification of duplicates or of conserved sequence features among a set of selected clones. Imported nucleotide sequences from DNA sequence analysis are automatically translated into the corresponding amino acid sequences and displayed, offering numerous options for selecting reading frames, highlighting of sequence features and graphical layout of the MSA. The MSA complexity can be reduced by hiding the conserved nucleotide and/or amino acid residues, thus putting emphasis on the relevant mutated positions. ANTIC ALIGN: is also able to handle suppressed stop codons or even to incorporate non-natural amino acids into a coding sequence. We demonstrate crucial functions of ANTIC ALIGN: in an example of Anticalins selected from a lipocalin random library against the fibronectin extradomain B (ED-B), an established marker of tumor vasculature. Apart from engineered protein scaffolds, ANTIC ALIGN: provides a powerful tool in the area of antibody engineering and for directed enzyme evolution. PMID:27261456

  17. Formation Sequences of Iron Minerals in the Acidic Alteration Products and Variation of Hydrothermal Fluid Conditions

    NASA Astrophysics Data System (ADS)

    Isobe, H.; Yoshizawa, M.

    2008-12-01

    Iron minerals have important role in environmental issues not only on the Earth but also other terrestrial planets. Iron mineral species related to alteration products of primary minerals with surface or subsurface fluids are characterized by temperature, acidity and redox conditions of the fluids. We can see various iron- bearing alteration products in alteration products around fumaroles in geothermal/volcanic areas. In this study, zonal structures of iron minerals in alteration products of the geothermal area are observed to elucidate temporal and spatial variation of hydrothermal fluids. Alteration of the pyroxene-amphibole andesite of Garan-dake volcano, Oita, Japan occurs by the acidic hydrothermal fluid to form cristobalite leaching out elements other than Si. Hand specimens with unaltered or weakly altered core and cristobalite crust show various sequences of layers. XRD analysis revealed that the alteration degree is represented by abundance of cristobalite. Intermediately altered layers are characterized by occurrence including alunite, pyrite, kaolinite, goethite and hematite. A specimen with reddish brown core surrounded by cristobalite-rich white crust has brown colored layers at the boundary of core and the crust. Reddish core is characterized by occurrence of crystalline hematite by XRD. Another hand specimen has light gray core, which represents reduced conditions, and white cristobalite crust with light brown and reddish brown layers of ferric iron minerals between the core and the crust. On the other hand, hornblende crystals, typical ferrous iron-bearing mineral of the host rock, are well preserved in some samples with strongly decolorized cristobalite-rich groundmass. Hydrothermal alteration experiments of iron-rich basaltic material shows iron mineral species depend on acidity and temperature of the fluid. Oxidation states of the iron-bearing mineral species are strongly influenced by the acidity and redox conditions. Variations of alteration

  18. Analysis of the indices of acidity in the soil profile and their relationship with pedogenesis

    NASA Astrophysics Data System (ADS)

    Kokotov, Yu. A.; Sukhacheva, E. Yu.; Aparin, B. F.

    2016-01-01

    A new notion—an acidic trace of pedogenesis in the field of soil acidity—is suggested. This notion implies a three-dimensional representation of the distribution of soil acidity in the soil profile and can be graphically shown in three two-dimensional projections that can be combined on a common V-diagram. Such V-diagrams are individual for each particular soil profile. At the same time, they have some common phenomenology in their shapes and in the position in the acidity field. A tendency for the S-shaped form of acidic trace is manifested by a sharp decrease in pH upon the reduction of base saturation at the high and low values of this index and by small changes in pH at the moderate values of base saturation in the area of acid buffering of the soil profile. This phenomenon is related to the weak acidity and polyfunctionality of the soils as ionite systems. An acidic trace can be subdivided into several characteristic parts related to different pedogenetic processes in their interaction. Its position in the field of acidity is largely determined by the acidity of parent material. Acidic traces of different types of soils in the northwestern Russia are discussed. It is argued that V-diagrams should be analyzed together with other soil characteristics.

  19. Multiple Amino Acid Sequence Alignment Nitrogenase Component 1: Insights into Phylogenetics and Structure-Function Relationships

    PubMed Central

    Howard, James B.; Kechris, Katerina J.; Rees, Douglas C.; Glazer, Alexander N.

    2013-01-01

    Amino acid residues critical for a protein's structure-function are retained by natural selection and these residues are identified by the level of variance in co-aligned homologous protein sequences. The relevant residues in the nitrogen fixation Component 1 α- and β-subunits were identified by the alignment of 95 protein sequences. Proteins were included from species encompassing multiple microbial phyla and diverse ecological niches as well as the nitrogen fixation genotypes, anf, nif, and vnf, which encode proteins associated with cofactors differing at one metal site. After adjusting for differences in sequence length, insertions, and deletions, the remaining >85% of the sequence co-aligned the subunits from the three genotypes. Six Groups, designated Anf, Vnf , and Nif I-IV, were assigned based upon genetic origin, sequence adjustments, and conserved residues. Both subunits subdivided into the same groups. Invariant and single variant residues were identified and were defined as “core” for nitrogenase function. Three species in Group Nif-III, Candidatus Desulforudis audaxviator, Desulfotomaculum kuznetsovii, and Thermodesulfatator indicus, were found to have a seleno-cysteine that replaces one cysteinyl ligand of the 8Fe:7S, P-cluster. Subsets of invariant residues, limited to individual groups, were identified; these unique residues help identify the gene of origin (anf, nif, or vnf) yet should not be considered diagnostic of the metal content of associated cofactors. Fourteen of the 19 residues that compose the cofactor pocket are invariant or single variant; the other five residues are highly variable but do not correlate with the putative metal content of the cofactor. The variable residues are clustered on one side of the cofactor, away from other functional centers in the three dimensional structure. Many of the invariant and single variant residues were not previously recognized as potentially critical and their identification provides the bases

  20. Draft Genome Sequences of Gluconobacter cerinus CECT 9110 and Gluconobacter japonicus CECT 8443, Acetic Acid Bacteria Isolated from Grape Must

    PubMed Central

    Sainz, Florencia

    2016-01-01

    We report here the draft genome sequences of Gluconobacter cerinus strain CECT9110 and Gluconobacter japonicus CECT8443, acetic acid bacteria isolated from grape must. Gluconobacter species are well known for their ability to oxidize sugar alcohols into the corresponding acids. Our objective was to select strains to oxidize effectively d-glucose. PMID:27365351

  1. Swfoldrate: predicting protein folding rates from amino acid sequence with sliding window method.

    PubMed

    Cheng, Xiang; Xiao, Xuan; Wu, Zhi-cheng; Wang, Pu; Lin, Wei-zhong

    2013-01-01

    Protein folding is the process by which a protein processes from its denatured state to its specific biologically active conformation. Understanding the relationship between sequences and the folding rates of proteins remains an important challenge. Most previous methods of predicting protein folding rate require the tertiary structure of a protein as an input. In this study, the long-range and short-range contact in protein were used to derive extended version of the pseudo amino acid composition based on sliding window method. This method is capable of predicting the protein folding rates just from the amino acid sequence without the aid of any structural class information. We systematically studied the contributions of individual features to folding rate prediction. The optimal feature selection procedures are adopted by means of combining the forward feature selection and sequential backward selection method. Using the jackknife cross validation test, the method was demonstrated on the large dataset. The predictor was achieved on the basis of multitudinous physicochemical features and statistical features from protein using nonlinear support vector machine (SVM) regression model, the method obtained an excellent agreement between predicted and experimentally observed folding rates of proteins. The correlation coefficient is 0.9313 and the standard error is 2.2692. The prediction server is freely available at http://www.jci-bioinfo.cn/swfrate/input.jsp. PMID:22933332

  2. From amino acid sequence to bioactivity: The biomedical potential of antitumor peptides.

    PubMed

    Blanco-Míguez, Aitor; Gutiérrez-Jácome, Alberto; Pérez-Pérez, Martín; Pérez-Rodríguez, Gael; Catalán-García, Sandra; Fdez-Riverola, Florentino; Lourenço, Anália; Sánchez, Borja

    2016-06-01

    Chemoprevention is the use of natural and/or synthetic substances to block, reverse, or retard the process of carcinogenesis. In this field, the use of antitumor peptides is of interest as, (i) these molecules are small in size, (ii) they show good cell diffusion and permeability, (iii) they affect one or more specific molecular pathways involved in carcinogenesis, and (iv) they are not usually genotoxic. We have checked the Web of Science Database (23/11/2015) in order to collect papers reporting on bioactive peptide (1691 registers), which was further filtered searching terms such as "antiproliferative," "antitumoral," or "apoptosis" among others. Works reporting the amino acid sequence of an antiproliferative peptide were kept (60 registers), and this was complemented with the peptides included in CancerPPD, an extensive resource for antiproliferative peptides and proteins. Peptides were grouped according to one of the following mechanism of action: inhibition of cell migration, inhibition of tumor angiogenesis, antioxidative mechanisms, inhibition of gene transcription/cell proliferation, induction of apoptosis, disorganization of tubulin structure, cytotoxicity, or unknown mechanisms. The main mechanisms of action of those antiproliferative peptides with known amino acid sequences are presented and finally, their potential clinical usefulness and future challenges on their application is discussed. PMID:27010507

  3. The amino acid sequences and activities of synergistic hemolysins from Staphylococcus cohnii.

    PubMed

    Mak, Pawel; Maszewska, Agnieszka; Rozalska, Malgorzata

    2008-10-01

    Staphylococcus cohnii ssp. cohnii and S. cohnii ssp. urealyticus are a coagulase-negative staphylococci considered for a long time as unable to cause infections. This situation changed recently and pathogenic strains of these bacteria were isolated from hospital environments, patients and medical staff. Most of the isolated strains were resistant to many antibiotics. The present work describes isolation and characterization of several synergistic peptide hemolysins produced by these bacteria and acting as virulence factors responsible for hemolytic and cytotoxic activities. Amino acid sequences of respective hemolysins from S. cohnii ssp. cohnii (named as H1C, H2C and H3C) and S. cohnii ssp. urealyticus (H1U, H2U and H3U) were identical. Peptides H1 and H3 possessed significant amino acid homology to three synergistic hemolysins secreted by Staphylococcus lugdunensis and to putative antibacterial peptide produced by Staphylococcus saprophyticus ssp. saprophyticus. On the other hand, hemolysin H2 had a unique sequence. All isolated peptides lysed red cells from different mammalian species and exerted a cytotoxic effect on human fibroblasts. PMID:18752624

  4. Clostridium sticklandii, a specialist in amino acid degradation:revisiting its metabolism through its genome sequence

    PubMed Central

    2010-01-01

    Background Clostridium sticklandii belongs to a cluster of non-pathogenic proteolytic clostridia which utilize amino acids as carbon and energy sources. Isolated by T.C. Stadtman in 1954, it has been generally regarded as a "gold mine" for novel biochemical reactions and is used as a model organism for studying metabolic aspects such as the Stickland reaction, coenzyme-B12- and selenium-dependent reactions of amino acids. With the goal of revisiting its carbon, nitrogen, and energy metabolism, and comparing studies with other clostridia, its genome has been sequenced and analyzed. Results C. sticklandii is one of the best biochemically studied proteolytic clostridial species. Useful additional information has been obtained from the sequencing and annotation of its genome, which is presented in this paper. Besides, experimental procedures reveal that C. sticklandii degrades amino acids in a preferential and sequential way. The organism prefers threonine, arginine, serine, cysteine, proline, and glycine, whereas glutamate, aspartate and alanine are excreted. Energy conservation is primarily obtained by substrate-level phosphorylation in fermentative pathways. The reactions catalyzed by different ferredoxin oxidoreductases and the exergonic NADH-dependent reduction of crotonyl-CoA point to a possible chemiosmotic energy conservation via the Rnf complex. C. sticklandii possesses both the F-type and V-type ATPases. The discovery of an as yet unrecognized selenoprotein in the D-proline reductase operon suggests a more detailed mechanism for NADH-dependent D-proline reduction. A rather unusual metabolic feature is the presence of genes for all the enzymes involved in two different CO2-fixation pathways: C. sticklandii harbours both the glycine synthase/glycine reductase and the Wood-Ljungdahl pathways. This unusual pathway combination has retrospectively been observed in only four other sequenced microorganisms. Conclusions Analysis of the C. sticklandii genome and

  5. Complete amino acid sequence of the myoglobin from the Pacific spotted dolphin, Stenella attenuata graffmani.

    PubMed

    Jones, B N; Wang, C C; Dwulet, F E; Lehman, L D; Meuth, J L; Bogardt, R A; Gurd, F R

    1979-04-25

    The complete amino acid sequence of the major component myoglobin from the Pacific spotted dolphin, Stenella attenuata graffmani, was determined by the automated Edman degradation of several large peptides obtained by specific cleavage of the protein. The acetimidated apomyoglobin was selectively cleaved at its two methionyl residues with cyanogen bromide and at its three arginyl residues by trypsin. By subjecting four of these peptides and the apomyoglobin to automated Edman degradation, over 80% of the primary structure of the protein was obtained. The remainder of the covalent structure was determined by the sequence analysis of peptides that resulted from further digestion of the central cyanogen bromide fragment. This fragment was cleaved at its glutamyl residues with staphylococcal protease and its lysyl residues with trypsin. The action of trypsin was restricted to the lysyl residues by chemical modification of the single arginyl residue of the fragment with 1,2-cyclohexanedione. The primary structure of this myoglobin proved to be identical with that from the Atlantic bottlenosed dolphin and Pacific common dolphin but differs from the myoglobins of the killer whale and pilot whale at two positions. The above sequence identities and differences reflect the close taxonomic relationship of these five species of Cetacea. PMID:454657

  6. Isolation and amino acid sequences of squirrel monkey (Saimiri sciurea) insulin and glucagon.

    PubMed Central

    Yu, J H; Eng, J; Yalow, R S

    1990-01-01

    It was reported two decades ago that insulin was not detectable in the glucose-stimulated state in Saimiri sciurea, the New World squirrel monkey, by a radioimmunoassay system developed with guinea pig anti-pork insulin antibody and labeled pork insulin. With the same system, reasonable levels were observed in rhesus monkeys and chimpanzees. This suggested that New World monkeys, like the New World hystricomorph rodents such as the guinea pig and the coypu, might have insulins whose sequences differ markedly from those of Old World mammals. In this report we describe the purification and amino acid sequences of squirrel monkey insulin and glucagon. We demonstrate that the substitutions at B29, B27, A2, A4, and A17 of squirrel monkey insulin are identical with those previously found in another New World primate, the owl monkey (Aotus trivirgatus). The immunologic cross-reactivity of this insulin in our immunoassay system is only a few percent of that of human insulin. Squirrel monkey glucagon is identical with the usual glucagon found in Old World mammals, which predicts that the glucagons of other New World monkeys would not differ from the usual Old World mammalian glucagon. It appears that the peptides of the New World monkeys have diverged less from those of the Old World mammals than have those of the New World hystricomorph rodents. The striking improvements in peptide purification and sequencing have the potential for adding new information concerning the evolutionary divergence of species. PMID:2263627

  7. Isolation and amino acid sequences of squirrel monkey (Saimiri sciurea) insulin and glucagon

    SciTech Connect

    Yu, Jinghua ); Eng, J.; Yalow, R.S. City Univ. of New York, NY )

    1990-12-01

    It was reported two decades ago that insulin was not detectable in the glucose-stimulated state in Saimiri sciurea, the New World squirrel monkey, by a radioimmunoassay system developed with guinea pig anti-pork insulin antibody and labeled park insulin. With the same system, reasonable levels were observed in rhesus monkeys and chimpanzees. This suggested that New World monkeys, like the New World hystricomorph rodents such as the guinea pig and the coypu, might have insulins whose sequences differ markedly from those of Old World mammals. In this report the authors describe the purification and amino acid sequences of squirrel monkey insulin and glucagon. They demonstrate that the substitutions at B29, B27, A2, A4, and A17 of squirrel monkey insulin are identical with those previously found in another New World primate, the owl monkey (Aotus trivirgatus). The immunologic cross-reactivity of this insulin in their immunoassay system is only a few percent of that of human insulin. It appears that the peptides of the New World monkeys have diverged less from those of the Old World mammals than have those of the New World hystricomorph rodents. The striking improvements in peptide purification and sequencing have the potential for adding new information concerning the evolutionary divergence of species.

  8. Binding site discovery from nucleic acid sequences by discriminative learning of hidden Markov models

    PubMed Central

    Maaskola, Jonas; Rajewsky, Nikolaus

    2014-01-01

    We present a discriminative learning method for pattern discovery of binding sites in nucleic acid sequences based on hidden Markov models. Sets of positive and negative example sequences are mined for sequence motifs whose occurrence frequency varies between the sets. The method offers several objective functions, but we concentrate on mutual information of condition and motif occurrence. We perform a systematic comparison of our method and numerous published motif-finding tools. Our method achieves the highest motif discovery performance, while being faster than most published methods. We present case studies of data from various technologies, including ChIP-Seq, RIP-Chip and PAR-CLIP, of embryonic stem cell transcription factors and of RNA-binding proteins, demonstrating practicality and utility of the method. For the alternative splicing factor RBM10, our analysis finds motifs known to be splicing-relevant. The motif discovery method is implemented in the free software package Discrover. It is applicable to genome- and transcriptome-scale data, makes use of available repeat experiments and aside from binary contrasts also more complex data configurations can be utilized. PMID:25389269

  9. Nucleotide and derived amino acid sequences of the major porin of Comamonas acidovorans and comparison of porin primary structures.

    PubMed Central

    Gerbl-Rieger, S; Peters, J; Kellermann, J; Lottspeich, F; Baumeister, W

    1991-01-01

    The DNA sequence of the gene which codes for the major outer membrane porin (Omp32) of Comamonas acidovorans has been determined. The structural gene encodes a precursor consisting of 351 amino acid residues with a signal peptide of 19 amino acid residues. Comparisons with amino acid sequences of outer membrane proteins and porins from several other members of the class Proteobacteria and of the Chlamydia trachomatis porin and the Neurospora crassa mitochondrial porin revealed a motif of eight regions of local homology. The results of this analysis are discussed with regard to common structural features of porins. PMID:1848840

  10. High-fat diet lowers the nutritional status indicators of pantothenic acid in weaning rats.

    PubMed

    Yoshida, Erina; Fukuwatari, Tsutomu; Ohtsubo, Masako; Shibata, Katsumi

    2010-01-01

    Weaning rats were fed a 5% or 30% fat diet containing limited calcium pantothenate for 28 d. The plasma, liver and adrenal pantothenic acid levels in the rats fed on the 30% fat diet were significantly lower than with the 5% fat diet. The results suggest that the high-fat diet affected pantothenic acid metabolism. PMID:20699566

  11. Sedimentological indicators of paleoenvironments and siliciclastic stratigraphic sequences in some Miocene deposits of the Calvert Cliffs, southern Maryland

    USGS Publications Warehouse

    Shideler, G.L.

    1994-01-01

    Middle Miocene siliciclastic deposits comprising the Calvert Cliffs section at the Baltimore Gas and Electric Company's (BG&E) nuclear power plant site in southern Maryland were analyzed in terms of lithostratigraphy, sedimentary structures, and granulometric parameters, to interprete paleo-environments within a sequence-stratigraphic framework. In terms of sequence-stratigraphic models, the BG&E section can be interpreted as consisting of two genetic stratigraphic sequences (Galloway model), namely, a shelf sequence and an overlying deltaic sequence. Using the Exxon model, the section consists of two third-order (1-5 m.y. duration) depositional sequences. The stratigraphic sequences of the BG&E section reflect both relatively short-term eustatic transgressive events, as well as a long-term regressive trend with associated local deltation and coastal progradation. The regression probably signified a regional basinward shift of depocenters within the Salisbury embayment during Miocene time. -from Author

  12. The Baltic Sea: Geophysical and geochemical properties of Holocene sediment sequences as indicators of past environmental variability

    NASA Astrophysics Data System (ADS)

    Lenz, Conny; Reinholdsson, Maja; Zillén, Lovisa; Conley, Daniel J.; Snowball, Ian

    2010-05-01

    The Baltic Sea has undergone large environmental changes since the retreat of the Weischselian Ice-sheet. In the Late Glacial Period and the early Holocene these changes were most likely caused by natural environmental changes (i.e. changes in the morphology and depths of the Baltic basin and the sills). In more recent time anthropogenic impacts have become more important as a possible and likely cause for changes. During the whole Holocene period climate variability played an important role. However, the relative importance between humans and nature is largely unknown. Here we present the results of a combined geophysical and geochemical study on selected sediment sequences from the Baltic Sea within the two BONUS (Baltic Organisations Network For Funding Science) funded projects HYPER (HYPoxia mitigation for Baltic Sea Ecosystem Restoration) and Baltic GAS (GAS storage and effects of climate change and eutrophication). The over-all aim of these projects is to understand large-scale Baltic Sea ecosystem responses to environmental, climate and anthropogenic forcing. During two Baltic Sea research cruises in 2009 long sediment cores from 8 different locations were recovered. We present preliminary results from one site (LL19) located in the north central Baltic Proper at 169 m water depth. The Littorina Sea sediment record (i.e. the last c. 8000 years) is characterised by alternating periods of homogenised sediments (indicative of oxic conditions) and laminated sediments (indicative of hypoxic/anoxic conditions). Mineral magnetic properties illustrate clear changes between laminated and non-laminated sections of the core. The concentration of ferrimagnetic minerals, as revealed by initial magnetic susceptibility (χ) and saturation isothermal remanent magnetization (SIRM) is variable. The laminated sections in particular show high concentrations and to reveal the origin of the ferrimagnetic signal additional magnetic properties were measured, specifically the

  13. Amino acid sequence analysis and characterization of a ribonuclease from starfish Asterias amurensis.

    PubMed

    Motoyoshi, Naomi; Kobayashi, Hiroko; Itagaki, Tadashi; Inokuchi, Norio

    2016-09-01

    The aim of this study was to phylogenetically characterize the location of the RNase T2 enzyme in the starfish (Asterias amurensis). We isolated an RNase T2 ribonuclease (RNase Aa) from the ovaries of starfish and determined its amino acid sequence by protein chemistry and cloning cDNA encoding RNase Aa. The isolated protein had 231 amino acid residues, a predicted molecular mass of 25,906 Da, and an optimal pH of 5.0. RNase Aa preferentially released guanylic acid from the RNA. The catalytic sites of the RNase T2 family are conserved in RNase Aa; furthermore, the distribution of the cysteine residues in RNase Aa is similar to that in other animal and plant T2 RNases. RNase Aa is cleaved at two points: 21 residues from the N-terminus and 29 residues from the C-terminus; however, both fragments may remain attached to the protein via disulfide bridges, leading to the maintenance of its conformation, as suggested by circular dichroism spectrum analysis. The phylogenetic analysis revealed that starfish RNase Aa is evolutionarily an intermediate between protozoan and oyster RNases. PMID:26920046

  14. Analysis of a nucleotide-binding site of 5-lipoxygenase by affinity labelling: binding characteristics and amino acid sequences.

    PubMed Central

    Zhang, Y Y; Hammarberg, T; Radmark, O; Samuelsson, B; Ng, C F; Funk, C D; Loscalzo, J

    2000-01-01

    5-Lipoxygenase (5LO) catalyses the first two steps in the biosynthesis of leukotrienes, which are inflammatory mediators derived from arachidonic acid. 5LO activity is stimulated by ATP; however, a consensus ATP-binding site or nucleotide-binding site has not been found in its protein sequence. In the present study, affinity and photoaffinity labelling of 5LO with 5'-p-fluorosulphonylbenzoyladenosine (FSBA) and 2-azido-ATP showed that 5LO bound to the ATP analogues quantitatively and specifically and that the incorporation of either analogue inhibited ATP stimulation of 5LO activity. The stoichiometry of the labelling was 1.4 mol of FSBA/mol of 5LO (of which ATP competed with 1 mol/mol) or 0.94 mol of 2-azido-ATP/mol of 5LO (of which ATP competed with 0.77 mol/mol). Labelling with FSBA prevented further labelling with 2-azido-ATP, indicating that the same binding site was occupied by both analogues. Other nucleotides (ADP, AMP, GTP, CTP and UTP) also competed with 2-azido-ATP labelling, suggesting that the site was a general nucleotide-binding site rather than a strict ATP-binding site. Ca(2+), which also stimulates 5LO activity, had no effect on the labelling of the nucleotide-binding site. Digestion with trypsin and peptide sequencing showed that two fragments of 5LO were labelled by 2-azido-ATP. These fragments correspond to residues 73-83 (KYWLNDDWYLK, in single-letter amino acid code) and 193-209 (FMHMFQSSWNDFADFEK) in the 5LO sequence. Trp-75 and Trp-201 in these peptides were modified by the labelling, suggesting that they were immediately adjacent to the C-2 position of the adenine ring of ATP. Given the stoichiometry of the labelling, the two peptide sequences of 5LO were probably near each other in the enzyme's tertiary structure, composing or surrounding the ATP-binding site of 5LO. PMID:11042125

  15. Full Genome Virus Detection in Fecal Samples Using Sensitive Nucleic Acid Preparation, Deep Sequencing, and a Novel Iterative Sequence Classification Algorithm

    PubMed Central

    Cotten, Matthew; Oude Munnink, Bas; Canuti, Marta; Deijs, Martin; Watson, Simon J.; Kellam, Paul; van der Hoek, Lia

    2014-01-01

    We have developed a full genome virus detection process that combines sensitive nucleic acid preparation optimised for virus identification in fecal material with Illumina MiSeq sequencing and a novel post-sequencing virus identification algorithm. Enriched viral nucleic acid was converted to double-stranded DNA and subjected to Illumina MiSeq sequencing. The resulting short reads were processed with a novel iterative Python algorithm SLIM for the identification of sequences with homology to known viruses. De novo assembly was then used to generate full viral genomes. The sensitivity of this process was demonstrated with a set of fecal samples from HIV-1 infected patients. A quantitative assessment of the mammalian, plant, and bacterial virus content of this compartment was generated and the deep sequencing data were sufficient to assembly 12 complete viral genomes from 6 virus families. The method detected high levels of enteropathic viruses that are normally controlled in healthy adults, but may be involved in the pathogenesis of HIV-1 infection and will provide a powerful tool for virus detection and for analyzing changes in the fecal virome associated with HIV-1 progression and pathogenesis. PMID:24695106

  16. Deep Sequencing of HIV-1 RNA and DNA in Newly Diagnosed Patients with Baseline Drug Resistance Showed No Indications for Hidden Resistance and Is Biased by Strong Interference of Hypermutation.

    PubMed

    Dauwe, Kenny; Staelens, Delfien; Vancoillie, Leen; Mortier, Virginie; Verhofstede, Chris

    2016-06-01

    Deep sequencing of plasma RNA or proviral DNA may be an interesting alternative to population sequencing for the detection of baseline transmitted HIV-1 drug resistance. Using a Roche 454 GS Junior HIV-1 prototype kit, we performed deep sequencing of the HIV-1 protease and reverse transcriptase genes on paired plasma and buffy coat samples from newly diagnosed HIV-1-positive individuals. Selection was based on the outcome of population sequencing and included 12 patients with either a revertant amino acid at codon 215 of the reverse transcriptase or a singleton resistance mutation, 4 patients with multiple resistance mutations, and 4 patients with wild-type virus. Deep sequencing of RNA and DNA detected 6 and 43 mutations, respectively, that were not identified by population sequencing. A subsequently performed hypermutation analysis, however, revealed hypermutation in 61.19% of 3,188 DNA reads with a resistance mutation. The removal of hypermutated reads dropped the number of additional mutations in DNA from 43 to 17. No hypermutation evidence was found in the RNA reads. Five of the 6 additional RNA mutations and all additional DNA mutations, after full exclusion of hypermutation bias, were observed in the 3 individuals with multiple resistance mutations detected by population sequencing. Despite focused selection of patients with T215 revertants or singleton mutations, deep sequencing failed to identify the resistant T215Y/F or M184V or any other resistance mutation, indicating that in most of these cases there is no hidden resistance and that the virus detected at diagnosis by population sequencing is the original infecting variant. PMID:27076656

  17. Immunoreactivity of polyclonal antibodies generated against the carboxy terminus of the predicted amino acid sequence of the Huntington disease gene

    SciTech Connect

    Alkatib, G.; Graham, R.; Pelmear-Telenius, A.

    1994-09-01

    A cDNA fragment spanning the 3{prime}-end of the Huntington disease gene (from 8052 to 9252) was cloned into a prokaryotic expression vector containing the E. Coli lac promoter and a portion of the coding sequence for {beta}-galactosidase. The truncated {beta}-galactosidase gene was cleaved with BamHl and fused in frame to the BamHl fragment of the Huntington disease gene 3{prime}-end. Expression analysis of proteins made in E. Coli revealed that 20-30% of the total cellular proteins was represented by the {beta}-galactosidase-huntingtin fusion protein. The identity of the Huntington disease protein amino acid sequences was confirmed by protein sequence analysis. Affinity chromatography was used to purify large quantities of the fusion protein from bacterial cell lysates. Affinity-purified proteins were used to immunize New Zealand white rabbits for antibody production. The generated polyclonal antibodies were used to immunoprecipitate the Huntington disease gene product expressed in a neuroblastoma cell line. In this cell line the antibodies precipitated two protein bands of apparent gel migrations of 200 and 150 kd which together, correspond to the calculated molecular weight of the Huntington disease gene product (350 kd). Immunoblotting experiments revealed the presence of a large precursor protein in the range of 350-750 kd which is in agreement with the predicted molecular weight of the protein without post-translational modifications. These results indicate that the huntingtin protein is cleaved into two subunits in this neuroblastoma cell line and implicate that cleavage of a large precursor protein may contribute to its biological activity. Experiments are ongoing to determine the precursor-product relationship and to examine the synthesis of the huntingtin protein in freshly isolated rat brains, and to determine cellular and subcellular distribution of the gene product.

  18. Functional Analysis and RNA Sequencing Indicate the Regulatory Role of Argonaute1 in Tomato Compound Leaf Development.

    PubMed

    Wang, Tian; Li, Rui; Wen, Liwei; Fu, Daqi; Zhu, Benzhong; Luo, Yunbo; Zhu, Hongliang

    2015-01-01

    Regardless of whether a leaf is simple or compound, the mechanism underlying its development will give rise to a full comprehension of plant morphogenesis. The role of Argonaute1 (AGO1) in the development of simple leaves has been established, but its role in the development of compound leaves remains to be characterized. In this paper, a virus-induced gene silencing (VIGS) strategy was used to dramatically down-regulate the expression of AGO1 ortholog in tomatoes, a model plant for research into compound leaves. AGO1-silenced tomato compound leaves exhibited morphological defects of leaf adaxial-abaxial and trichome development. Analysis of global gene expression profiles indicated that the silencing of AGO1 in tomato compound leaf caused significant changes in the expression of several critical genes, including Auxin Response Factor 4 (ARF4) and Non-expressor of PR5 (NPR5), which were involved in adaxial-abaxial formation and IAA15 that was found to contribute to growth of trichomes as well as Gibberellic Acid Insensitive (GAI) which participated in hormone regulation. Collectively, these results shed light on the complicated mechanism by which AGO1 regulates compound leaf development. PMID:26479258

  19. Evolutionary connections of biological kingdoms based on protein and nucleic acid sequence evidence

    NASA Technical Reports Server (NTRS)

    Dayhoff, M. O.

    1983-01-01

    Prokaryotic and eukaryotic evolutionary trees are developed from protein and nucleic-acid sequences by the methods of numerical taxonomy. Trees are presented for bacterial ferredoxins, 5S ribosomal RNA, c-type cytochromes , cytochromes c2 and c', and 5.8S ribosomal RNA; the implications for early evolution are discussed; and a composite tree showing the branching of the anaerobes, aerobes, archaebacteria, and eukaryotes is shown. Single lines are found for all oxygen-evolving photosynthetic forms and for the salt-loving and high-temperature forms of archaebacteria. It is argued that the eukaryote mitochondria, chloroplasts, and cytoplasmic host material are descended from free-living prokaryotes that formed symbiotic associations, with more than one symbiotic event involved in the evolution of each organelle.

  20. The amino acid alphabet and the architecture of the protein sequence-structure map. I. Binary alphabets.

    PubMed

    Ferrada, Evandro

    2014-12-01

    The correspondence between protein sequences and structures, or sequence-structure map, relates to fundamental aspects of structural, evolutionary and synthetic biology. The specifics of the mapping, such as the fraction of accessible sequences and structures, or the sequences' ability to fold fast, are dictated by the type of interactions between the monomers that compose the sequences. The set of possible interactions between monomers is encapsulated by the potential energy function. In this study, I explore the impact of the relative forces of the potential on the architecture of the sequence-structure map. My observations rely on simple exact models of proteins and random samples of the space of potential energy functions of binary alphabets. I adopt a graph perspective and study the distribution of viable sequences and the structures they produce, as networks of sequences connected by point mutations. I observe that the relative proportion of attractive, neutral and repulsive forces defines types of potentials, that induce sequence-structure maps of vastly different architectures. I characterize the properties underlying these differences and relate them to the structure of the potential. Among these properties are the expected number and relative distribution of sequences associated to specific structures and the diversity of structures as a function of sequence divergence. I study the types of binary potentials observed in natural amino acids and show that there is a strong bias towards only some types of potentials, a bias that seems to characterize the folding code of natural proteins. I discuss implications of these observations for the architecture of the sequence-structure map of natural proteins, the construction of random libraries of peptides, and the early evolution of the natural amino acid alphabet. PMID:25473967

  1. The Amino Acid Alphabet and the Architecture of the Protein Sequence-Structure Map. I. Binary Alphabets

    PubMed Central

    Ferrada, Evandro

    2014-01-01

    The correspondence between protein sequences and structures, or sequence-structure map, relates to fundamental aspects of structural, evolutionary and synthetic biology. The specifics of the mapping, such as the fraction of accessible sequences and structures, or the sequences' ability to fold fast, are dictated by the type of interactions between the monomers that compose the sequences. The set of possible interactions between monomers is encapsulated by the potential energy function. In this study, I explore the impact of the relative forces of the potential on the architecture of the sequence-structure map. My observations rely on simple exact models of proteins and random samples of the space of potential energy functions of binary alphabets. I adopt a graph perspective and study the distribution of viable sequences and the structures they produce, as networks of sequences connected by point mutations. I observe that the relative proportion of attractive, neutral and repulsive forces defines types of potentials, that induce sequence-structure maps of vastly different architectures. I characterize the properties underlying these differences and relate them to the structure of the potential. Among these properties are the expected number and relative distribution of sequences associated to specific structures and the diversity of structures as a function of sequence divergence. I study the types of binary potentials observed in natural amino acids and show that there is a strong bias towards only some types of potentials, a bias that seems to characterize the folding code of natural proteins. I discuss implications of these observations for the architecture of the sequence-structure map of natural proteins, the construction of random libraries of peptides, and the early evolution of the natural amino acid alphabet. PMID:25473967

  2. Trypsin inhibitors from ridged gourd (Luffa acutangula Linn.) seeds: purification, properties, and amino acid sequences.

    PubMed

    Haldar, U C; Saha, S K; Beavis, R C; Sinha, N K

    1996-02-01

    Two trypsin inhibitors, LA-1 and LA-2, have been isolated from ridged gourd (Luffa acutangula Linn.) seeds and purified to homogeneity by gel filtration followed by ion-exchange chromatography. The isoelectric point is at pH 4.55 for LA-1 and at pH 5.85 for LA-2. The Stokes radius of each inhibitor is 11.4 A. The fluorescence emission spectrum of each inhibitor is similar to that of the free tyrosine. The biomolecular rate constant of acrylamide quenching is 1.0 x 10(9) M-1 sec-1 for LA-1 and 0.8 x 10(9) M-1 sec-1 for LA-2 and that of K2HPO4 quenching is 1.6 x 10(11) M-1 sec-1 for LA-1 and 1.2 x 10(11) M-1 sec-1 for LA-2. Analysis of the circular dichroic spectra yields 40% alpha-helix and 60% beta-turn for La-1 and 45% alpha-helix and 55% beta-turn for LA-2. Inhibitors LA-1 and LA-2 consist of 28 and 29 amino acid residues, respectively. They lack threonine, alanine, valine, and tryptophan. Both inhibitors strongly inhibit trypsin by forming enzyme-inhibitor complexes at a molar ratio of unity. A chemical modification study suggests the involvement of arginine of LA-1 and lysine of LA-2 in their reactive sites. The inhibitors are very similar in their amino acid sequences, and show sequence homology with other squash family inhibitors. PMID:8924202

  3. Microfluidic platform for isolating nucleic acid targets using sequence specific hybridization

    PubMed Central

    Wang, Jingjing; Morabito, Kenneth; Tang, Jay X.; Tripathi, Anubhav

    2013-01-01

    The separation of target nucleic acid sequences from biological samples has emerged as a significant process in today's diagnostics and detection strategies. In addition to the possible clinical applications, the fundamental understanding of target and sequence specific hybridization on surface modified magnetic beads is of high value. In this paper, we describe a novel microfluidic platform that utilizes a mobile magnetic field in static microfluidic channels, where single stranded DNA (ssDNA) molecules are isolated via nucleic acid hybridization. We first established efficient isolation of biotinylated capture probe (BP) using streptavidin-coated magnetic beads. Subsequently, we investigated the hybridization of target ssDNA with BP bound to beads and explained these hybridization kinetics using a dual-species kinetic model. The number of hybridized target ssDNA molecules was determined to be about 6.5 times less than that of BP on the bead surface, due to steric hindrance effects. The hybridization of target ssDNA with non-complementary BP bound to bead was also examined, and non-specific hybridization was found to be insignificant. Finally, we demonstrated highly efficient capture and isolation of target ssDNA in the presence of non-target ssDNA, where as low as 1% target ssDNA can be detected from mixture. The microfluidic method described in this paper is significantly relevant and is broadly applicable, especially towards point-of-care biological diagnostic platforms that require binding and separation of known target biomolecules, such as RNA, ssDNA, or protein. PMID:24404041

  4. Establishment of reliable mass spectra and retention indices library: identification of fatty acids in human plasma without authentic standards.

    PubMed

    Zhang, Liangxiao; Tan, Binbin; Zeng, Maomao; Lu, Hongmei; Liang, Yizeng

    2012-01-15

    Gas chromatography mass spectrometry (GC-MS) is routinely employed to analyze small molecules in various samples. The more challenge of GC-MS data processing is to identify the unknown compounds in samples. Mass spectra and retention indices library searching are commonly used method. However, the current libraries are often built through collecting data from different groups. To unknown compounds with similar mass spectra and retention indices (e.g. geometric (cis/trans) isomers), the inaccurate results sometime are supplied. In this case, the costly standard compounds have to be used in every analysis. In this report, taking identification of fatty acids as an example, we proposed a strategy of establishment of special database constructed by equivalent chain length (ECL) values in uniform conditions and mass spectra of fatty acid methyl esters (FAMEs). The mass spectral characteristics were firstly used to identify all expected straight saturated fatty acids, and subsequently calculate the ECL for fatty acids in the sample. Finally, the ECL values of fatty acids in the sample were compared with those of fatty acids in the customized database to identify their structures. The results showed that the method developed in this report could effectively identify similar unknown compounds (FAMEs in the human plasma) after validated by the authentic standards. PMID:22265504

  5. Characterization of N-glycosylation and amino acid sequence features of immunoglobulins from swine.

    PubMed

    Lopez, Paul G; Girard, Lauren; Buist, Marjorie; de Oliveira, Andrey Giovanni Gomes; Bodnar, Edward; Salama, Apolline; Soulillou, Jean-Paul; Perreault, Hélène

    2016-02-01

    The primary goal of this study was to develop a method to study the N-glycosylation of IgG from swine in order to detect epitopes containing N-glycolylneuraminic acid (Neu5Gc) and/or terminal galactose residues linked in α1-3 susceptible to cause xenograft-related problems. Samples of immunoglobulin were isolated from porcine serum using protein-A affinity chromatography. The eluate was then separated on electrophoretic gel, and bands corresponding to the N-glycosylated heavy chains were cut off the gel and subjected to tryptic digestion. Peptides and glycopeptides were separated by reversed phase liquid chromatography and fractions were collected for matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOF-MS) analysis. Overall no α1-3 galactose was detected, as demonstrated by complete susceptibility of terminal galactose residues to β-galactosidase digestion. Neu5Gc was detected on singly sialylated structures. Two major N-glycopeptides were found, EEQFNSTYR and EAQFNSTYR as determined by tandem MS (MS/MS), as previously reported by Butler et al. (Immunogenetics, 61, 2009, 209-230), who found 11 subclasses for porcine IgG. Out of the 11, ten include the sequence corresponding to EEQFNSTYR, and only one codes for EAQFNSTYR. In this study, glycosylation patterns associated with both chains were slightly different, in that EEQFNSTYR had a higher content of galactose. The last step of this study consisted of peptide-mapping the 11 reported porcine IgG sequences. Although there was considerable overlap, at least one unique tryptic peptide was found per IgG sequence. The workflow presented in this manuscript constitutes the first study to use MALDI-TOF-MS in the investigation of porcine IgG structural features. PMID:26586247

  6. Human Retroviruses and AIDS. A compilation and analysis of nucleic acid and amino acid sequences: I--II; III--V

    SciTech Connect

    Myers, G.; Korber, B.; Wain-Hobson, S.; Smith, R.F.; Pavlakis, G.N.

    1993-12-31

    This compendium and the accompanying floppy diskettes are the result of an effort to compile and rapidly publish all relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses. The scope of the compendium and database is best summarized by the five parts that it comprises: (I) HIV and SIV Nucleotide Sequences; (II) Amino Acid Sequences; (III) Analyses; (IV) Related Sequences; and (V) Database Communications. Information within all the parts is updated at least twice in each year, which accounts for the modes of binding and pagination in the compendium.

  7. Methylmalonic Acid and Homocysteine as Indicators of Vitamin B-12 Deficiency in Cancer

    PubMed Central

    Vashi, Pankaj; Edwin, Persis; Popiel, Brenten; Lammersfeld, Carolyn; Gupta, Digant

    2016-01-01

    Background/Aims Normal or high serum vitamin B-12 levels can sometimes be seen in a B-12 deficient state, and can therefore be misleading. High levels of Methymalonic Acid (MMA) and Homocysteine (HC) have been identified as better indicators of B-12 deficiency than the actual serum B-12 level itself. We evaluated the prevalence of vitamin B-12 deficiency using appropriate cut-off levels of vitamin B-12, MMA and HC, and determined the relationship between serum levels of vitamin B-12, MMA and HC in cancer. Methods This is a cross-sectional study using a consecutive case series of 316 cancer patients first seen at Cancer Treatment Centers of America® (CTCA) at Midwestern Regional Medical Center between April 2014 and June 2014. All patients were evaluated at baseline for vitamin B-12 (pg/mL), MMA (nmol/L) and HC (μmol/L) levels. In accordance with previously published research, the following cut-offs were used to define vitamin B-12 deficiency: <300 pg/mL for vitamin B-12, >260 nmol/L for MMA and >12 μmol/L for HC. The relationship between B-12, MMA and HC was evaluated using Spearman's rho correlation coefficient and cross-tabulation analysis. Receiver Operating Characteristic (ROC) curves were estimated using the non-parametric method to further evaluate the diagnostic accuracy of vitamin B-12 using Fedosov quotient as the "gold standard". Results Mean age at presentation was 52.5 years. 134 (42.4%) patients were males while 182 (57.6%) were females. Median vitamin B-12, MMA and HC levels were 582.5 pg/mL, 146.5 nmol/L and 8.4 μmol/L respectively. Of 316 patients, 28 (8.9%) were vitamin B-12 deficient based on vitamin B-12 (<300pg/mL), 34 (10.8%) were deficient based on MMA (>260 nmol/L) while 55 (17.4%) were deficient based on HC (>12 μmol/L). Correlation analysis revealed a significant weak negative correlation between vitamin B-12 and MMA (rho = -0.22) as well as B-12 and HC (rho = -0.35). ROC curves suggested MMA to have the best discriminatory power in

  8. Lactic acid production from potato peel waste by anaerobic sequencing batch fermentation using undefined mixed culture.

    PubMed

    Liang, Shaobo; McDonald, Armando G; Coats, Erik R

    2015-11-01

    Lactic acid (LA) is a necessary industrial feedstock for producing the bioplastic, polylactic acid (PLA), which is currently produced by pure culture fermentation of food carbohydrates. This work presents an alternative to produce LA from potato peel waste (PPW) by anaerobic fermentation in a sequencing batch reactor (SBR) inoculated with undefined mixed culture from a municipal wastewater treatment plant. A statistical design of experiments approach was employed using set of 0.8L SBRs using gelatinized PPW at a solids content range from 30 to 50 g L(-1), solids retention time of 2-4 days for yield and productivity optimization. The maximum LA production yield of 0.25 g g(-1) PPW and highest productivity of 125 mg g(-1) d(-1) were achieved. A scale-up SBR trial using neat gelatinized PPW (at 80 g L(-1) solids content) at the 3 L scale was employed and the highest LA yield of 0.14 g g(-1) PPW and a productivity of 138 mg g(-1) d(-1) were achieved with a 1 d SRT. PMID:25708409

  9. Bacterial community compositions in sediment polluted by perfluoroalkyl acids (PFAAs) using Illumina high-throughput sequencing.

    PubMed

    Sun, Yajun; Wang, Tieyu; Peng, Xiawei; Wang, Pei; Lu, Yonglong

    2016-06-01

    The characterization of bacterial community compositions and the change in perfluoroalkyl acids (PFAAs) along a natural river distribution system were explored in the present study. Illumina high-throughput sequencing was used to explore bacterial community diversity and structure in sediment polluted by PFAAs from the Xiaoqing River, the area with concentrated fluorochemical facilities in China. The concentration of PFAAs was in the range of 8.44-465.60 ng/g dry weight (dw) in sediment. Perfluorooctanoic acid (PFOA) was the dominant PFAA in all samples, which accounted for 94.2 % of total PFAAs. High-level PFOA could lead to an obvious increase in relative abundance of Proteobacteria, ε-Proteobacteria, Thiobacillus, and Sulfurimonas and the decrease in relative abundance of other bacteria. Redundancy analysis revealed that PFOA played an important role in the formation of bacterial community, and PFOA at higher concentration could reduce the diversity of bacterial community. When the concentration of PFOA was below 100 ng/g dw in sediment, no significant effect on microbial community structure was observed. Thiobacillus and Sulfurimonas were positively correlated with the concentration of PFOA, suggesting that both genera were resistant to PFOA contamination. PMID:26780047

  10. Mass spectrometric detection of the amino acid sequence polymorphism of the hepatitis C virus antigen.

    PubMed

    Kaysheva, A L; Ivanov, Yu D; Frantsuzov, P A; Krohin, N V; Pavlova, T I; Uchaikin, V F; Konev, V А; Kovalev, O B; Ziborov, V S; Archakov, A I

    2016-03-01

    A method for detection and identification of the hepatitis C virus antigen (HCVcoreAg) in human serum with consideration for possible amino acid substitutions is proposed. The method is based on a combination of biospecific capturing and concentrating of the target protein on the surface of the chip for atomic force microscope (AFM chip) with subsequent protein identification by tandem mass spectrometric (MS/MS) analysis. Biospecific AFM-capturing of viral particles containing HCVcoreAg from serum samples was performed by use of AFM chips with monoclonal antibodies (anti-HCVcore) covalently immobilized on the surface. Biospecific complexes were registered and counted by AFM. Further MS/MS analysis allowed to reliably identify the HCVcoreAg in the complexes formed on the AFM chip surface. Analysis of MS/MS spectra, with the account taken of the possible polymorphisms in the amino acid sequence of the HCVcoreAg, enabled us to increase the number of identified peptides. PMID:26773170

  11. {Delta}-Aminolevulinic acid dehydratase: A sensitive indicator of lead exposure in broiler chicks: (Gallus domesticus)

    SciTech Connect

    Bakalli, R.I.; Pesti, G.M.; Konjufca, V.

    1995-12-01

    Delta-aminolevulinic acid dehydratase, EC 4.2.1.24 (ALAD) is one of the enzymes participating in heme synthesis. The study reported in this paper was designed to determine the activity of erythrocyte ALAD anbd the relationship between this enzyme and tissue lead levels in chickens, during Pb intake and after withdrawing Bv from the feed. 20 refs., 3 tabs.

  12. 76 FR 81363 - Temperature-Indicating Devices; Thermally Processed Low-Acid Foods Packaged in Hermetically...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-28

    ... (76 FR 11892). The final rule amended FDA's regulations for thermally processed low-acid foods..., 5100 Paint Branch Pkwy., College Park, MD 20740, (240) 205-1165. SUPPLEMENTARY INFORMATION: In FR Doc... HUMAN SERVICES Food and Drug Administration 21 CFR Part 113 (formerly 2007N-0026)...

  13. FATTY ACID STABLE ISOTOPE INDICATORS OF MICROBIAL CARBON SOURCE IN TROPICAL SOILS

    EPA Science Inventory

    The soil microbial community plays an important role in tropical ecosystem functioning because of its importance in the soil organic matter (SOM) cycle. We have measured the stable carbon isotopic ratio (delta13C) of individual phospholipid fatty acids (PLFAs) in a variety of tr...

  14. Peptide sequencing by using a combination of partial acid hydrolysis and fast-atom-bombardment mass spectrometry.

    PubMed Central

    De Angelis, F; Botta, M; Ceccarelli, S; Nicoletti, R

    1986-01-01

    To overcome the limit of the intensity of ions carrying sequence information in structural determinations of peptides by fast-atom-bombardment m.s., we have developed a method that consists in taking spectra of the peptide acid hydrolysates at different hydrolysis times. Peaks correspond to the oligomers arising from the peptide partial hydrolysis. The sequence can then be identified from the structurally overlapping fragments. PMID:2428356

  15. Canine preprorelaxin: nucleic acid sequence and localization within the canine placenta.

    PubMed

    Klonisch, T; Hombach-Klonisch, S; Froehlich, C; Kauffold, J; Steger, K; Steinetz, B G; Fischer, B

    1999-03-01

    Employing uteroplacental tissue at Day 35 of gestation, we determined the nucleic acid sequence of canine preprorelaxin using reverse transcription- and rapid amplification of cDNA ends-polymerase chain reaction. Canine preprorelaxin cDNA consisted of 534 base pairs encoding a protein of 177 amino acids with a signal peptide of 25 amino acids (aa), a B domain of 35 aa, a C domain of 93 aa, and an A domain of 24 aa. The putative receptor binding region in the N'-terminal part of the canine relaxin B domain GRDYVR contained two substitutions from the classical motif (E-->D and L-->Y). Canine preprorelaxin shared highest homology with porcine and equine preprorelaxin. Northern analysis revealed a 1-kilobase transcript present in total RNA of canine uteroplacental tissue but not of kidney tissue. Uteroplacental tissue from two bitches each at Days 30 and 35 of gestation were studied by in situ hybridization to localize relaxin mRNA. Immunohistochemistry for relaxin, cytokeratin, vimentin, and von Willebrand factor was performed on uteroplacental tissue at Day 30 of gestation. The basal cell layer at the core of the chorionic villi was devoid of relaxin mRNA and immunoreactive relaxin or vimentin but was immunopositive for cytokeratin and identified as cytotrophoblast cells. The cell layer surrounding the chorionic villi displayed specific hybridization signals for relaxin mRNA and immunoreactivity for relaxin and cytokeratin but not for vimentin, and was identified as syncytiotrophoblast. Those areas of the chorioallantoic tissue with most intense relaxin immunoreactivity were highly vascularized as demonstrated by immunoreactive von Willebrand factor expressed on vascular endothelium. The uterine glands and nonplacental uterine areas of the canine zonary girdle placenta were devoid of relaxin mRNA and relaxin. We conclude that the syncytiotrophoblast is the source of relaxin in the canine placenta. PMID:10026098

  16. Sequence-Specific Electrical Purification of Nucleic Acids with Nanoporous Gold Electrodes.

    PubMed

    Daggumati, Pallavi; Appelt, Sandra; Matharu, Zimple; Marco, Maria L; Seker, Erkin

    2016-06-22

    Nucleic-acid-based biosensors have enabled rapid and sensitive detection of pathogenic targets; however, these devices often require purified nucleic acids for analysis since the constituents of complex biological fluids adversely affect sensor performance. This purification step is typically performed outside the device, thereby increasing sample-to-answer time and introducing contaminants. We report a novel approach using a multifunctional matrix, nanoporous gold (np-Au), which enables both detection of specific target sequences in a complex biological sample and their subsequent purification. The np-Au electrodes modified with 26-mer DNA probes (via thiol-gold chemistry) enabled sensitive detection and capture of complementary DNA targets in the presence of complex media (fetal bovine serum) and other interfering DNA fragments in the range of 50-1500 base pairs. Upon capture, the noncomplementary DNA fragments and serum constituents of varying sizes were washed away. Finally, the surface-bound DNA-DNA hybrids were released by electrochemically cleaving the thiol-gold linkage, and the hybrids were iontophoretically eluted from the nanoporous matrix. The optical and electrophoretic characterization of the analytes before and after the detection-purification process revealed that low target DNA concentrations (80 pg/μL) can be successfully detected in complex biological fluids and subsequently released to yield pure hybrids free of polydisperse digested DNA fragments and serum biomolecules. Taken together, this multifunctional platform is expected to enable seamless integration of detection and purification of nucleic acid biomarkers of pathogens and diseases in miniaturized diagnostic devices. PMID:27244455

  17. Homology analyses of the protein sequences of fatty acid synthases from chicken liver, rat mammary gland, and yeast

    SciTech Connect

    Chang, Soo-Ik ); Hammes, G.G. )

    1989-11-01

    Homology analyses of the protein sequences of chicken liver and rat mammary gland fatty acid synthases were carried out. The amino acid sequences of the chicken and rat enzymes are 67% identical. If conservative substitutions are allowed, 78% of the amino acids are matched. A region of low homologies exists between the functional domains, in particular around amino acid residues 1059-1264 of the chicken enzyme. Homologies between the active sites of chicken and rat and of chicken and yeast enzymes have been analyzed by an alignment method. A high degree of homology exists between the active sites of the chicken and rat enzymes. However, the chicken and yeast enzymes show a lower degree of homology. The DADPH-binding dinucleotide folds of the {beta}-ketoacyl reductase and the enoyl reductase sites were identified by comparison with a known consensus sequence for the DADP- and FAD-binding dinucleotide folds. The active sites of all of the enzymes are primarily in hydrophobic regions of the protein. This study suggests that the genes for the functional domains of fatty acid synthase were originally separated, and these genes were connected to each other by using different connecting nucleotide sequences in different species. An alternative explanation for the differences in rat and chicken is a common ancestry and mutations in the joining regions during evolution.

  18. Complete mitochondrial control region sequences indicate a distinct variety of brown trout Salmo trutta in the Aral Sea.

    PubMed

    Griffiths, A M; Bright, D; Stevens, J R

    2009-04-01

    Complete sequencing of the mtDNA control region (CR) from five specimens of brown trout Salmo trutta from the Amu Darya River identified two novel haplotypes belonging to the Danubian lineage. This finding supports the long-standing hypothesis that brown trout in the Aral Sea represent a distinct genetic stock and also illustrates the benefits that complete sequencing of the CR can provide for elucidating phylogeographic relationships. PMID:20735624

  19. Enantioselective separation of amino acids as biomarkers indicating life in extraterrestrial environments.

    PubMed

    Pietrogrande, Maria Chiara

    2013-10-01

    Traces of prebiotic amino acids, i.e., the building blocks of proteins, are excellent biomarkers that could provide evidence of extinct or extant life in extra-terrestrial environments. In particular, characterization of the enantiomeric excess of amino acids gives relevant information about the biotic or abiotic origin of molecules, because it is generally assumed that life elsewhere could be based on either L or D amino acids, but not both. The analytical procedures used in in-situ space missions for chiral discrimination of amino acids must meet severe requirements imposed by flight conditions: short analysis time, low energy consumption, robustness, storage for long periods under extreme conditions, high efficiency and sensitivity, automation, and remote-control operation. Such methods are based on gas chromatography, high-pressure liquid chromatography, and capillary electrophoresis, usually coupled with mass spectrometry; of these, gas chromatography-mass spectrometry (GC-MS) is the only such combination yet used in space missions. Preliminary in-situ sample derivatization is required before GC-MS analysis to convert amino acids into volatile and thermally stable compounds. The silylation reagent most commonly used, N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide, is unsuitable for detection of homochirality, and alternative derivatization techniques have been developed that preserve the stereochemical configuration of the original compounds and are compatible with spaceflight conditions. These include the reagent N,N-dimethylformamide dimethylacetal, which has already been used in the Rosetta mission, a mixture of alkyl chloroformate, ethanol, and pyridine, a mixture of perfluorinated anhydrides and perfluoro alcohols, and hexafluoroacetone, the first gaseous derivatizing agent. In all the space instruments, solvent extraction of organic matter and chemical derivatization have been combined in a single automatic and remote-controlled procedure in a

  20. Molecular cloning of the. alpha. -subunit of human prolyl 4-hydroxylase: The complete cDNA-derived amino acid sequence and evidence for alternative splicing of RNA transcripts

    SciTech Connect

    Helaakoski, T.; Vuori, K.; Myllylae, R.; Kivirikko, K.I.; Pihlajaniemi, T. )

    1989-06-01

    Prolyl 4-hydroxylase an {alpha}{sub 2}{beta}{sub 2} tetramer, catalyzes the formation of 4-hydroxyproline in collagens by the hydroxylation of proline residues in peptide linkages. The authors report here on the isolation of cDNA clones encoding the {alpha}-subunit of the enzyme from human tumor HT-1080, placenta, and fibroblast cDNA libraries. Eight overlapping clones covering almost all of the corresponding 3,000-nucleotide mRNA, including all the coding sequences, were characterized. These clones encode a polypeptide of 517 amino acid residues and a signal peptide of 17 amino acids. Previous characterization of cDNA clones for the {beta}-subunit of prolyl 4-hydroxylase has indicated that its C terminus has the amino acid sequence Lys-Asp-Gly-Leu, which, it has been suggested, is necessary for the retention of a polypeptide within the lumen of the endoplasmic reticulum. The {alpha}-subunit does not have this C-terminal sequence, and thus one function of the {beta}-subunit in the prolyl 4-hydroxylase tetramer appears to be to retain the enzyme within this cell organelle. Southern blot analyses of human genomic DNA with a cDNA probe for the {alpha}-subunit suggested the presence of only one gene encoding the two types of mRNA, which appear to result from mutually exclusive alternative splicing of primary transcripts of one gene.

  1. Cleavage of nucleic acids

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor L.; Brow, Mary Ann D.; Dahlberg, James E.

    2007-12-11

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  2. Cleavage of nucleic acids

    SciTech Connect

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow; Mary Ann D.; Dahlberg, James E.

    2010-11-09

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  3. Cleavage of nucleic acids

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann D.; Dahlberg, James E.

    2000-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  4. Nucleic acid detection assays

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann; Dahlberg, James E.

    2005-04-05

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  5. Nucleic acid detection compositions

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann; Dahlberg, James L.

    2008-08-05

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  6. Complete amino acid sequence of the medium-chain S-acyl fatty acid synthetase thio ester hydrolase from rat mammary gland

    SciTech Connect

    Randhawa, Z.I.; Smith, S.

    1987-03-10

    The complete amino acid sequence of the medium-chain S-acyl fatty acid synthetase thio ester hydrolase (thioesterase II) from rat mammary gland is presented. Most of the sequence was derived by analysis of (/sup 14/C)-labelled peptide fragments produced by cleavage at methionyl, glutamyl, lysyl, arginyl, and tryptophanyl residues. A small section of the sequence was deduced from a previously analyzed cDNA clone. The protein consists of 260 residues and has a blocked amino-terminal methionine and calculated M/sub r/ of 29,212. The carboxy-terminal sequence, verified by Edman degradation of the carboxy-terminal cyanogen bromide fragment and carboxypeptidase Y digestion of the intact thioesterase II, terminates with a serine residue and lacks three additional residues predicted by the cDNA sequence. The native enzyme contains three cysteine residues but no disulfide bridges. The active site serine residue is located at position 101. The rat mammary gland thioesterase II exhibits approximately 40% homology with a thioesterase from mallard uropygial gland, the sequence of which was recently determined by cDNA analysis. Thus the two enzymes may share similar structural features and a common evolutionary origin. The location of the active site in these thioesterases differs from that of other serine active site esterases; indeed, the enzymes do not exhibit any significant homology with other serine esterases, suggesting that they may constitute a separate new family of serine active site enzymes.

  7. Amino acid sequence of the ligand-binding domain of the aryl hydrocarbon receptor 1 predicts sensitivity of wild birds to effects of dioxin-like compounds.

    PubMed

    Farmahin, Reza; Manning, Gillian E; Crump, Doug; Wu, Dongmei; Mundy, Lukas J; Jones, Stephanie P; Hahn, Mark E; Karchner, Sibel I; Giesy, John P; Bursian, Steven J; Zwiernik, Matthew J; Fredricks, Timothy B; Kennedy, Sean W

    2013-01-01

    The sensitivity of avian species to the toxic effects of dioxin-like compounds (DLCs) varies up to 1000-fold among species, and this variability has been associated with interspecies differences in aryl hydrocarbon receptor 1 ligand-binding domain (AHR1 LBD) sequence. We previously showed that LD(50) values, based on in ovo exposures to DLCs, were significantly correlated with in vitro EC(50) values obtained with a luciferase reporter gene (LRG) assay that measures AHR1-mediated induction of cytochrome P4501A in COS-7 cells transfected with avian AHR1 constructs. Those findings suggest that the AHR1 LBD sequence and the LRG assay can be used to predict avian species sensitivity to DLCs. In the present study, the AHR1 LBD sequences of 86 avian species were studied, and differences at amino acid sites 256, 257, 297, 324, 337, and 380 were identified. Site-directed mutagenesis, the LRG assay, and homology modeling highlighted the importance of each amino acid site in AHR1 sensitivity to 2,3,7,8-tetrachlorodibenzo-p-dioxin and other DLCs. The results of the study revealed that (1) only amino acids at sites 324 and 380 affect the sensitivity of AHR1 expression constructs of the 86 avian species to DLCs and (2) in vitro luciferase activity of AHR1 constructs containing only the LBD of the species of interest is significantly correlated (r (2) = 0.93, p < 0.0001) with in ovo toxicity data for those species. These results indicate promise for the use of AHR1 LBD amino acid sequences independently, or combined with the LRG assay, to predict avian species sensitivity to DLCs. PMID:22923492

  8. The complete amino acid sequence of the A-chain of human plasma alpha 2HS-glycoprotein.

    PubMed

    Yoshioka, Y; Gejyo, F; Marti, T; Rickli, E E; Bürgi, W; Offner, G D; Troxler, R F; Schmid, K

    1986-02-01

    Normal human plasma alpha 2HS-glycoprotein has earlier been shown to be comprised of two polypeptide chains. Recently, the amino acid and carbohydrate sequences of the short chain were elucidated (Gejyo, F., Chang, J.-L., Bürgi, W., Schmid, K., Offner, G. D., Troxler, R.F., van Halbeck, H., Dorland, L., Gerwig, G. J., and Vliegenthart, J.F.G. (1983) J. Biol. Chem. 258, 4966-4971). In the present study, the amino acid sequence of the long chain of this protein, designated A-chain, was determined and found to consist of 282 amino acid residues. Twenty-four amino acid doublets were found; the most abundant of these are Pro-Pro and Ala-Ala which each occur five times. Of particular interest is the presence of three Gly-X-Pro and one Gly-Pro-X sequences that are characteristic of the repeating sequences of collagens. Chou-Fasman evaluation of the secondary structure suggested that the A-chain contains 29% alpha-helix, 24% beta-pleated sheet, and 26% reverse turns and, thus, approximately 80% of the polypeptide chain may display ordered structure. Four glycosylation sites were identified. The two N-glycosidic oligosaccharides were found in the center region (residues 138 and 158), whereas the two O-glycosidic heterosaccharides, both linked to threonine (residues 238 and 252), occur within the carboxyl-terminal region. The N-glycans are linked to Asn residues in beta-turns, while the O-glycans are located in short random segments. Comparison of the sequence of the amino- and carboxyl-terminal 30 residues with protein sequences in a data bank demonstrated that the A-chain is not significantly related to any known proteins. However, the proline-rich carboxyl-terminal region of the A-chain displays some sequence similarity to collagens and the collagen-like domains of complement subcomponent C1q. PMID:3944104

  9. Omega-3 fatty acid deficiency selectively up-regulates delta6-desaturase expression and activity indices in rat liver: prevention by normalization of omega-3 fatty acid status.

    PubMed

    Hofacer, Rylon; Jandacek, Ronald; Rider, Therese; Tso, Patrick; Magrisso, I Jack; Benoit, Stephen C; McNamara, Robert K

    2011-09-01

    This study investigated the effects of perinatal dietary omega-3 (n-3) fatty acid depletion and subsequent repletion on the expression of genes that regulate long-chain (LC) polyunsaturated fatty acid biosynthesis in rat liver and brain. It was hypothesized that chronic n-3 fatty acid deficiency would increase liver Fads1 and Fads2 messenger RNA (mRNA) expression/activity and that n-3 fatty acid repletion would normalize this response. Adult rats fed the n-3-free diet during perinatal development exhibited significantly lower erythrocyte, liver, and frontal cortex LCn-3 fatty acid composition and reciprocal elevations in LC omega-6 (n-6) fatty acid composition compared with controls (CONs) and repleted rats. Liver Fads2, but not Fads1, Elovl2, or Elovl5, mRNA expression was significantly greater in n-3-deficient (DEF) rats compared with CONs and was partially normalized in repleted rats. The liver 18:3n-6/18:2n-6 ratio, an index of delta6-desturase activity, was significantly greater in DEF rats compared with CON and repleted rats and was positively correlated with Fads2 mRNA expression among all rats. The liver 18:3n-6/18:2n-6 ratio, but not Fads2 mRNA expression, was also positively correlated with erythrocyte and frontal cortex LCn-6 fatty acid compositions. Neither Fads1 or Fads2 mRNA expression was altered in brain cortex of DEF rats. These results confirm previous findings that liver, but not brain, delta6-desaturase expression and activity indices are negatively regulated by dietary n-3 fatty acids. PMID:22024496

  10. Effect of acidity on the polarization sensitivity of azo-indicator based recording media*

    NASA Astrophysics Data System (ADS)

    Shaverdova, V. G.; Petrova, S. S.; Purtseladze, A. L.; Tarasashvili, V. I.; Obolashvili, N. Z.

    2013-01-01

    This is an experimental study of the photoanisotropic gyrotropic properties of recording media based on azoindicators — homologs (five dyes) of methyl orange-- introduced into the polymer matrix. Samples were prepared by a technology we have developed employing solvents with different acidities (pH 1.68-12.48). The samples were exposed to actinic radiation (λ = 488 nm) from an argon laser, and the photoinduced anisotropy measured in real time. The circular dichroism and circular birefringence in the layers under study are calculated for a neutral medium and at different pH levels.

  11. Indices of fatty acid desaturase activity in healthy human subjects: effects of different types of dietary fat.

    PubMed

    Vessby, Bengt; Gustafsson, Inga-Britt; Tengblad, Siv; Berglund, Lars

    2013-09-14

    Δ9-Desaturase (stearoyl-CoA desaturase 1, SCD-1) regulates the desaturation of SFA, mainly stearic and palmitic, to MUFA. Δ6-Desaturase (D6D) and Δ5-desaturase (D5D) are involved in the metabolism of linoleic and α-linolenic acid to polyunsaturated metabolites. The objective of the present study was to study the effects of different types of dietary fat on indices of fatty acid desaturase (FADS) activity (evaluated as product:precursor ratios) in plasma and skeletal muscle in human subjects. A high SCD-1 index has been related to obesity and metabolic disorders, while the D5D index is associated with insulin sensitivity. Fatty acid composition of serum and skeletal muscle lipids was analysed by GLC during a randomised, controlled, 3-month dietary intervention in healthy subjects. A comparison of the effects of a diet containing butter fat (SFA, n 17) with a diet containing monounsaturated fat (MUFA, n 17), keeping all other dietary components constant, showed a reduced SCD-1 activity index by 20% on the MUFA diet compared with the SFA diet assessed in serum cholesteryl esters. The D6D and D5D indices remained unaffected. Supplementation with long-chain n-3 fatty acids reduced the SCD-1 index by a similar magnitude while the D6D index decreased and the D5D index increased. It is concluded that changes in the type of fat in the diet affect the indices of FADS activity in serum and skeletal muscle in human subjects. The desaturase activity indices estimated from the serum lipid ester composition are significantly related to corresponding indices studied in skeletal muscle phospholipids. PMID:23414551

  12. Method for the detection of specific nucleic acid sequences by polymerase nucleotide incorporation

    DOEpatents

    Castro, Alonso

    2004-06-01

    A method for rapid and efficient detection of a target DNA or RNA sequence is provided. A primer having a 3'-hydroxyl group at one end and having a sequence of nucleotides sufficiently homologous with an identifying sequence of nucleotides in the target DNA is selected. The primer is hybridized to the identifying sequence of nucleotides on the DNA or RNA sequence and a reporter molecule is synthesized on the target sequence by progressively binding complementary nucleotides to the primer, where the complementary nucleotides include nucleotides labeled with a fluorophore. Fluorescence emitted by fluorophores on single reporter molecules is detected to identify the target DNA or RNA sequence.

  13. A systematic study of fundamentals in α-helical coiled coil mimicry by alternating sequences of β- and γ-amino acids.

    PubMed

    Rezaei Araghi, Raheleh; Baldauf, Carsten; Gerling, Ulla I M; Cadicamo, Cosimo Damiano; Koksch, Beate

    2011-08-01

    Aimed at understanding the crucially important structural features for the integrity of α-helical mimicry by βγ-sequences, an α-amino acid sequence in a native peptide was substituted by differently arranged βγ-sequences. The self- and hetero-assembly of a series of αβγ-chimeric sequences based on a 33-residue GCN4-derived peptide was investigated by means of molecular dynamics, circular dichroism, and a disulfide exchange assay. Despite the native-like behavior of βγ alternating sequences such as retention of α-helix dipole and the formation of 13-membered α-helix turns, the αβγ-chimeras with different βγ substitution patterns do not equally mimic the structural behavior of the native parent peptide in solution. The preservation of the key residue contacts such as van der Waals interactions and intrahelical H-bonding, which can be met only by particular substitution patterns, thermodynamically favor the adoption of coiled coil folding motif. In this study, we show how successfully the destabilizing structural consequences of α → βγ modification can be harnessed by reducing the solvent-exposed hydrophobic surface area and placing of suitably long and bulky helix-forming side chains at the hydrophobic core. The pairing of αβγ-chimeric sequences with the native wild-type are thermodynamically allowed in the case of ideal arrangement of β- and γ-residues. This indicates a similarity in local side chain packing of β- and γ-amino acids at the helical interface of αβγ-chimeras and the native α-peptide. Consequently, the backbone extended residues are able to participate in classical "knob-into-hole" packing with native α-peptide. PMID:21638022

  14. Genome Sequence of a Candidate World Health Organization Reference Strain of Zika Virus for Nucleic Acid Testing

    PubMed Central

    Trösemeier, Jan-Hendrik; Musso, Didier; Blümel, Johannes; Thézé, Julien; Pybus, Oliver G.

    2016-01-01

    We report here the sequence of a candidate reference strain of Zika virus (ZIKV) developed on behalf of the World Health Organization (WHO). The ZIKV reference strain is intended for use in nucleic acid amplification (NAT)-based assays for the detection and quantification of ZIKV RNA. PMID:27587826

  15. Genome Sequence of Schizochytrium sp. CCTCC M209059, an Effective Producer of Docosahexaenoic Acid-Rich Lipids

    PubMed Central

    Ji, Xiao-Jun; Mo, Kai-Qiang; Ren, Lu-Jing; Li, Gan-Lu; Huang, Jian-Zhong

    2015-01-01

    Schizochytrium is an effective species for producing omega-3 docosahexaenoic acid (DHA). Here, we report a genome sequence of Schizochytrium sp. CCTCC M209059, which has a genome size of 39.09 Mb. It will provide the genomic basis for further insights into the metabolic and regulatory mechanisms underlying the DHA formation. PMID:26251485

  16. Evolutionary Distance of Amino Acid Sequence Orthologs across Macaque Subspecies: Identifying Candidate Genes for SIV Resistance in Chinese Rhesus Macaques

    PubMed Central

    Ross, Cody T.; Roodgar, Morteza; Smith, David Glenn

    2015-01-01

    We use the Reciprocal Smallest Distance (RSD) algorithm to identify amino acid sequence orthologs in the Chinese and Indian rhesus macaque draft sequences and estimate the evolutionary distance between such orthologs. We then use GOanna to map gene function annotations and human gene identifiers to the rhesus macaque amino acid sequences. We conclude methodologically by cross-tabulating a list of amino acid orthologs with large divergence scores with a list of genes known to be involved in SIV or HIV pathogenesis. We find that many of the amino acid sequences with large evolutionary divergence scores, as calculated by the RSD algorithm, have been shown to be related to HIV pathogenesis in previous laboratory studies. Four of the strongest candidate genes for SIVmac resistance in Chinese rhesus macaques identified in this study are CDK9, CXCL12, TRIM21, and TRIM32. Additionally, ANKRD30A, CTSZ, GORASP2, GTF2H1, IL13RA1, MUC16, NMDAR1, Notch1, NT5M, PDCD5, RAD50, and TM9SF2 were identified as possible candidates, among others. We failed to find many laboratory experiments contrasting the effects of Indian and Chinese orthologs at these sites on SIVmac pathogenesis, but future comparative studies might hold fertile ground for research into the biological mechanisms underlying innate resistance to SIVmac in Chinese rhesus macaques. PMID:25884674

  17. Evolutionary distance of amino acid sequence orthologs across macaque subspecies: identifying candidate genes for SIV resistance in Chinese rhesus macaques.

    PubMed

    Ross, Cody T; Roodgar, Morteza; Smith, David Glenn

    2015-01-01

    We use the Reciprocal Smallest Distance (RSD) algorithm to identify amino acid sequence orthologs in the Chinese and Indian rhesus macaque draft sequences and estimate the evolutionary distance between such orthologs. We then use GOanna to map gene function annotations and human gene identifiers to the rhesus macaque amino acid sequences. We conclude methodologically by cross-tabulating a list of amino acid orthologs with large divergence scores with a list of genes known to be involved in SIV or HIV pathogenesis. We find that many of the amino acid sequences with large evolutionary divergence scores, as calculated by the RSD algorithm, have been shown to be related to HIV pathogenesis in previous laboratory studies. Four of the strongest candidate genes for SIVmac resistance in Chinese rhesus macaques identified in this study are CDK9, CXCL12, TRIM21, and TRIM32. Additionally, ANKRD30A, CTSZ, GORASP2, GTF2H1, IL13RA1, MUC16, NMDAR1, Notch1, NT5M, PDCD5, RAD50, and TM9SF2 were identified as possible candidates, among others. We failed to find many laboratory experiments contrasting the effects of Indian and Chinese orthologs at these sites on SIVmac pathogenesis, but future comparative studies might hold fertile ground for research into the biological mechanisms underlying innate resistance to SIVmac in Chinese rhesus macaques. PMID:25884674

  18. Draft Genome Sequence of Lactobacillus delbrueckii subsp. bulgaricus CFL1, a Lactic Acid Bacterium Isolated from French Handcrafted Fermented Milk.

    PubMed

    Meneghel, Julie; Dugat-Bony, Eric; Irlinger, Françoise; Loux, Valentin; Vidal, Marie; Passot, Stéphanie; Béal, Catherine; Layec, Séverine; Fonseca, Fernanda

    2016-01-01

    Lactobacillus delbrueckii subsp. bulgaricus (L. bulgaricus) is a lactic acid bacterium widely used for the production of yogurt and cheeses. Here, we report the genome sequence of L. bulgaricus CFL1 to improve our knowledge on its stress-induced damages following production and end-use processes. PMID:26941141

  19. Draft Genome Sequence of Cutaneotrichosporon curvatus DSM 101032 (Formerly Cryptococcus curvatus), an Oleaginous Yeast Producing Polyunsaturated Fatty Acids.

    PubMed

    Hofmeyer, Thomas; Hackenschmidt, Silke; Nadler, Florian; Thürmer, Andrea; Daniel, Rolf; Kabisch, Johannes

    2016-01-01

    Cutaneotrichosporon curvatus DSM 101032 is an oleaginous yeast that can be isolated from various habitats and is capable of producing substantial amounts of polyunsaturated fatty acids. Here, we present the first draft genome sequence of any C. curvatus species. PMID:27174275

  20. Complete genome sequence of Lactobacillus plantarum ZS2058, a probiotic strain with high conjugated linoleic acid production ability.

    PubMed

    Yang, Bo; Chen, Haiqin; Tian, Fengwei; Zhao, Jianxin; Gu, Zhennan; Zhang, Hao; Chen, Yong Q; Chen, Wei

    2015-11-20

    Lactobacillus plantarum ZS2058 was isolated from sauerkraut and identified to synthesize the beneficial metabolite conjugated linoleic acid. The genome contains a 319,7363-bp chromosome and three plasmids. The sequence will facilitate identification and characterization of the genetic determinants for its putative biological benefits. PMID:26439428

  1. Draft Genome Sequence of Burkholderia stabilis LA20W, a Trehalose Producer That Uses Levulinic Acid as a Substrate

    PubMed Central

    Sato, Yuya; Koike, Hideaki; Kondo, Susumu; Hori, Tomoyuki; Kanno, Manabu; Kimura, Nobutada; Morita, Tomotake; Kirimura, Kohtaro

    2016-01-01

    Burkholderia stabilis LA20W produces trehalose using levulinic acid (LA) as a substrate. Here, we report the 7.97-Mb draft genome sequence of B. stabilis LA20W, which will be useful in investigations of the enzymes involved in LA metabolism and the mechanism of LA-induced trehalose production. PMID:27491978

  2. Draft Genome Sequence of Acetobacter tropicalis Type Strain NBRC16470, a Producer of Optically Pure d-Glyceric Acid.

    PubMed

    Koike, Hideaki; Sato, Shun; Morita, Tomotake; Fukuoka, Tokuma; Habe, Hiroshi

    2014-01-01

    Here we report the 3.7-Mb draft genome sequence of Acetobacter tropicalis NBRC16470(T), which can produce optically pure d-glyceric acid (d-GA; 99% enantiomeric excess) from raw glycerol feedstock derived from biodiesel fuel production processes. PMID:25523780

  3. Genome Sequence of a Candidate World Health Organization Reference Strain of Zika Virus for Nucleic Acid Testing.

    PubMed

    Trösemeier, Jan-Hendrik; Musso, Didier; Blümel, Johannes; Thézé, Julien; Pybus, Oliver G; Baylis, Sally A

    2016-01-01

    We report here the sequence of a candidate reference strain of Zika virus (ZIKV) developed on behalf of the World Health Organization (WHO). The ZIKV reference strain is intended for use in nucleic acid amplification (NAT)-based assays for the detection and quantification of ZIKV RNA. PMID:27587826

  4. Draft Genome Sequence of Burkholderia stabilis LA20W, a Trehalose Producer That Uses Levulinic Acid as a Substrate.

    PubMed

    Sato, Yuya; Koike, Hideaki; Kondo, Susumu; Hori, Tomoyuki; Kanno, Manabu; Kimura, Nobutada; Morita, Tomotake; Kirimura, Kohtaro; Habe, Hiroshi

    2016-01-01

    Burkholderia stabilis LA20W produces trehalose using levulinic acid (LA) as a substrate. Here, we report the 7.97-Mb draft genome sequence of B. stabilis LA20W, which will be useful in investigations of the enzymes involved in LA metabolism and the mechanism of LA-induced trehalose production. PMID:27491978

  5. Draft Genome Sequence of Cutaneotrichosporon curvatus DSM 101032 (Formerly Cryptococcus curvatus), an Oleaginous Yeast Producing Polyunsaturated Fatty Acids

    PubMed Central

    Hofmeyer, Thomas; Hackenschmidt, Silke; Nadler, Florian; Thürmer, Andrea; Daniel, Rolf

    2016-01-01

    Cutaneotrichosporon curvatus DSM 101032 is an oleaginous yeast that can be isolated from various habitats and is capable of producing substantial amounts of polyunsaturated fatty acids. Here, we present the first draft genome sequence of any C. curvatus species. PMID:27174275

  6. Ultra high-throughput nucleic acid sequencing as a tool for virus discovery in the turkey gut.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recently, the use of the next generation of nucleic acid sequencing technology (i.e., 454 pyrosequencing, as developed by Roche/454 Life Sciences) has allowed an in-depth look at the uncultivated microorganisms present in complex environmental samples, including samples with agricultural importance....

  7. Draft Genome Sequence of Lactobacillus delbrueckii subsp. bulgaricus CFL1, a Lactic Acid Bacterium Isolated from French Handcrafted Fermented Milk

    PubMed Central

    Meneghel, Julie; Irlinger, Françoise; Loux, Valentin; Vidal, Marie; Passot, Stéphanie; Béal, Catherine; Layec, Séverine

    2016-01-01

    Lactobacillus delbrueckii subsp. bulgaricus (L. bulgaricus) is a lactic acid bacterium widely used for the production of yogurt and cheeses. Here, we report the genome sequence of L. bulgaricus CFL1 to improve our knowledge on its stress-induced damages following production and end-use processes. PMID:26941141

  8. Identification of G and P genotype-specific motifs in the predicted VP7 and VP4 amino acid sequences.

    PubMed

    Ma, Yongping

    2015-12-01

    Equine rotavirus (ERV) strain L338 (G13P[18]) has a unique G and P genotype. However, the evolutionary relationship of L338 with other ERVs is still unknown. Here whole genome analysis of the L338 ERV strain was independently performed. Its genotype constellations were determined as G13-P[18]-I6-R9-C9-M6-A6-N9-T12-E14-H11, confirming previous genotype assignments. The L338 strain only shared the P[18] and I6 genotypes with other ERVs. The nucleotide sequences of the other 9 RNA segments were different from those of cogent genes of all other group A rotavirus (RVA) strains including ERVs and formed unique phylogenetic lineages. The L338 evolutionary footprints were tentatively identified in both VP7 and VP4 amino acid sequences: two regions were found in VP7 and twelve in VP4. The conserved regions shared between L338 and other group A rotavirus strains (RVAs) indicated that L338 was more closely related genomically to animal and human RVAs other than ERVs, suggesting that L338 may not be an endogenous equine RV but have emerged as an interspecies reassortant with other RVA strains. Furthermore, genotype-specific motifs of all 27 G and 37 P types were identified in regions 7-1a (aa 91-100) of VP7 and regions 8-1 (aa146-151) and 8-3 (aa113-118 and 125-135) of VP4 (VP8*). PMID:26321159

  9. Sequence-Specific Recognition of MicroRNAs and Other Short Nucleic Acids with Solid-State Nanopores.

    PubMed

    Zahid, Osama K; Wang, Fanny; Ruzicka, Jan A; Taylor, Ethan W; Hall, Adam R

    2016-03-01

    The detection and quantification of short nucleic acid sequences has many potential applications in studying biological processes, monitoring disease initiation and progression, and evaluating environmental systems, but is challenging by nature. We present here an assay based on the solid-state nanopore platform for the identification of specific sequences in solution. We demonstrate that hybridization of a target nucleic acid with a synthetic probe molecule enables discrimination between duplex and single-stranded molecules with high efficacy. Our approach requires limited preparation of samples and yields an unambiguous translocation event rate enhancement that can be used to determine the presence and abundance of a single sequence within a background of nontarget oligonucleotides. PMID:26824296

  10. Isolation, characterization, and amino acid sequences of auracyanins, blue copper proteins from the green photosynthetic bacterium Chloroflexus aurantiacus

    NASA Technical Reports Server (NTRS)

    McManus, J. D.; Brune, D. C.; Han, J.; Sanders-Loehr, J.; Meyer, T. E.; Cusanovich, M. A.; Tollin, G.; Blankenship, R. E.

    1992-01-01

    Three small blue copper proteins designated auracyanin A, auracyanin B-1, and auracyanin B-2 have been isolated from the thermophilic green gliding photosynthetic bacterium Chloroflexus aurantiacus. All three auracyanins are peripheral membrane proteins. Auracyanin A was described previously (Trost, J. T., McManus, J. D., Freeman, J. C., Ramakrishna, B. L., and Blankenship, R. E. (1988) Biochemistry 27, 7858-7863) and is not glycosylated. The two B forms are glycoproteins and have almost identical properties to each other, but are distinct from the A form. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis apparent monomer molecular masses are 14 (A), 18 (B-2), and 22 (B-1) kDa. The amino acid sequences of the B forms are presented. All three proteins have similar absorbance, circular dichroism, and resonance Raman spectra, but the electron spin resonance signals are quite different. Laser flash photolysis kinetic analysis of the reactions of the three forms of auracyanin with lumiflavin and flavin mononucleotide semiquinones indicates that the site of electron transfer is negatively charged and has an accessibility similar to that found in other blue copper proteins. Copper analysis indicates that all three proteins contain 1 mol of copper per mol of protein. All three auracyanins exhibit a midpoint redox potential of +240 mV. Light-induced absorbance changes and electron spin resonance signals suggest that auracyanin A may play a role in photosynthetic electron transfer. Kinetic data indicate that all three proteins can donate electrons to cytochrome c-554, the electron donor to the photosynthetic reaction center.

  11. Human parainfluenza type 3 virus hemagglutinin-neuraminidase glycoprotein: nucleotide sequence of mRNA and limited amino acid sequence of the purified protein.

    PubMed Central

    Elango, N; Coligan, J E; Jambou, R C; Venkatesan, S

    1986-01-01

    The nucleotide sequence of mRNA for the hemagglutinin-neuraminidase (HN) protein of human parainfluenza type 3 virus obtained from the corresponding cDNA clone had a single long open reading frame encoding a putative protein of 64,254 daltons consisting of 572 amino acids. The deduced protein sequence was confirmed by limited N-terminal amino acid microsequencing of CNBr cleavage fragments of native HN that was purified by immunoprecipitation. The HN protein is moderately hydrophobic and has four potential sites (Asn-X-Ser/Thr) of N-glycosylation in the C-terminal half of the molecule. It is devoid of both the N-terminal signal sequence and the C-terminal membrane anchorage domain characteristic of the hemagglutinin of influenza virus and the fusion (F0) protein of the paramyxoviruses. Instead, it has a single prominent hydrophobic region capable of membrane insertion beginning at 32 residues from the N terminus. This N-terminal membrane insertion is similar to that of influenza virus neuraminidase and the recently reported structures of HN proteins of Sendai virus and simian virus 5. Images PMID:3003381

  12. Sequence of cDNA for rat cystathionine gamma-lyase and comparison of deduced amino acid sequence with related Escherichia coli enzymes.

    PubMed Central

    Erickson, P F; Maxwell, I H; Su, L J; Baumann, M; Glode, L M

    1990-01-01

    A cDNA clone for cystathionine gamma-lyase was isolated from a rat cDNA library in lambda gt11 by screening with a monospecific antiserum. The identity of this clone, containing 600 bp proximal to the 3'-end of the gene, was confirmed by positive hybridization selection. Northern-blot hybridization showed the expected higher abundance of the corresponding mRNA in liver than in brain. Two further cDNA clones from a plasmid pcD library were isolated by colony hybridization with the first clone and were found to contain inserts of 1600 and 1850 bp. One of these was confirmed as encoding cystathionine gamma-lyase by hybridization with two independent pools of oligodeoxynucleotides corresponding to partial amino acid sequence information for cystathionine gamma-lyase. The other clone (estimated to represent all but 8% of the 5'-end of the mRNA) was sequenced and its deduced amino acid sequence showed similarity to those of the Escherichia coli enzymes cystathionine beta-lyase and cystathionine gamma-synthase throughout its length, especially to that of the latter. Images Fig. 1. Fig. 2. Fig. 3. Fig. 5. PMID:2201285

  13. Sequence dependent N-terminal rearrangement and degradation of peptide nucleic acid (PNA) in aqueous solution

    NASA Technical Reports Server (NTRS)

    Eriksson, M.; Christensen, L.; Schmidt, J.; Haaima, G.; Orgel, L.; Nielsen, P. E.

    1998-01-01

    The stability of the PNA (peptide nucleic acid) thymine monomer inverted question markN-[2-(thymin-1-ylacetyl)]-N-(2-aminoaminoethyl)glycine inverted question mark and those of various PNA oligomers (5-8-mers) have been measured at room temperature (20 degrees C) as a function of pH. The thymine monomer undergoes N-acyl transfer rearrangement with a half-life of 34 days at pH 11 as analyzed by 1H NMR; and two reactions, the N-acyl transfer and a sequential degradation, are found by HPLC analysis to occur at measurable rates for the oligomers at pH 9 or above. Dependent on the amino-terminal sequence, half-lives of 350 h to 163 days were found at pH 9. At pH 12 the half-lives ranged from 1.5 h to 21 days. The results are discussed in terms of PNA as a gene therapeutic drug as well as a possible prebiotic genetic material.

  14. Snake venoms. The amino acid sequences of two proteinase inhibitor homologues from Dendroaspis angusticeps venom.

    PubMed

    Joubert, F J; Taljaard, N

    1980-05-01

    Toxins C13S1C3 and C13S2C3 from D. angusticeps venom were purified by gel filtration and ion exchange chromatography. Whereas C13S1C3 contains 57 amino acids, C13S2C3 contains 59 but each include six half-cystine residues. The complete primary structure of the low toxicity proteins have been elucidated. The sequences and the invariant residues of toxins C13S1C3 and C13S2C3 from D. angusticeps venom resemble, respectively, those of the proteinase inhibitor homologues K and I from D. polylepis polylepis venom and they are also homologous to the active proteinase inhibitors from various sources. In C13S1C3 and K the active site lysyl residue of active bovine pancreatic proteinase inhibitor is conserved but the site residue alanine, is replaced by lysine. In C13S2C3 and I the active site residue is replaced by tyrosine. PMID:7429422

  15. The amino acid sequences of two alpha chains of hemoglobins from Komodo dragon Varanus komodoensis and phylogenetic relationships of amniotes.

    PubMed

    Fushitani, K; Higashiyama, K; Moriyama, E N; Imai, K; Hosokawa, K

    1996-09-01

    To elucidate phylogenetic relationships among amniotes and the evolution of alpha globins, hemoglobins were analyzed from the Komodo dragon (Komodo monitor lizard) Varanus komodoensis, the world's largest extant lizard, inhabiting Komodo Islands, Indonesia. Four unique globin chains (alpha A, alpha D, beta B, and beta C) were isolated in an equal molar ratio by high performance liquid chromatography from the hemolysate. The amino acid sequences of two alpha chains were determined. The alpha D chain has a glutamine at E7 as does an alpha chain of a snake, Liophis miliaris, but the alpha A chain has a histidine at E7 like the majority of hemoglobins. Phylogenetic analyses of 19 globins including two alpha chains of Komodo dragon and ones from representative amniotes showed the following results: (1) The a chains of squamates (snakes and lizards), which have a glutamine at E7, are clustered with the embryonic alpha globin family, which typically includes the alpha D chain from birds; (2) birds form a sister group with other reptiles but not with mammals; (3) the genes for embryonic and adult types of alpha globins were possibly produced by duplication of the ancestral alpha gene before ancestral amniotes diverged, indicating that each of the present amniotes might carry descendants of the two types of alpha globin genes; (4) squamates first split off from the ancestor of other reptiles and birds. PMID:8752011

  16. Nucleotide and predicted amino acid sequence of a cDNA clone encoding part of human transketolase.

    PubMed

    Abedinia, M; Layfield, R; Jones, S M; Nixon, P F; Mattick, J S

    1992-03-31

    Transketolase is a key enzyme in the pentose-phosphate pathway which has been implicated in the latent human genetic disease, Wernicke-Korsakoff syndrome. Here we report the cloning and partial characterisation of the coding sequences encoding human transketolase from a human brain cDNA library. The library was screened with oligonucleotide probes based on the amino acid sequence of proteolytic fragments of the purified protein. Northern blots showed that the transketolase mRNA is approximately 2.2 kb, close to the minimum expected, of which approximately 60% was represented in the largest cDNA clone. Sequence analysis of the transketolase coding sequences reveals a number of homologies with related enzymes from other species. PMID:1567394

  17. 5S ribosomal ribonucleic acid sequences in Bacteroides and Fusobacterium: evolutionary relationships within these genera and among eubacteria in general

    NASA Technical Reports Server (NTRS)

    Van den Eynde, H.; De Baere, R.; Shah, H. N.; Gharbia, S. E.; Fox, G. E.; Michalik, J.; Van de Peer, Y.; De Wachter, R.

    1989-01-01

    The 5S ribosomal ribonucleic acid (rRNA) sequences were determined for Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides capillosus, Bacteroides veroralis, Porphyromonas gingivalis, Anaerorhabdus furcosus, Fusobacterium nucleatum, Fusobacterium mortiferum, and Fusobacterium varium. A dendrogram constructed by a clustering algorithm from these sequences, which were aligned with all other hitherto known eubacterial 5S rRNA sequences, showed differences as well as similarities with respect to results derived from 16S rRNA analyses. In the 5S rRNA dendrogram, Bacteroides clustered together with Cytophaga and Fusobacterium, as in 16S rRNA analyses. Intraphylum relationships deduced from 5S rRNAs suggested that Bacteroides is specifically related to Cytophaga rather than to Fusobacterium, as was suggested by 16S rRNA analyses. Previous taxonomic considerations concerning the genus Bacteroides, based on biochemical and physiological data, were confirmed by the 5S rRNA sequence analysis.

  18. Urinary t,t-muconic acid as an indicator of exposure to benzene.

    PubMed Central

    Inoue, O; Seiji, K; Nakatsuka, H; Watanabe, T; Yin, S N; Li, G L; Cai, S X; Jin, C; Ikeda, M

    1989-01-01

    A method for rapidly determining t,t-muconic acid (MA) by high performance liquid chromatography was developed and successfully applied to urine samples from 152 workers exposed to benzene (64 men, 88 women) and 213 non-exposed controls (113 men, 100 women). The MA concentrations in urine correlated linearly with time weighted average benzene concentrations in the breath zone air of workers. A cross sectional balance study showed that about 2% of benzene inhaled is excreted into the urine as MA. The MA concentrations in the urine of the non-exposed was below the detection limit (less than 0.1 mg/l) in most cases, and the 95% lower confidence limit of MA for those exposed to benzene at 5 ppm (5.0 mg/l as a non-corrected value) was higher than the 97.5%-tile values for the non-exposed (1.4 mg/l). In practice, it was possible to separate those exposed to 6-7 ppm benzene from the non-exposed by means of urine analysis for MA. The urinary MA concentration was suppressed by coexposure to toluene. PMID:2923822

  19. Sample Prep, Workflow Automation and Nucleic Acid Fractionation for Next Generation Sequencing

    SciTech Connect

    Roskey, Mark

    2010-06-03

    Mark Roskey of Caliper LifeSciences discusses how the company's technologies fit into the next generation sequencing workflow on June 3, 2010 at the "Sequencing, Finishing, Analysis in the Future" meeting in Santa Fe, NM

  20. Evolution of vertebrate IgM: complete amino acid sequence of the constant region of Ambystoma mexicanum mu chain deduced from cDNA sequence.

    PubMed

    Fellah, J S; Wiles, M V; Charlemagne, J; Schwager, J

    1992-10-01

    cDNA clones coding for the constant region of the Mexican axolotl (Ambystoma mexicanum) mu heavy immunoglobulin chain were selected from total spleen RNA, using a cDNA polymerase chain reaction technique. The specific 5'-end primer was an oligonucleotide homologous to the JH segment of Xenopus laevis mu chain. One of the clones, JHA/3, corresponded to the complete constant region of the axolotl mu chain, consisting of a 1362-nucleotide sequence coding for a polypeptide of 454 amino acids followed in 3' direction by a 179-nucleotide untranslated region and a polyA+ tail. The axolotl C mu is divided into four typical domains (C mu 1-C mu 4) and can be aligned with the Xenopus C mu with an overall identity of 56% at the nucleotide level. Percent identities were particularly high between C mu 1 (59%) and C mu 4 (71%). The C-terminal 20-amino acid segment which constitutes the secretory part of the mu chain is strongly homologous to the equivalent sequences of chondrichthyans and of other tetrapods, including a conserved N-linked oligosaccharide, the penultimate cysteine and the C-terminal lysine. The four C mu domains of 13 vertebrate species ranging from chondrichthyans to mammals were aligned and compared at the amino acid level. The significant number of mu-specific residues which are conserved into each of the four C mu domains argues for a continuous line of evolution of the vertebrate mu chain. This notion was confirmed by the ability to reconstitute a consistent vertebrate evolution tree based on the phylogenic parsimony analysis of the C mu 4 sequences. PMID:1382992

  1. Neurobehavioral effects during exposures to propionic acid--an indicator of chemosensory distraction?

    PubMed

    Hey, Kathrin; Juran, Stephanie; Schäper, Michael; Kleinbeck, Stefan; Kiesswetter, Ernst; Blaszkewicz, Meinolf; Golka, Klaus; Brüning, Thomas; van Thriel, Christoph

    2009-11-01

    The chemosensory effects of propionic acid (PA) in humans have not been conclusively studied and there is no established occupational exposure limit (OEL) in Germany. In addition to sensory irritation caused by PA, it was hypothesized that the annoying odor of PA might interfere with the performance results. There were 23 consenting healthy participants (12 female, 11 male) in the group studied. They were exposed for 4 h to PA in concentrations of 0.3, 5 and 10 ppm in a cross-over design. During these exposures, performance was recorded with four cognitive tests measuring response-inhibition, working memory, set-shifting, and divided attention. Odor annoyance, other chemosensory sensations, and acute symptoms were assessed before, during, and after exposure with standardized rating scales. Moderate odor annoyance and weak sensory irritation were reported during 5 and 10 ppm exposure conditions. The different levels of exposure to PA had no impact upon the results of three out of the four behavioral tests. The difficulties of the task were reflected in the results. However in the fourth, which was the response-inhibition task, there was significant increase in the error rates which corresponded to the exposure levels. Results from previous experiments suggested high odor annoyance at the investigated concentrations. Our findings showed that odor annoyance and reported sensory irritations were low. In conclusion, the hypothesis of a distractive effect due to the malodor of PA could not be confirmed. Only in concentrations as high as 10 ppm acute PA exposure affected the response accuracy of one of the four neurobehavioral task. For other more annoying substances, a neurobehavioral effect influenced by an indirect mechanism of resources competition is still conceivable. PMID:19733590

  2. Low levels of haptoglobin and putative amino acid sequence in Taiwanese Lanyu miniature pigs.

    PubMed

    Yueh, Sunny C H; Wang, Yao Horng; Lin, Kuan Yu; Tseng, Chi Feng; Chu, Hsien Pin; Chen, Kuen Jaw; Wang, Shih Sheng; Lai, I Hsiang; Mao, Simon J T

    2008-04-01

    Porcine haptoglobin (Hp) is an acute phase protein. Its plasma level increases significantly during inflammation and infection. One of the main functions of Hp is to bind free hemoglobin (Hb) and inhibit its oxidative activity. In the present report, we studied the Hp phenotype of Taiwanese Lanyu miniature pigs (TLY minipigs; n=43) and found their Hp structure to be a homodimer (beta-alpha-alpha-beta) similar to human Hp 1-1. Interestingly, Western blot and high performance liquid chromatographic (HPLC) analysis showed that 25% of the TLY minipigs possessed low or no plasma Hp level (<0.05 mg/ml). The Hp cDNA of these TLY minipigs was then cloned, and the translated amino acid sequence was analyzed. No sequences were found to be deficient; they showed a 99.7% identity with domestic pigs (NP_999165). The mean overall Hp level of the TLY minipigs (0.21 +/- 0.25 mg/ml; n=43) determined by enzyme-linked immunosorbent assay (ELISA) was markedly lower than that of domestic pigs (0.78 +/- 0.45 mg/ml; p<0.001), while 25% of the TLY minipigs had an Hp level that was extremely low (<0.05 mg/ml). In addition, the initial recovery rate (first 40 min) in the circulation of infused fluorescein isothiocyanate (FITC)-Hb was significantly higher in the TLY minipigs with extremely low Hp levels than those with high levels. This data suggests that the low concentration of Hp-Hb complex is responsible for the higher recovery rate of Hb in the circulation. TLY minipigs have been used as an experimental model for cardiovascular diseases; whether they can be used as a model for inflammatory diseases, with Hp as a marker, remains a topic of interest. However, since the Hp level varies significantly among individual TLY minipigs, it is necessary to prescreen the Hp levels of the animals to minimize variation in the experimental baseline. The present study may provide a reference value for future use of the TLY minipig as an animal model for inflammation-associated diseases. PMID:18460833

  3. The Genome Sequence of the Highly Acetic Acid-Tolerant Zygosaccharomyces bailii-Derived Interspecies Hybrid Strain ISA1307, Isolated From a Sparkling Wine Plant

    PubMed Central

    Mira, Nuno P.; Münsterkötter, Martin; Dias-Valada, Filipa; Santos, Júlia; Palma, Margarida; Roque, Filipa C.; Guerreiro, Joana F.; Rodrigues, Fernando; Sousa, Maria João; Leão, Cecília; Güldener, Ulrich; Sá-Correia, Isabel

    2014-01-01

    In this work, it is described the sequencing and annotation of the genome of the yeast strain ISA1307, isolated from a sparkling wine continuous production plant. This strain, formerly considered of the Zygosaccharomyces bailii species, has been used to study Z. bailii physiology, in particular, its extreme tolerance to acetic acid stress at low pH. The analysis of the genome sequence described in this work indicates that strain ISA1307 is an interspecies hybrid between Z. bailii and a closely related species. The genome sequence of ISA1307 is distributed through 154 scaffolds and has a size of around 21.2 Mb, corresponding to 96% of the genome size estimated by flow cytometry. Annotation of ISA1307 genome includes 4385 duplicated genes (∼90% of the total number of predicted genes) and 1155 predicted single-copy genes. The functional categories including a higher number of genes are ‘Metabolism and generation of energy’, ‘Protein folding, modification and targeting’ and ‘Biogenesis of cellular components’. The knowledge of the genome sequence of the ISA1307 strain is expected to contribute to accelerate systems-level understanding of stress resistance mechanisms in Z. bailii and to inspire and guide novel biotechnological applications of this yeast species/strain in fermentation processes, given its high resilience to acidic stress. The availability of the ISA1307 genome sequence also paves the way to a better understanding of the genetic mechanisms underlying the generation and selection of more robust hybrid yeast strains in the stressful environment of wine fermentations. PMID:24453040

  4. Nucleotide sequences 1986/1987

    SciTech Connect

    Not Available

    1987-01-01

    These eight volumes are the third annual published compendium of nucleic acid sequences included in the European Molecular Biology Laboratory Nucleotide Sequence Data Library and the GenBank Genetic Sequences Data Bank. Each volume surveys one or more subdivisions of the database. The volume subtitles are: Primates; Rodents; Other Vertebrates and Invertebrates, Plants and Organelles, Bacteria and Bacteriophage, Viruses, Structural RNA, Synthetic and Unannotated Sequences, and Database Directory and Master Indices.

  5. Amino acid sequence of mouse nidogen, a multidomain basement membrane protein with binding activity for laminin, collagen IV and cells.

    PubMed Central

    Mann, K; Deutzmann, R; Aumailley, M; Timpl, R; Raimondi, L; Yamada, Y; Pan, T C; Conway, D; Chu, M L

    1989-01-01

    The whole amino acid sequence of nidogen was deduced from cDNA clones isolated from expression libraries and confirmed to approximately 50% by Edman degradation of peptides. The protein consists of some 1217 amino acid residues and a 28-residue signal peptide. The data support a previously proposed dumb-bell model of nidogen by demonstrating a large N-terminal globular domain (641 residues), five EGF-like repeats constituting the rod-like domain (248 residues) and a smaller C-terminal globule (328 residues). Two more EGF-like repeats interrupt the N-terminal and terminate the C-terminal sequences. Weak sequence homologies (25%) were detected between some regions of nidogen, the LDL receptor, thyroglobulin and the EGF precursor. Nidogen contains two consensus sequences for tyrosine sulfation and for asparagine beta-hydroxylation, two N-linked carbohydrate acceptor sites and, within one of the EGF-like repeats an Arg-Gly-Asp sequence. The latter was shown to be functional in cell attachment to nidogen. Binding sites for laminin and collagen IV are present on the C-terminal globule but not yet precisely localized. Images PMID:2496973

  6. Jack bean α-mannosidase: amino acid sequencing and N-glycosylation analysis of a valuable glycomics tool.

    PubMed

    Gnanesh Kumar, B S; Pohlentz, Gottfried; Schulte, Mona; Mormann, Michael; Siva Kumar, Nadimpalli

    2014-03-01

    Jack bean (Canavalia ensiformis) seeds contain several biologically important proteins among which α-mannosidase (EC 3.2.1.24) has been purified, its biochemical properties studied and widely used in glycan analysis. In the present study, we have used the purified enzyme and derived its amino acid sequence covering both the known subunits (molecular mass of ∼66,000 and ∼44,000 Da) hitherto not known in its entirety. Peptide de novo sequencing and structural elucidation of N-glycopeptides obtained either directly from proteolytic digestion or after zwitterionic hydrophilic interaction liquid chromatography solid phase extraction-based separation were performed by use of nanoelectrospray ionization quadrupole time-of-flight mass spectrometry and low-energy collision-induced dissociation experiments. De novo sequencing provided new insights into the disulfide linkage organization, intersection of subunits and complete N-glycan structures along with site specificities. The primary sequence suggests that the enzyme belongs to glycosyl hydrolase family 38 and the N-glycan sequence analysis revealed high-mannose oligosaccharides, which were found to be heterogeneous with varying number of hexoses viz, Man8-9GlcNAc2 and Glc1Man9GlcNAc2 in an evolutionarily conserved N-glycosylation site. This site with two proximal cysteines is present in all the acidic α-mannosidases reported so far in eukaryotes. Further, a truncated paucimannose type was identified to be lacking terminal two mannose, Man1(Xyl)GlcNAc2 (Fuc). PMID:24295789

  7. Complete Genome Sequence of Enterococcus mundtii QU 25, an Efficient l-(+)-Lactic Acid-Producing Bacterium

    PubMed Central

    Shiwa, Yuh; Yanase, Hiroaki; Hirose, Yuu; Satomi, Shohei; Araya-Kojima, Tomoko; Watanabe, Satoru; Zendo, Takeshi; Chibazakura, Taku; Shimizu-Kadota, Mariko; Yoshikawa, Hirofumi; Sonomoto, Kenji

    2014-01-01

    Enterococcus mundtii QU 25, a non-dairy bacterial strain of ovine faecal origin, can ferment both cellobiose and xylose to produce l-lactic acid. The use of this strain is highly desirable for economical l-lactate production from renewable biomass substrates. Genome sequence determination is necessary for the genetic improvement of this strain. We report the complete genome sequence of strain QU 25, primarily determined using Pacific Biosciences sequencing technology. The E. mundtii QU 25 genome comprises a 3 022 186-bp single circular chromosome (GC content, 38.6%) and five circular plasmids: pQY182, pQY082, pQY039, pQY024, and pQY003. In all, 2900 protein-coding sequences, 63 tRNA genes, and 6 rRNA operons were predicted in the QU 25 chromosome. Plasmid pQY024 harbours genes for mundticin production. We found that strain QU 25 produces a bacteriocin, suggesting that mundticin-encoded genes on plasmid pQY024 were functional. For lactic acid fermentation, two gene clusters were identified—one involved in the initial metabolism of xylose and uptake of pentose and the second containing genes for the pentose phosphate pathway and uptake of related sugars. This is the first complete genome sequence of an E. mundtii strain. The data provide insights into lactate production in this bacterium and its evolution among enterococci. PMID:24568933

  8. Gastropod arginine kinases from Cellana grata and Aplysia kurodai. Isolation and cDNA-derived amino acid sequences.

    PubMed

    Suzuki, T; Inoue, N; Higashi, T; Mizobuchi, R; Sugimura, N; Yokouchi, K; Furukohri, T

    2000-12-01

    Arginine kinase (AK) was isolated from the radular muscle of the gastropod molluscs Cellana grata (subclass Prosobranchia) and Aplysia kurodai (subclass Opisthobranchia), respectively, by ammonium sulfate fractionation, Sephadex G-75 gel filtration and DEAE-ion exchange chromatography. The denatured relative molecular mass values were estimated to be 40 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isolated enzyme from Aplysia gave a Km value of 0.6 mM for arginine and a Vmax value of 13 micromole Pi min(-1) mg protein(-1) for the forward reaction. These values are comparable to other molluscan AKs. The cDNAs encoding Cellana and Aplysia AKs were amplified by polymerase chain reaction, and the nucleotide sequences of 1,608 and 1,239 bp, respectively, were determined. The open reading frame for Cellana AK is 1044 nucleotides in length and encodes a protein with 347 amino acid residues, and that for A. kurodai is 1077 nucleotides and 354 residues. The cDNA-derived amino acid sequences were validated by chemical sequencing of internal lysyl endopeptidase peptides. The amino acid sequences of Cellana and Aplysia AKs showed the highest percent identity (66-73%) with those of the abalone Nordotis and turbanshell Battilus belonging to the same class Gastropoda. These AK sequences still have a strong homology (63-71%) with that of the chiton Liolophura (class Polyplacophora), which is believed to be one of the most primitive molluscs. On the other hand, these AK sequences are less homologous (55-57%) with that of the clam Pseudocardium (class Bivalvia), suggesting that the biological position of the class Polyplacophora should be reconsidered. PMID:11281267

  9. Studies on the high-sulphur proteins of reduced Merino wool. Amino acid sequence of protein SCMKB-IIIB4

    PubMed Central

    Swart, L. S.; Haylett, T.

    1971-01-01

    The complete amino acid sequence of protein SCMKB-IIIB4 is presented. It is closely related to the sequence of protein SCMKB-IIIB3 (Haylett, Swart & Parris, 1971) differing in only four positions. The peptic and thermolysin peptides of protein SCMKB-IIIB4 were analysed by the dansyl–Edman method (Gray, 1967) and by tritium-labelling of C-terminal residues (Matsuo, Fujimoto & Tatsuno, 1966). This protein is the third member of a group of high-sulphur wool proteins with molecular weight of about 11400. It consists of 98 residues and has acetylalanine and carboxymethylcysteine as N- and C-terminal residues respectively. PMID:4942536

  10. Complete Sequence of the mitochondrial genome of the tapeworm Hymenolepis diminuta: Gene arrangements indicate that platyhelminths are eutrochozoans

    SciTech Connect

    von Nickisch-Rosenegk, Markus; Brown, Wesley M.; Boore, Jeffrey L.

    2001-01-01

    Using ''long-PCR'' we have amplified in overlapping fragments the complete mitochondrial genome of the tapeworm Hymenolepis diminuta (Platyhelminthes: Cestoda) and determined its 13,900 nucleotide sequence. The gene content is the same as that typically found for animal mitochondrial DNA (mtDNA) except that atp8 appears to be lacking, a condition found previously for several other animals. Despite the small size of this mtDNA, there are two large non-coding regions, one of which contains 13 repeats of a 31 nucleotide sequence and a potential stem-loop structure of 25 base pairs with an 11-member loop. Large potential secondary structures are identified also for the non-coding regions of two other cestode mtDNAs. Comparison of the mitochondrial gene arrangement of H. diminuta with those previously published supports a phylogenetic position of flatworms as members of the Eutrochozoa, rather than being basal to either a clade of protostomes or a clade of coelomates.

  11. Exome Sequencing in an Admixed Isolated Population Indicates NFXL1 Variants Confer a Risk for Specific Language Impairment

    PubMed Central

    Villanueva, Pía; Nudel, Ron; Hoischen, Alexander; Fernández, María Angélica; Simpson, Nuala H.; Gilissen, Christian; Reader, Rose H.; Jara, Lillian; Echeverry, Maria Magdalena; Francks, Clyde; Baird, Gillian; Conti-Ramsden, Gina; O’Hare, Anne; Bolton, Patrick F.; Hennessy, Elizabeth R.; Palomino, Hernán; Carvajal-Carmona, Luis; Veltman, Joris A.; Cazier, Jean-Baptiste; De Barbieri, Zulema

    2015-01-01

    Children affected by Specific Language Impairment (SLI) fail to acquire age appropriate language skills despite adequate intelligence and opportunity. SLI is highly heritable, but the understanding of underlying genetic mechanisms has proved challenging. In this study, we use molecular genetic techniques to investigate an admixed isolated founder population from the Robinson Crusoe Island (Chile), who are affected by a high incidence of SLI, increasing the power to discover contributory genetic factors. We utilize exome sequencing in selected individuals from this population to identify eight coding variants that are of putative significance. We then apply association analyses across the wider population to highlight a single rare coding variant (rs144169475, Minor Allele Frequency of 4.1% in admixed South American populations) in the NFXL1 gene that confers a nonsynonymous change (N150K) and is significantly associated with language impairment in the Robinson Crusoe population (p = 2.04 × 10–4, 8 variants tested). Subsequent sequencing of NFXL1 in 117 UK SLI cases identified four individuals with heterozygous variants predicted to be of functional consequence. We conclude that coding variants within NFXL1 confer an increased risk of SLI within a complex genetic model. PMID:25781923

  12. Exome sequencing in an admixed isolated population indicates NFXL1 variants confer a risk for specific language impairment.

    PubMed

    Villanueva, Pía; Nudel, Ron; Hoischen, Alexander; Fernández, María Angélica; Simpson, Nuala H; Gilissen, Christian; Reader, Rose H; Jara, Lillian; Echeverry, María Magdalena; Echeverry, Maria Magdalena; Francks, Clyde; Baird, Gillian; Conti-Ramsden, Gina; O'Hare, Anne; Bolton, Patrick F; Hennessy, Elizabeth R; Palomino, Hernán; Carvajal-Carmona, Luis; Veltman, Joris A; Cazier, Jean-Baptiste; De Barbieri, Zulema; Fisher, Simon E; Newbury, Dianne F

    2015-03-01

    Children affected by Specific Language Impairment (SLI) fail to acquire age appropriate language skills despite adequate intelligence and opportunity. SLI is highly heritable, but the understanding of underlying genetic mechanisms has proved challenging. In this study, we use molecular genetic techniques to investigate an admixed isolated founder population from the Robinson Crusoe Island (Chile), who are affected by a high incidence of SLI, increasing the power to discover contributory genetic factors. We utilize exome sequencing in selected individuals from this population to identify eight coding variants that are of putative significance. We then apply association analyses across the wider population to highlight a single rare coding variant (rs144169475, Minor Allele Frequency of 4.1% in admixed South American populations) in the NFXL1 gene that confers a nonsynonymous change (N150K) and is significantly associated with language impairment in the Robinson Crusoe population (p = 2.04 × 10-4, 8 variants tested). Subsequent sequencing of NFXL1 in 117 UK SLI cases identified four individuals with heterozygous variants predicted to be of functional consequence. We conclude that coding variants within NFXL1 confer an increased risk of SLI within a complex genetic model. PMID:25781923

  13. A computer program for the estimation of protein and nucleic acid sequence diversity in random point mutagenesis libraries

    PubMed Central

    Volles, Michael J.; Lansbury, Peter T.

    2005-01-01

    A computer program for the generation and analysis of in silico random point mutagenesis libraries is described. The program operates by mutagenizing an input nucleic acid sequence according to mutation parameters specified by the user for each sequence position and type of point mutation. The program can mimic almost any type of random mutagenesis library, including those produced via error-prone PCR (ep-PCR), mutator Escherichia coli strains, chemical mutagenesis, and doped or random oligonucleotide synthesis. The program analyzes the generated nucleic acid sequences and/or the associated protein library to produce several estimates of library diversity (number of unique sequences, point mutations, and single point mutants) and the rate of saturation of these diversities during experimental screening or selection of clones. This information allows one to select the optimal screen size for a given mutagenesis library, necessary to efficiently obtain a certain coverage of the sequence-space. The program also reports the abundance of each specific protein mutation at each sequence position, which is useful as a measure of the level and type of mutation bias in the library. Alternatively, one can use the program to evaluate the relative merits of preexisting libraries, or to examine various hypothetical mutation schemes to determine the optimal method for creating a library that serves the screen/selection of interest. Simulated libraries of at least 109 sequences are accessible by the numerical algorithm with currently available personal computers; an analytical algorithm is also available which can rapidly calculate a subset of the numerical statistics in libraries of arbitrarily large size. A multi-type double-strand stochastic model of ep-PCR is developed in an appendix to demonstrate the applicability of the algorithm to amplifying mutagenesis procedures. Estimators of DNA polymerase mutation-type-specific error rates are derived using the model. Analyses of an

  14. DNA Sequence and Expression Variation of Hop (Humulus lupulus) Valerophenone Synthase (VPS), a Key Gene in Bitter Acid Biosynthesis

    PubMed Central

    Castro, Consuelo B.; Whittock, Lucy D.; Whittock, Simon P.; Leggett, Grey; Koutoulis, Anthony

    2008-01-01

    Background The hop plant (Humulus lupulus) is a source of many secondary metabolites, with bitter acids essential in the beer brewing industry and others having potential applications for human health. This study investigated variation in DNA sequence and gene expression of valerophenone synthase (VPS), a key gene in the bitter acid biosynthesis pathway of hop. Methods Sequence variation was studied in 12 varieties, and expression was analysed in four of the 12 varieties in a series across the development of the hop cone. Results Nine single nucleotide polymorphisms (SNPs) were detected in VPS, seven of which were synonymous. The two non-synonymous polymorphisms did not appear to be related to typical bitter acid profiles of the varieties studied. However, real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) analysis of VPS expression during hop cone development showed a clear link with the bitter acid content. The highest levels of VPS expression were observed in two triploid varieties, ‘Symphony’ and ‘Ember’, which typically have high bitter acid levels. Conclusions In all hop varieties studied, VPS expression was lowest in the leaves and an increase in expression was consistently observed during the early stages of cone development. PMID:18519445

  15. The amino acid sequence of protein SCMK-B2A from the high-sulphur fraction of wool keratin

    PubMed Central

    Elleman, T. C.

    1972-01-01

    1. The amino acid sequence of protein SCMK-B2A, a reduced and S-carboxymethylated protein from the high-sulphur fraction of wool, has been determined. 2. This protein of 171 amino acid residues displays both a high degree of internal homology and extensive external homology with other members of the SCMK-B2 group of proteins. 3. Evidence is presented which suggests that the SCMK-B2 group of proteins are produced by separate non-allelic genes. ImagesPLATE 1 PMID:4679226

  16. Transcriptome sequencing revealed the transcriptional organization at ribosome-mediated attenuation sites in Corynebacterium glutamicum and identified a novel attenuator involved in aromatic amino acid biosynthesis.

    PubMed

    Neshat, Armin; Mentz, Almut; Rückert, Christian; Kalinowski, Jörn

    2014-11-20

    The Gram-positive bacterium Corynebacterium glutamicum belongs to the order Corynebacteriales and is used as a producer of amino acids at industrial scales. Due to its economic importance, gene expression and particularly the regulation of amino acid biosynthesis has been investigated extensively. Applying the high-resolution technique of transcriptome sequencing (RNA-seq), recently a vast amount of data has been generated that was used to comprehensively analyze the C. glutamicum transcriptome. By analyzing RNA-seq data from a small RNA cDNA library of C. glutamicum, short transcripts in the known transcriptional attenuators sites of the trp operon, the ilvBNC operon and the leuA gene were verified. Furthermore, whole transcriptome RNA-seq data were used to elucidate the transcriptional organization of these three amino acid biosynthesis operons. In addition, we discovered and analyzed the novel attenuator aroR, located upstream of the aroF gene (cg1129). The DAHP synthase encoded by aroF catalyzes the first step in aromatic amino acid synthesis. The AroR leader peptide contains the amino acid sequence motif F-Y-F, indicating a regulatory effect by phenylalanine and tyrosine. Analysis by real-time RT-PCR suggests that the attenuator regulates the transcription of aroF in dependence of the cellular amount of tRNA loaded with phenylalanine when comparing a phenylalanine-auxotrophic C. glutamicum mutant fed with limiting and excess amounts of a phenylalanine-containing dipeptide. Additionally, the very interesting finding was made that all analyzed attenuators are leaderless transcripts. PMID:24910972

  17. Sterols and fatty acid biomarkers as indicators of changes in soil microbial communities in a uranium mine area.

    PubMed

    Guedes, Maria J; Pereira, Ruth; Duarte, Kátia; Rocha-Santos, Teresa A P; Antunes, Sara C; Gonçalves, Fernando; Duarte, Armando C; Freitas, Ana C

    2011-01-01

    Included in the 2nd tier of a site specific risk assessment that is being carried out in an abandoned uranium mine (Cunha Baixa uranium mine, Central Portugal), fatty acids biomarkers and sterols were analyzed to assess the impact of soil contamination with metals and radionuclides in the structure of the microbial community in seven sampling sites located at different distances from the mine. Surface soil samples were collected in those sampling sites in the four different seasons of the year. Principal component analysis (PCA) was performed on fatty acid biomarkers and sterols. Subsequently PCA scores obtained for both components were used to test the effect of sites and seasons, on soil samples collected in the Cunha Baixa uranium mine, through bi-factorial ANOVAs. Through PCA analysis, two distinct groups were set apart along the first two components. One group included sites at a great distance from the mine which were negatively correlated with higher contents of iC15:0 and iC17:0, both indicators of Gram-positive bacteria, as well as with ergosterol, cholestanol and cholesterol. The second group, in turn, was composed of the sampling sites most impacted by ore exploration, in situ leaching of poor ore, and spread of sludge from the effluent treatment pond. These sites were positively correlated with higher levels of iC16:0 (Gram-positive bacteria indicator), cyC17:0 (generally common in gram negative bacteria) and C18:0 and C17:0 biomarkers of non-specific bacteria. The profile of fatty acids obtained in the sampling sites revealed variable predominance of groups of bacteria which are a clear indication of differences in the soil microbial communities that are directly related to the environmental conditions prevailing in the uranium mine area. PMID:21547821

  18. DNA polymerase gene sequences indicate western and forest tent caterpillar viruses form a new taxonomic group within baculoviruses.

    PubMed

    Nielsen, Cydney B; Cooper, Dawn; Short, Steven M; Myers, Judith H; Suttle, Curtis A

    2002-11-01

    Baculoviruses infect larval lepidopterans, and thus have potential value as microbial controls of agricultural and forest pests. Understanding their genetic relatedness and host specificity is relevant to the risk assessment of viral insecticides if non-target impacts are to be avoided. DNA polymerase gene sequences have been demonstrated to be useful for inferring genetic relatedness among dsDNA viruses. We have adopted this approach to examine the relatedness among natural isolates of two uncharacterized caterpillar-infecting baculoviruses, Malacosoma californicum pluviale nucleopolyhedrovirus (McplMNPV) and Malacosoma disstria nucleopolyhedrovirus (MadiMNPV), which infect two closely related host species with little to no cross-infectivity. We designed two degenerate primers (BVP1 and BVP2) based on protein motifs conserved among baculoviruses. McplMNPV and MadiMNPV viral DNA was obtained from naturally infected caterpillars collected from geographically distinct sites in the Southern Gulf Islands and Prince George regions of British Columbia, Canada. Sequencing of 0.9 kb PCR amplicons from six McplMNPV and six MadiMNPV isolates obtained from a total of eight sites, revealed very low nucleotide variation among McplMNPV isolates (99.2-100% nucleotide identity) and among MadiMNPV isolates (98.9-100% nucleotide identity). Greater nucleotide variation was observed between viral isolates from the two different caterpillar species (only 84.7-86.1% nucleotide identity). Both maximum parsimony and maximum likelihood phylogenetic analyses support placement of McplMNPV and MadiMNPV in a clade that is distinct from other groups of baculoviruses. PMID:12507483

  19. A knowledge engineering approach to recognizing and extracting sequences of nucleic acids from scientific literature.

    PubMed

    García-Remesal, Miguel; Maojo, Victor; Crespo, José

    2010-01-01

    In this paper we present a knowledge engineering approach to automatically recognize and extract genetic sequences from scientific articles. To carry out this task, we use a preliminary recognizer based on a finite state machine to extract all candidate DNA/RNA sequences. The latter are then fed into a knowledge-based system that automatically discards false positives and refines noisy and incorrectly merged sequences. We created the knowledge base by manually analyzing different manuscripts containing genetic sequences. Our approach was evaluated using a test set of 211 full-text articles in PDF format containing 3134 genetic sequences. For such set, we achieved 87.76% precision and 97.70% recall respectively. This method can facilitate different research tasks. These include text mining, information extraction, and information retrieval research dealing with large collections of documents containing genetic sequences. PMID:21096556

  20. Ferredoxin:NADP oxidoreductase of Cyanophora paradoxa: purification, partial characterization, and N-terminal amino acid sequence.

    PubMed

    Gebhart, U B; Maier, T L; Stevanović, S; Bayer, M G; Schenk, H E

    1992-06-01

    The ferredoxin:NADP+ oxidoreductase of the protist Cyanophora paradoxa, as a descendant of a former symbiotic consortium, an important model organism in view of the Endosymbiosis Theory, is the first enzyme purified from a formerly original endocytobiont (cyanelle) that is found to be encoded in the nucleus of the host. This cyanoplast enzyme was isolated by FPLC (19% yield) and characterized with respect to the uv-vis spectrum, pH optimum (pH 9), molecular mass of 34 kDa, and an N-terminal amino acid sequence (24 residues). The enzyme shows, as known from other organisms, molecular heterogeneity. The N-terminus of a further ferredoxin:NADP+ oxidoreductase polypeptide represents a shorter sequence missing the first four amino acids of the mature enzyme. PMID:1392619

  1. Identification of novel rice low phytic acid mutations via TILLING by sequencing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Phytic acid (myo-inositol-1,2,3,4,5,6-hexakisphosphate or InsP6) accounts for 75-85% of the total phosphorus in seeds. Low phytic acid (lpa) mutants exhibit decreases in seed InsP6 with corresponding increases in inorganic P which, unlike phytic acid P, is readily utilized by humans and monogastric ...

  2. Snake venoms. The amino-acid sequence of trypsin inhibitor E of Dendroaspis polylepis polylepis (Black Mamba) venom.

    PubMed

    Joubert, F J; Strydom, D J

    1978-06-01

    Trypsin inhibitor E from black mamba venom comprises 59 amino acid residues in a single polypeptide chain, cross-linked by three intrachain disulphide bridges. The complete primary structure of inhibitor E was elucidated. The sequence is homologous with trypsin inhibitors from different sources. Unique among this homologous series of proteinase inhibitors, inhibitor E has an affinity for transition metal ions, exemplified here by Cu2 and Co2+. PMID:668688

  3. Draft Genome Sequence of Escherichia coli Strain VKPM B-10182, Producing the Enzyme for Synthesis of Cephalosporin Acids

    PubMed Central

    Mardanov, Andrey V.; Eldarov, Mikhail A.; Sklyarenko, Anna V.; Dumina, Maria V.; Beletsky, Alexey V.; Yarotsky, Sergey V.

    2014-01-01

    Escherichia coli strain VKPM B-10182, obtained by chemical mutagenesis from E. coli strain ATCC 9637, produces cephalosporin acid synthetase employed in the synthesis of β-lactam antibiotics, such as cefazolin. The draft genome sequence of strain VKPM B-10182 revealed 32 indels and 1,780 point mutations that might account for the improvement in antibiotic synthesis that we observed. PMID:25414512

  4. First draft genome sequencing of indole acetic acid producing and plant growth promoting fungus Preussia sp. BSL10.

    PubMed

    Khan, Abdul Latif; Asaf, Sajjad; Khan, Abdur Rahim; Al-Harrasi, Ahmed; Al-Rawahi, Ahmed; Lee, In-Jung

    2016-05-10

    Preussia sp. BSL10, family Sporormiaceae, was actively producing phytohormone (indole-3-acetic acid) and extra-cellular enzymes (phosphatases and glucosidases). The fungus was also promoting the growth of arid-land tree-Boswellia sacra. Looking at such prospects of this fungus, we sequenced its draft genome for the first time. The Illumina based sequence analysis reveals an approximate genome size of 31.4Mbp for Preussia sp. BSL10. Based on ab initio gene prediction, total 32,312 coding sequences were annotated consisting of 11,967 coding genes, pseudogenes, and 221 tRNA genes. Furthermore, 321 carbohydrate-active enzymes were predicted and classified into many functional families. PMID:26995610

  5. A simple ligation-based method to increase the information density in sequencing reactions used to deconvolute nucleic acid selections

    PubMed Central

    Childs-Disney, Jessica L.; Disney, Matthew D.

    2008-01-01

    Herein, a method is described to increase the information density of sequencing experiments used to deconvolute nucleic acid selections. The method is facile and should be applicable to any selection experiment. A critical feature of this method is the use of biotinylated primers to amplify and encode a BamHI restriction site on both ends of a PCR product. After amplification, the PCR reaction is captured onto streptavidin resin, washed, and digested directly on the resin. Resin-based digestion affords clean product that is devoid of partially digested products and unincorporated PCR primers. The product's complementary ends are annealed and ligated together with T4 DNA ligase. Analysis of ligation products shows formation of concatemers of different length and little detectable monomer. Sequencing results produced data that routinely contained three to four copies of the library. This method allows for more efficient formulation of structure-activity relationships since multiple active sequences are identified from a single clone. PMID:18065718

  6. A novel T-cell-defined HLA-DR polymorphism not predicted from the linear amino acid sequence.

    PubMed

    Termijtelen, A; van den Elsen, P; Koning, F; de Koster, S; Schroeijers, W; Vanderkerckhove, B

    1989-09-01

    Recent investigations have shown that alloreactive T cells are capable of responding to structures defined by specific linear amino acid sequences on class II molecules. In the present study we show that also a polymorphism can be recognized that is not defined by such linear amino acid sequences. Two human T-cell clones, sensitized to DRw13 haplotypes, are described. The description of clone c50 serves to exemplify the first model. This DRB1-specific clone responds to stimulator cells that carry DR molecules, different in their DRB1 first and second hypervariable regions (HV1 and HV2) but identical in their HV3 regions (i.e., DRw13,Dw18; DRw13,Dw19; DR4,Dw10; and DRw11,LDVII). The second clone, c1443, behaves nonconventionally. It responds to DRw13,Dw18; DRw13,Dw19; and DR4,Dw4 stimulator cells, although no specific amino acid sequence is shared between these specificities. The latter pattern of reactivity suggests the existence of a novel polymorphism recognized by alloreactive T cells. This particular polymorphism may also be biologically significant. PMID:2476425

  7. cDNA-derived amino-acid sequence of a land turtle (Geochelone carbonaria) beta-chain hemoglobin.

    PubMed

    Bordin, S; Meza, A N; Saad, S T; Ogo, S H; Costa, F F

    1997-06-01

    The cDNA sequence encoding the turtle Geochelone carbonaria beta-chain was determinated. The isolation of hemoglobin mRNA was based on degenerate primers' PCR in combination with 5'- and 3'-RACE protocol. The full length cDNA is 615 bp with the ATG start codon at position 53 and TGA stop codon at position 495; The AATAAA polyadenylation signal is found at position 599. The deduced polypeptyde contains 146 amino-acid residues. The predicted amino acid sequence shares 83% identity with the beta-globin of a related specie, the aquatic turtle C. p. belli. Otherwise, identity is higher when compared with chicken beta-Hb (80%) than with other reptilian orders (Squamata, 69%, and Crocodilia, 61%). Compared with human HbA, there is 67% identity, and at least three amino acid substitutions could be of some functional significance (Glu43 beta-->Ser, His116 beta-->Thr and His143 beta-->Leu). To our knowledge this represents the first cDNA sequence of a reptile globin gene described. PMID:9238523

  8. Amino acid sequence of the serine-repeat antigen (SERA) of Plasmodium falciparum determined from cloned cDNA.

    PubMed

    Bzik, D J; Li, W B; Horii, T; Inselburg, J

    1988-09-01

    We report the isolation of cDNA clones for a Plasmodium falciparum gene that encodes the complete amino acid sequence of a previously identified exported blood stage antigen. The Mr of this antigen protein had been determined by sodium dodecylsulphate-polyacrylamide gel electrophoresis analysis, by different workers, to be 113,000, 126,000, and 140,000. We show, by cDNA nucleotide sequence analysis, that this antigen gene encodes a 989 amino acid protein (111 kDa) that contains a potential signal peptide, but not a membrane anchor domain. In the FCR3 strain the serine content of the protein was 11%, of which 57% of the serine residues were localized within a 201 amino acid sequence that included 35 consecutive serine residues. The protein also contained three possible N-linked glycosylation sites and numerous possible O-linked glycosylation sites. The mRNA was abundant during late trophozoite-schizont parasite stages. We propose to identity this antigen, which had been called p126, by the acronym SERA, serine-repeat antigen, based on its complete structure. The usefulness of the cloned cDNA as a source of a possible malaria vaccine is considered in view of the previously demonstrated ability of the antigen to induce parasite-inhibitory antibodies and a protective immune response in Saimiri monkeys. PMID:2847041

  9. Amino acid sequences of lysozymes newly purified from invertebrates imply wide distribution of a novel class in the lysozyme family.

    PubMed

    Ito, Y; Yoshikawa, A; Hotani, T; Fukuda, S; Sugimura, K; Imoto, T

    1999-01-01

    Lysozymes were purified from three invertebrates: a marine bivalve, a marine conch, and an earthworm. The purified lysozymes all showed a similar molecular weight of 13 kDa on SDS/PAGE. Their N-terminal sequences up to the 33rd residue determined here were apparently homologous among them; in addition, they had a homology with a partial sequence of a starfish lysozyme which had been reported before. The complete sequence of the bivalve lysozyme was determined by peptide mapping and subsequent sequence analysis. This was composed of 123 amino acids including as many as 14 cysteine residues and did not show a clear homology with the known types of lysozymes. However, the homology search of this protein on the protein or nucleic acid database revealed two homologous proteins. One of them was a gene product, CELF22 A3.6 of C. elegans, which was a functionally unknown protein. The other was an isopeptidase of a medicinal leech, named destabilase. Thus, a new type of lysozyme found in at least four species across the three classes of the invertebrates demonstrates a novel class of protein/lysozyme family in invertebrates. The bivalve lysozyme, first characterized here, showed extremely high protein stability and hen lysozyme-like enzymatic features. PMID:9914527

  10. Complete Genome Sequences of Escherichia coli O157:H7 Strains SRCC 1675 and 28RC, Which Vary in Acid Resistance

    PubMed Central

    Baranzoni, Gian Marco; Reichenberger, Erin R.; Kim, Gwang-Hee; Breidt, Frederick; Kay, Kathryn; Oh, Deog-Hwan

    2016-01-01

    The level of acid resistance among Escherichia coli O157:H7 strains varies, and strains with higher resistance to acid may have a lower infectious dose. The complete genome sequences belonging to two strains of Escherichia coli O157:H7 with different levels of acid resistance are presented here. PMID:27469964

  11. Complete Genome Sequences of Escherichia coli O157:H7 Strains SRCC 1675 and 28RC, Which Vary in Acid Resistance.

    PubMed

    Baranzoni, Gian Marco; Fratamico, Pina M; Reichenberger, Erin R; Kim, Gwang-Hee; Breidt, Frederick; Kay, Kathryn; Oh, Deog-Hwan

    2016-01-01

    The level of acid resistance among Escherichia coli O157:H7 strains varies, and strains with higher resistance to acid may have a lower infectious dose. The complete genome sequences belonging to two strains of Escherichia coli O157:H7 with different levels of acid resistance are presented here. PMID:27469964

  12. Complete genome sequences of Escherichia coli O157:H7 strains SRCC 1675 and 28RC that vary in acid resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The level of acid resistance among Escherichia coli O157:H7 strains varies, and strains with higher resistance to acid may have a lower infectious dose. The complete genome sequences belonging to two strains of Escherichia coli O157:H7 with different levels of acid resistance are presented....

  13. Comparisons of Ribosomal Protein Gene Promoters Indicate Superiority of Heterologous Regulatory Sequences for Expressing Transgenes in Phytophthora infestans

    PubMed Central

    Khachatoorian, Careen; Judelson, Howard S.

    2015-01-01

    Molecular genetics approaches in Phytophthora research can be hampered by the limited number of known constitutive promoters for expressing transgenes and the instability of transgene activity. We have therefore characterized genes encoding the cytoplasmic ribosomal proteins of Phytophthora and studied their suitability for expressing transgenes in P. infestans. Phytophthora spp. encode a standard complement of 79 cytoplasmic ribosomal proteins. Several genes are duplicated, and two appear to be pseudogenes. Half of the genes are expressed at similar levels during all stages of asexual development, and we discovered that the majority share a novel promoter motif named the PhRiboBox. This sequence is enriched in genes associated with transcription, translation, and DNA replication, including tRNA and rRNA biogenesis. Promoters from the three P. infestans genes encoding ribosomal proteins S9, L10, and L23 and their orthologs from P. capsici were tested for their ability to drive transgenes in stable transformants of P. infestans. Five of the six promoters yielded strong expression of a GUS reporter, but the stability of expression was higher using the P. capsici promoters. With the RPS9 and RPL10 promoters of P. infestans, about half of transformants stopped making GUS over two years of culture, while their P. capsici orthologs conferred stable expression. Since cross-talk between native and transgene loci may trigger gene silencing, we encourage the use of heterologous promoters in transformation studies. PMID:26716454

  14. ERAP140/Nbla10993 is a novel favorable prognostic indicator for neuroblastoma induced in response to retinoic acid.

    PubMed

    Arai, Hiroshi; Ozaki, Toshinori; Niizuma, Hidetaka; Nakamura, Yohko; Ohira, Miki; Takano, Kunio; Matsumoto, Masahiko; Nakagawara, Akira

    2008-06-01

    In the present study, we identified a gene termed Nbla10993 whose expression levels are higher in favorable neuroblastomas versus unfavorable ones. Structural analysis showed that Nbla10993 is a novel splicing variant of the ER-associated protein of 140 kDa (ERAP140), which lacks the central acidic as well as the COOH-terminal Cys/His-rich domain. Similarly, ERAP140 was preferentially expressed in favorable neuroblastomas relative to unfavorable ones. During the all-trans-retinoic acid (ATRA)-mediated neuronal differentiation in neuroblastoma-derived RTBM1 cells, the expression levels of ERAP140/Nbla10993 increased at the mRNA level. Consistent with these observations, the luciferase reporter analysis demonstrated that the ERAP140/Nbla10993 promoter responds to ATRA. In addition, the immunoprecipitation/immunoblotting experiments showed that ERAP140 forms a stable complex with RARalpha but not with RXRalpha in cells, suggesting that ERAP140 is involved in RAR-mediated transcriptional regulation. Furthermore, the quantitative real-time PCR analysis using 109 primary neuroblastoma samples demonstrated that the expression levels of ERAP140/Nbla10993 significantly correlate with a better clinical outcome of neuroblastomas. Taken together, our present findings indicate that ERAP140/Nbla10993 plays an important role in the regulation of ATRA-mediated neuronal differentiation, and is a novel member of prognostic indicators for neuroblastoma. PMID:18497940

  15. Comparative studies on tree pollen allergens. X. Further purification and N-terminal amino acid sequence analyses of the major allergen of birch pollen (Betula verrucosa).

    PubMed

    Vik, H; Elsayed, S

    1986-01-01

    The previously isolated major allergen of birch pollen (fraction BV45), Int. Archs Allergy appl. Immun. 68: 70-78 (1982), was further purified by recycling chromatography. The purified preparation was run on a high-performance liquid chromatography (HPLC) TSK-G-2000 gel filtration chromatography column and, finally, on paper high-volt electrophoresis. The protein recovered met the homogeneity criteria required for performing the N-terminal sequence analysis. The allergenic and antigenic reactivities of the HPLC-purified protein, designated BV45B, was examined. A single homogeneous precipitation line in crossed immunoelectrophoresis (CIE) was shown. Specific IgE-inhibition tests and immuno-autoradiographic prints indicated that this allergen could bind reaginic IgE specificially and with good affinity. The homogeneity of BV45B was examined by isoelectric focusing (IEF). Several minor bands of pI differences of less than 0.1 units were visible, demonstrating the existence of some molecular variants of this protein. The N-terminal sequence analysis of the molecule was performed, and the following four amino acids were tentatively shown by sequential cleavage: NH2-Ala-Gly-Ile-Val-. The demonstration of one dominant N-terminal 1-dimethyl-amino-5-naphthalene sulphonyl (DNS)-amino acid by polyamide thin-layer chromatography at each sequence step confirmed that the N-terminal residue of the protein was not blocked; the heterogeneity shown by the IEF system was merely due to the presence of several homologous polymorphic proteins with identical N-terminal amino acid, the adequacy of the purification repertoire used. PMID:3957444

  16. Sequence analysis of the chitin synthase A gene of the Dutch elm pathogen Ophiostoma novo-ulmi indicates a close association with the human pathogen Sporothrix schenckii.

    PubMed

    Hintz, W E

    1999-09-01

    Degenerate oligonucleotide primers were designed according to conserved regions of the chitin synthase gene family and used to amplify a 621 basepair (bp) fragment from genomic DNA of Ophiostoma novo-ulmi, the causal agent of Dutch elm disease. The amplification product was used as a hybridization probe to screen a library of genomic DNA sequences and to retrieve a full-length chitin synthase gene (chsA). The putative coding region of the gene was 2619 bp long, lacked introns, and encoded a polypeptide of 873 amino acids. Based on the similarity of the predicted amino acid sequence to the full-length chsC gene of Aspergillus nidulans and chsA gene of Ampelomyces quisqualis, the O. novo-ulmi chsA was classified as a Class I chitin synthase. The phylogenies constructed, according to a subregion of all available chitin synthases, showed that O. novo-ulmi consistently clustered most closely with the human pathogen Sporothrix schenckii, recently classified as a member of the mitosporic Ophiostomataceae. Disruption of the chsA gene locus had no obvious effects on the growth or morphology of the fungus. PMID:10524253

  17. Sediment amino acids as indicators of anthropogenic activities and potential environmental risk in Erhai Lake, Southwest China.

    PubMed

    Ni, Zhaokui; Wang, Shengrui; Zhang, Mianmian

    2016-05-01

    Total hydrolysable amino acids (THAAs) constitute the most important fraction of labile nitrogen. Anthropogenic activities directly influence various biogeochemical cycles and then accelerate lake ecosystem deterioration. This is the first study that has established the relationship between sediment THAAs and anthropogenic activities using dated sediment cores, and evaluated the possibility of THAAs release at the sediment interface based on changes in environmental conditions in Erhai Lake. The results showed that historical distribution and fractions of THAAs could be divided into three stages: a stable period before the 1970s, a clear increasing period from the 1970s to 1990s, and a gradually steady period that started after the 1990s. The chemical fraction, aromatic and sulfur amino acids (AAs) accounted for only ≤3% of THAAs. Basic AAs accounted for 5-17% of THAAs, and remained at a relatively stable level. However, acidic and neutral AAs, which accounted for 19-44% and 35-69% of THAAs, respectively, were the predominant factors causing THAAs to increase due to rapid agricultural intensification and intensification of contemporary sedimentation of phytoplankton or macrophytes since the 1970s. These trends were closely related to both anthropogenic activities and natural processes, which implied that sediment THAAs could act as an effective indicator that reflects anthropogenic activities and aquatic environmental characteristics. The current contributions of sediment THAAs on TN and TOC were <5% and 1.5%, respectively. However, the dramatic increase in THAAs in the sediment cores indicated that there was a huge potential source of labile nitrogen for the overlying water under certain environmental conditions. Correlation analysis suggested that the release of THAAs was negatively correlated with pH, whereas positively correlated with bacterial number and degree of OM mineralization, which particularly depend on the stability of HFOM. Therefore, the risk of

  18. The phosphate clamp: sequence selective nucleic acid binding profiles and conformational induction of endonuclease inhibition by cationic Triplatin complexes

    PubMed Central

    Prisecaru, Andreea; Molphy, Zara; Kipping, Ralph G.; Peterson, Erica J.; Qu, Yun; Kellett, Andrew; Farrell, Nicholas P.

    2014-01-01

    The substitution-inert polynuclear platinum(II) complex (PPC) series, [{trans-Pt(NH3)2(NH2(CH2)nNH3)}2-μ-(trans-Pt(NH3)2(NH2(CH2)nNH2)2}](NO3)8, where n = 5 (AH78P), 6 (AH78 TriplatinNC) and 7 (AH78H), are potent non-covalent DNA binding agents where nucleic acid recognition is achieved through use of the ‘phosphate clamp' where the square-planar tetra-am(m)ine Pt(II) coordination units all form bidentate N–O–N complexes through hydrogen bonding with phosphate oxygens. The modular nature of PPC–DNA interactions results in high affinity for calf thymus DNA (Kapp ∼5 × 107 M−1). The phosphate clamp–DNA interactions result in condensation of superhelical and B-DNA, displacement of intercalated ethidium bromide and facilitate cooperative binding of Hoechst 33258 at the minor groove. The effect of linker chain length on DNA conformational changes was examined and the pentane-bridged complex, AH78P, was optimal for condensing DNA with results in the nanomolar region. Analysis of binding affinity and conformational changes for sequence-specific oligonucleotides by ITC, dialysis, ICP-MS, CD and 2D-1H NMR experiments indicate that two limiting modes of phosphate clamp binding can be distinguished through their conformational changes and strongly suggest that DNA condensation is driven by minor-groove spanning. Triplatin-DNA binding prevents endonuclease activity by type II restriction enzymes BamHI, EcoRI and SalI, and inhibition was confirmed through the development of an on-chip microfluidic protocol. PMID:25414347

  19. Fad7 gene identification and fatty acids phenotypic variation in an olive collection by EcoTILLING and sequencing approaches.

    PubMed

    Sabetta, Wilma; Blanco, Antonio; Zelasco, Samanta; Lombardo, Luca; Perri, Enzo; Mangini, Giacomo; Montemurro, Cinzia

    2013-08-01

    The ω-3 fatty acid desaturases (FADs) are enzymes responsible for catalyzing the conversion of linoleic acid to α-linolenic acid localized in the plastid or in the endoplasmic reticulum. In this research we report the genotypic and phenotypic variation of Italian Olea europaea L. germoplasm for the fatty acid composition. The phenotypic oil characterization was followed by the molecular analysis of the plastidial-type ω-3 FAD gene (fad7) (EC 1.14.19), whose full-length sequence has been here identified in cultivar Leccino. The gene consisted of 2635 bp with 8 exons and 5'- and 3'-UTRs of 336 and 282 bp respectively, and showed a high level of heterozygousity (1/110 bp). The natural allelic variation was investigated both by a LiCOR EcoTILLING assay and the PCR product direct sequencing. Only three haplotypes were identified among the 96 analysed cultivars, highlighting the strong degree of conservation of this gene. PMID:23685785

  20. Nucleotide sequences and organization of the genes for carotovoricin (Ctv) from Erwinia carotovora indicate that Ctv evolved from the same ancestor as Salmonella typhi prophage.

    PubMed

    Yamada, Kazuteru; Hirota, Morihiko; Niimi, Yoshiko; Nguyen, Hoa Anh; Takahara, Yoshiyuki; Kamio, Yoshiyuki; Kaneko, Jun

    2006-09-01

    Carotovoricin Er (CtvEr), which is produced by a plant soft rot disease causative agent, Erwinia carotovora subsp. carotovora Er, is a high-molecular-weight bacteriocin showing Myoviridae phage-tail-like morphology with contractile sheath and plural tail fibers. We determined the complete nucleotide sequences of CtvEr genes on the E. carotovora Er chromosome and report that CtvEr genes consist of lysis cassette, major and minor structural protein gene clusters. Four promoters were identified. The lysis gene cassette, which is composed of the genes for lysis enzyme and holin, was also identified and characterized. The nucleotide sequences and organization of the genes for CtvCGE, which is produced by E. carotovora strain CGE234-M403 with the morphology similar to CtvEr, were also determined and compared to that of CtvEr, and it was found that CtvCGE is almost identical to CtvEr except for tail fibers which are involved in the killing spectra of both bacteriocins. We also explain that the gene organization and the deduced amino acid sequences of both carotovoricins are very close to those of prophage, which is lysogenized in the chromosome on Salmonella enterica serovar Typhi CT18. These findings strongly suggest that Ctv evolved as a phage tail-like bacteriocin from a common ancestor with Salmonella typhi prophage. PMID:16960352

  1. Enzymatic generation of peptides flanked by basic amino acids to obtain MS/MS spectra with 2× sequence coverage

    PubMed Central

    Ebhardt, H Alexander; Nan, Jie; Chaulk, Steven G; Fahlman, Richard P; Aebersold, Ruedi

    2014-01-01

    RATIONALE Tandem mass (MS/MS) spectra generated by collision-induced dissociation (CID) typically lack redundant peptide sequence information in the form of e.g. b- and y-ion series due to frequent use of sequence-specific endopeptidases cleaving C- or N-terminal to Arg or Lys residues. METHODS Here we introduce arginyl-tRNA protein transferase (ATE, EC 2.3.2.8) for proteomics. ATE recognizes acidic amino acids or oxidized Cys at the N-terminus of a substrate peptide and conjugates an arginine from an aminoacylated tRNAArg onto the N-terminus of the substrate peptide. This enzymatic reaction is carried out under physiological conditions and, in combination with Lys-C/Asp-N double digest, results in arginylated peptides with basic amino acids on both termini. RESULTS We demonstrate that in vitro arginylation of peptides using yeast arginyl tRNA protein transferase 1 (yATE1) is a robust enzymatic reaction, specific to only modifying N-terminal acidic amino acids. Precursors originating from arginylated peptides generally have an increased protonation state compared with their non-arginylated forms. Furthermore, the product ion spectra of arginylated peptides show near complete 2× fragment ladders within the same MS/MS spectrum using commonly available electrospray ionization peptide fragmentation modes. Unexpectedly, arginylated peptides generate complete y- and c-ion series using electron transfer dissociation (ETD) despite having an internal proline residue. CONCLUSIONS We introduce a rapid enzymatic method to generate peptides flanked on either terminus by basic amino acids, resulting in a rich, redundant MS/MS fragment pattern. © 2014 The Authors. Rapid Communications in Mass Spectrometry published by John Wiley & Sons Ltd. PMID:25380496

  2. Site-directed gene mutation at mixed sequence targets by psoralen-conjugated pseudo-complementary peptide nucleic acids.

    PubMed

    Kim, Ki-Hyun; Nielsen, Peter E; Glazer, Peter M

    2007-01-01

    Sequence-specific DNA-binding molecules such as triple helix-forming oligonucleotides (TFOs) provide a means for inducing site-specific mutagenesis and recombination at chromosomal sites in mammalian cells. However, the utility of TFOs is limited by the requirement for homopurine stretches in the target duplex DNA. Here, we report the use of pseudo-complementary peptide nucleic acids (pcPNAs) for intracellular gene targeting at mixed sequence sites. Due to steric hindrance, pcPNAs are unable to form pcPNA-pcPNA duplexes but can bind to complementary DNA sequences by Watson-Crick pairing via double duplex-invasion complex formation. We show that psoralen-conjugated pcPNAs can deliver site-specific photoadducts and mediate targeted gene modification within both episomal and chromosomal DNA in mammalian cells without detectable off-target effects. Most of the induced psoralen-pcPNA mutations were single-base substitutions and deletions at the predicted pcPNA-binding sites. The pcPNA-directed mutagenesis was found to be dependent on PNA concentration and UVA dose and required matched pairs of pcPNAs. Neither of the individual pcPNAs alone had any effect nor did complementary PNA pairs of the same sequence. These results identify pcPNAs as new tools for site-specific gene modification in mammalian cells without purine sequence restriction, thereby providing a general strategy for designing gene targeting molecules. PMID:17977869

  3. Comparison of the nucleotide and amino acid sequences of the RsrI and EcoRI restriction endonucleases.

    PubMed

    Stephenson, F H; Ballard, B T; Boyer, H W; Rosenberg, J M; Greene, P J

    1989-12-21

    The RsrI endonuclease, a type-II restriction endonuclease (ENase) found in Rhodobacter sphaeroides, is an isoschizomer of the EcoRI ENase. A clone containing an 11-kb BamHI fragment was isolated from an R. sphaeroides genomic DNA library by hybridization with synthetic oligodeoxyribonucleotide probes based on the N-terminal amino acid (aa) sequence of RsrI. Extracts of E. coli containing a subclone of the 11-kb fragment display RsrI activity. Nucleotide sequence analysis reveals an 831-bp open reading frame encoding a polypeptide of 277 aa. A 50% identity exists within a 266-aa overlap between the deduced aa sequences of RsrI and EcoRI. Regions of 75-100% aa sequence identity correspond to key structural and functional regions of EcoRI. The type-II ENases have many common properties, and a common origin might have been expected. Nevertheless, this is the first demonstration of aa sequence similarity between ENases produced by different organisms. PMID:2695392

  4. Invasive cleavage of nucleic acids

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann D.; Dahlberg, James E.

    1999-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  5. Invasive cleavage of nucleic acids

    DOEpatents

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann D.; Dahlberg, James E.

    2002-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  6. Complete amino acid sequence of human plasma Zn-. cap alpha. /sub 2/-glycoprotein and its homology to histocompatibility antigens

    SciTech Connect

    Araki, T.; Gejyo, F.; Takagaki, K.; Haupt, H.; Schwick, H.G.; Buergi, W.; Marti, T.; Schaller, J.; Rickli, E.; Brossmer, R.

    1988-02-01

    In the present study the complete amino acid sequence of human plasma Zn-..cap alpha../sub 2/-glycoprotein was determined. This protein whose biological function is unknown consists of a single polypeptide chain of 276 amino acid residues including 8 tryptophan residues and has a pyroglutamyl residue at the amino terminus. The location of the two disulfide bonds in the polypeptide chain was also established. The three glycans, whose structure was elucidated with the aid of 500 MHz /sup 1/H NMR spectroscopy, were sialylated N-biantennas. The molecular weight calculated from the polypeptide and carbohydrate structure is 38,478, which is close to the reported value of approx. = 41,000 based on physicochemical measurements. The predicted secondary structure appeared to comprised of 23% ..cap alpha..-helix, 27% ..beta..-sheet, and 22% ..beta..-turns. The three N-glycans were found to be located in ..beta..-turn regions. An unexpected finding was made by computer analysis of the sequence data; this revealed that Zn-..cap alpha../sub 2/-glycoprotein is closely related to antigens of the major histocompatibility complex in amino acid sequence and in domain structure. There was an unusually high degree of sequence homology with the ..cap alpha.. chains of class I histocompatibility antigens. Moreover, this plasma protein was shown to be a member of the immunoglobulin gene superfamily. Zn-..cap alpha../sub 2/-glycoprotein appears to be truncated secretory major histocompatibility complex-related molecule, and it may have a role in the expression of the immune response.

  7. ENTPRISE: An Algorithm for Predicting Human Disease-Associated Amino Acid Substitutions from Sequence Entropy and Predicted Protein Structures

    PubMed Central

    Zhou, Hongyi; Gao, Mu; Skolnick, Jeffrey

    2016-01-01

    The advance of next-generation sequencing technologies has made exome sequencing rapid and relatively inexpensive. A major application of exome sequencing is the identification of genetic variations likely to cause Mendelian diseases. This requires processing large amounts of sequence information and therefore computational approaches that can accurately and efficiently identify the subset of disease-associated variations are needed. The accuracy and high false positive rates of existing computational tools leave much room for improvement. Here, we develop a boosted tree regression machine-learning approach to predict human disease-associated amino acid variations by utilizing a comprehensive combination of protein sequence and structure features. On comparing our method, ENTPRISE, to the state-of-the-art methods SIFT, PolyPhen-2, MUTATIONASSESSOR, MUTATIONTASTER, FATHMM, ENTPRISE exhibits significant improvement. In particular, on a testing dataset consisting of only proteins with balanced disease-associated and neutral variations defined as having the ratio of neutral/disease-associated variations between 0.3 and 3, the Mathews Correlation Coefficient by ENTPRISE is 0.493 as compared to 0.432 by PPH2-HumVar, 0.406 by SIFT, 0.403 by MUTATIONASSESSOR, 0.402 by PPH2-HumDiv, 0.305 by MUTATIONTASTER, and 0.181 by FATHMM. ENTPRISE is then applied to nucleic acid binding proteins in the human proteome. Disease-associated predictions are shown to be highly correlated with the number of protein-protein interactions. Both these predictions and the ENTPRISE server are freely available for academic users as a web service at http://cssb.biology.gatech.edu/entprise/. PMID:26982818

  8. Fatty acid profile and Unigene-derived simple sequence repeat markers in tung tree (Vernicia fordii)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Tung tree (Vernicia fordii) provides the sole source of tung oil widely used in industry. Lack of fatty acid composition and molecular markers hinders biochemical, genetic and breeding research. The objectives of this study were to determine fatty acid profiles and develop unigene-derived simple se...

  9. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... Director of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 51. Copies of WIPO... 37 Patents, Trademarks, and Copyrights 1 2010-07-01 2010-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid...

  10. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Director of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 51. Copies of WIPO... 37 Patents, Trademarks, and Copyrights 1 2011-07-01 2011-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid...

  11. Indications of human activity from amino acid and amino sugar analyses on Holocene sediments from lake Lonar, central India

    NASA Astrophysics Data System (ADS)

    Menzel, P.; Gaye, B.; Wiesner, M.; Prasad, S.; Basavaiah, N.; Stebich, M.; Anoop, A.; Riedel, N.; Brauer, A.

    2012-04-01

    The DFG funded HIMPAC (Himalaya: Modern and Past Climates) programme aims to reconstruct Holocene Indian Monsoon climate using a multi-proxy and multi-archive approach. First investigations made on sediments from a ca. 10 m long core covering the whole Holocene taken from the lake Lonar in central India's state Maharashtra, Buldhana District, serve to identify changes in sedimentation, lake chemistry, local vegetation and regional to supra-regional climate patterns. Lake Lonar occupies the floor of an impact crater that formed on the ~ 65 Ma old basalt flows of the Deccan Traps. It covers an area of ca. 1 km2 and is situated in India's core monsoon area. The modern lake has a maximum depth of about 5 m, is highly alkaline, and hyposaline, grouped in the Na-Cl-CO3 subtype of saline lakes. No out-flowing stream is present and only three small streams feed the lake, resulting in a lake level highly sensitive to precipitation and evaporation. The lake is eutrophic and stratified throughout most of the year with sub- to anoxic waters below 2 m depth. In this study the core sediments were analysed for their total amino acid (AA) and amino sugar (AS) content, the amino acid bound C and N percentage of organic C and total N in the sediment and the distribution of individual amino acids. The results roughly show three zones within the core separated by distinct changes in their AA content and distribution. (i) The bottom part of the core from ca. 12000 cal a BP to 11400 cal a BP with very low AA and AS percentage indicating high lithogenic contribution, most probably related to dry conditions. (ii) From 11400 cal a BP to 1200 cal a BP the sediments show moderate AA and AS percentages and low values for the ratios of proteinogenic AAs to their non-proteinogenic degradation products (e.g. ASP/β-ALA; GLU/γ-ABA). (iii) The top part of the core (< 1200 cal a BP) is characterised by an intense increase in total AA and AS, AA-C/Corg and AA-N/Ntotas well as in the ratio of

  12. Shared Segment Analysis and Next-Generation Sequencing Implicates the Retinoic Acid Signaling Pathway in Total Anomalous Pulmonary Venous Return (TAPVR)

    PubMed Central

    Nash, Dustin; Arrington, Cammon B.; Kennedy, Brett J.; Yandell, Mark; Wu, Wilfred; Zhang, Wenying; Ware, Stephanie; Jorde, Lynn B.; Gruber, Peter J.; Yost, H. Joseph

    2015-01-01

    Most isolated congenital heart defects are thought to be sporadic and are often ascribed to multifactorial mechanisms with poorly understood genetics. Total Anomalous Pulmonary Venous Return (TAPVR) occurs in 1 in 15,000 live-born infants and occurs either in isolation or as part of a syndrome involving aberrant left-right development. Previously, we reported causative links between TAVPR and the PDGFRA gene. TAPVR has also been linked to the ANKRD1/CARP genes. However, these genes only explain a small fraction of the heritability of the condition. By examination of phased single nucleotide polymorphism genotype data from 5 distantly related TAPVR patients we identified a single 25 cM shared, Identical by Descent genomic segment on the short arm of chromosome 12 shared by 3 of the patients and their obligate-carrier parents. Whole genome sequence (WGS) analysis identified a non-synonymous variant within the shared segment in the retinol binding protein 5 (RBP5) gene. The RBP5 variant is predicted to be deleterious and is overrepresented in the TAPVR population. Gene expression and functional analysis of the zebrafish orthologue, rbp7, supports the notion that RBP5 is a TAPVR susceptibility gene. Additional sequence analysis also uncovered deleterious variants in genes associated with retinoic acid signaling, including NODAL and retinol dehydrogenase 10. These data indicate that genetic variation in the retinoic acid signaling pathway confers, in part, susceptibility to TAPVR. PMID:26121141

  13. Halogen bonding from a hard and soft acids and bases perspective: investigation by using density functional theory reactivity indices.

    PubMed

    Pinter, Balazs; Nagels, Nick; Herrebout, Wouter A; De Proft, Frank

    2013-01-01

    Halogen bonds between the trifluoromethyl halides CF(3)Cl, CF(3)Br and CF(3)I, and dimethyl ether, dimethyl sulfide, trimethylamine and trimethyl phosphine were investigated using Pearson's hard and soft acids and bases (HSAB) concept with conceptual DFT reactivity indices, the Ziegler-Rauk-type energy-decomposition analysis, the natural orbital for chemical valence (NOCV) framework and the non-covalent interaction (NCI) index. It is found that the relative importance of electrostatic and orbital (charge transfer) interactions varies as a function of both the donor and acceptor molecules. Hard and soft interactions were distinguished and characterised by atomic charges, electrophilicity and local softness indices. Dual-descriptor plots indicate an orbital σ hole on the halogen similar to the electrostatic σ hole manifested in the molecular electrostatic potential. The predicted high halogen-bond-acceptor affinity of N-heterocyclic carbenes was evidenced in the highest complexation energy for the hitherto unknown CF(3) I·NHC complex. The dominant NOCV orbital represents an electron-density deformation according to a n→σ*-type interaction. The characteristic signal found in the reduced density gradient versus electron-density diagram corresponds to the non-covalent interaction between contact atoms in the NCI plots, which is the manifestation of halogen bonding within the NCI theory. The unexpected C-X bond strengthening observed in several cases was rationalised within the molecular orbital framework. PMID:23169478

  14. "De-novo" amino acid sequence elucidation of protein G'e by combined "Top-Down" and "Bottom-Up" mass spectrometry

    NASA Astrophysics Data System (ADS)

    Yefremova, Yelena; Al-Majdoub, Mahmoud; Opuni, Kwabena F. M.; Koy, Cornelia; Cui, Weidong; Yan, Yuetian; Gross, Michael L.; Glocker, Michael O.

    2015-03-01

    Mass spectrometric de-novo sequencing was applied to review the amino acid sequence of a commercially available recombinant protein Ǵ with great scientific and economic importance. Substantial deviations to the published amino acid sequence (Uniprot Q54181) were found by the presence of 46 additional amino acids at the N-terminus, including a so-called "His-tag" as well as an N-terminal partial α- N-gluconoylation and α- N-phosphogluconoylation, respectively. The unexpected amino acid sequence of the commercial protein G' comprised 241 amino acids and resulted in a molecular mass of 25,998.9 ± 0.2 Da for the unmodified protein. Due to the higher mass that is caused by its extended amino acid sequence compared with the original protein G' (185 amino acids), we named this protein "protein G'e." By means of mass spectrometric peptide mapping, the suggested amino acid sequence, as well as the N-terminal partial α- N-gluconoylations, was confirmed with 100% sequence coverage. After the protein G'e sequence was determined, we were able to determine the expression vector pET-28b from Novagen with the Xho I restriction enzyme cleavage site as the best option that was used for cloning and expressing the recombinant protein G'e in E. coli. A dissociation constant ( K d ) value of 9.4 nM for protein G'e was determined thermophoretically, showing that the N-terminal flanking sequence extension did not cause significant changes in the binding affinity to immunoglobulins.

  15. "De-novo" amino acid sequence elucidation of protein G'e by combined "top-down" and "bottom-up" mass spectrometry.

    PubMed

    Yefremova, Yelena; Al-Majdoub, Mahmoud; Opuni, Kwabena F M; Koy, Cornelia; Cui, Weidong; Yan, Yuetian; Gross, Michael L; Glocker, Michael O

    2015-03-01

    Mass spectrometric de-novo sequencing was applied to review the amino acid sequence of a commercially available recombinant protein G´ with great scientific and economic importance. Substantial deviations to the published amino acid sequence (Uniprot Q54181) were found by the presence of 46 additional amino acids at the N-terminus, including a so-called "His-tag" as well as an N-terminal partial α-N-gluconoylation and α-N-phosphogluconoylation, respectively. The unexpected amino acid sequence of the commercial protein G' comprised 241 amino acids and resulted in a molecular mass of 25,998.9 ± 0.2 Da for the unmodified protein. Due to the higher mass that is caused by its extended amino acid sequence compared with the original protein G' (185 amino acids), we named this protein "protein G'e." By means of mass spectrometric peptide mapping, the suggested amino acid sequence, as well as the N-terminal partial α-N-gluconoylations, was confirmed with 100% sequence coverage. After the protein G'e sequence was determined, we were able to determine the expression vector pET-28b from Novagen with the Xho I restriction enzyme cleavage site as the best option that was used for cloning and expressing the recombinant protein G'e in E. coli. A dissociation constant (K(d)) value of 9.4 nM for protein G'e was determined thermophoretically, showing that the N-terminal flanking sequence extension did not cause significant changes in the binding affinity to immunoglobulins. PMID:25560987

  16. Dynamic behavior of an intrinsically unstructured linker domain is conserved in the face of negligible amino acid sequence conservation.

    PubMed

    Daughdrill, Gary W; Narayanaswami, Pranesh; Gilmore, Sara H; Belczyk, Agniezka; Brown, Celeste J

    2007-09-01

    Proteins or regions of proteins that do not form compact globular structures are classified as intrinsically unstructured proteins (IUPs). IUPs are common in nature and have essential molecular functions, but even a limited understanding of the evolution of their dynamic behavior is lacking. The primary objective of this work was to test the evolutionary conservation of dynamic behavior for a particular class of IUPs that form intrinsically unstructured linker domains (IULD) that tether flanking folded domains. This objective was accomplished by measuring the backbone flexibility of several IULD homologues using nuclear magnetic resonance (NMR) spectroscopy. The backbone flexibility of five IULDs, representing three kingdoms, was measured and analyzed. Two IULDs from animals, one IULD from fungi, and two IULDs from plants showed similar levels of backbone flexibility that were consistent with the absence of a compact globular structure. In contrast, the amino acid sequences of the IULDs from these three taxa showed no significant similarity. To investigate how the dynamic behavior of the IULDs could be conserved in the absence of detectable sequence conservation, evolutionary rate studies were performed on a set of nine mammalian IULDs. The results of this analysis showed that many sites in the IULD are evolving neutrally, suggesting that dynamic behavior can be maintained in the absence of natural selection. This work represents the first experimental test of the evolutionary conservation of dynamic behavior and demonstrates that amino acid sequence conservation is not required for the conservation of dynamic behavior and presumably molecular function. PMID:17721672

  17. Nucleotide sequence of the fadR gene, a multifunctional regulator of fatty acid metabolism in Escherichia coli.

    PubMed Central

    DiRusso, C C

    1988-01-01

    The Escherichia coli fadR gene is a multifunctional regulator of fatty acid and acetate metabolism. In the present work the nucleotide sequence of the 1.3 kb DNA fragment which encodes FadR has been determined. The coding sequence of the fadR gene is 714 nucleotides long and is preceded by a typical E. coli ribosome binding site and is followed by a sequence predicted to be sufficient for factor-independent chain termination. Primer extension experiments demonstrated that the transcription of the fadR gene initiates with an adenine nucleotide 33 nucleotides upstream from the predicted start of translation. The derived fadR peptide has a calculated molecular weight of 26,972. This is in reasonable agreement with the apparent molecular weight of 29,000 previously estimated on the basis of maxi-cell analysis of plasmid encoded proteins. There is a segment of twenty amino acids within the predicted peptide which resembles the DNA recognition and binding site of many transcriptional regulatory proteins. Images PMID:2843809

  18. The amino acid sequence of protein SCMK-B2C from the high-sulphur fraction of wool keratin

    PubMed Central

    Elleman, T. C.

    1972-01-01

    1. The amino acid sequence of a protein from the reduced and carboxymethylated high-sulphur fraction of wool has been determined. 2. The sequence of this S-carboxymethylkerateine (SCMK-B2C) of 151 amino acid residues displays much internal homology and an unusual residue distribution. Thus a ten-residue sequence occurs four times near the N-terminus and five times near the C-terminus with few changes. These regions contain much of the molecule's half-cystine, whereas between them there is a region of 19 residues that are mainly small and devoid of cystine and proline. 3. Certain models of the wool fibre based on its mechanical and physical properties propose a matrix of small compact globular units linked together to form beaded chains. The unusual distribution of the component residues of protein SCMK-B2C suggests structures in the wool-fibre matrix compatible with certain features of the proposed models. PMID:4678578

  19. Validating a Stability Indicating HPLC Method for Kinetic Study of Cetirizine Degradation in Acidic and Oxidative Conditions

    PubMed Central

    Souri, Effat; Hatami, Ali; Shabani Ravari, Nazanin; Alvandifar, Farhad; Barazandeh Tehrani, Maliheh

    2013-01-01

    A stability indicating High-Performance Liquid Chromatography (HPLC) method was validated and used to study the degradation of cetirizine dihydrochloride in acidic and oxidative conditions. The separation was carried out on a Symmetry C18 column and a mixture of 50 mM KH2PO4 and acetonitrile (60:40 v/v, pH = 3.5) was used as the mobile phase. The method was linear over the range of 1-20 μg/mL of cetirizine dihydrochloride (r2 > 0.999) and the within-day and between-day precision values were less than 1.5%. The results showed that cetirizine dihydrochloride was unstable in 2 M HCl and 0.5% H2O2. The kinetics of the acidic degradation showed a pseudo-first-order reaction in the temperature range of 70-90°C. In addition, the kinetics of hydrogen peroxide mediated degradation was pseudo-first-order in the temperature range of 50-80°C. PMID:24250602

  20. Heuman indices of hydrophobicity of bile acids and their comparison with a newly developed and conventional molecular descriptors.

    PubMed

    Poša, Mihalj

    2014-02-01

    Bile salts (BSs), in addition to their physiological role in the digestion of lipids in vertebrates, are also of significant importance in biomedical investigations. For predicting biological-pharmacological activity and physico-chemical properties of BSs it is important to develop such molecular descriptors that adequately describe the structural characteristics of the steroid skeleton. The present study encompassed the following bile acids (BAs): cholic, chenodeoxycholic, deoxycholic, hyodeoxycholic, ursodeoxycholic, hyocholic, and ursocholic acid, as well as oxo derivatives of certain BAs. For all of them, Heuman hydrophobicity indices (HI(BA)) (RP-HPLC parameters) were determined, and a detailed conformational analysis of the steroid skeleton showed that HI(BA) has the discrimination power for BAs based on the size of the hydrophobic surface on the β side and the lateral L7 and L12 sides of the steroid skeleton. Also, HI(BA) discerns the regiochemical characteristics of OH and oxo groups. Based on a survey of the structural factors of the steroid skeleton that influence the HI(BA) values of the tested BAs, we constructed a new molecular descriptor, CHIBA, with the characteristics of 2D and 3D topological descriptors. In respect of the structure of the steroid skeleton, the descriptor CHIBA behaves as a reversed-phase chromatographic descriptor of BAs. PMID:24076126

  1. Advanced stability indicating chemometric methods for quantitation of amlodipine and atorvastatin in their quinary mixture with acidic degradation products

    NASA Astrophysics Data System (ADS)

    Darwish, Hany W.; Hassan, Said A.; Salem, Maissa Y.; El-Zeany, Badr A.

    2016-02-01

    Two advanced, accurate and precise chemometric methods are developed for the simultaneous determination of amlodipine besylate (AML) and atorvastatin calcium (ATV) in the presence of their acidic degradation products in tablet dosage forms. The first method was Partial Least Squares (PLS-1) and the second was Artificial Neural Networks (ANN). PLS was compared to ANN models with and without variable selection procedure (genetic algorithm (GA)). For proper analysis, a 5-factor 5-level experimental design was established resulting in 25 mixtures containing different ratios of the interfering species. Fifteen mixtures were used as calibration set and the other ten mixtures were used as validation set to validate the prediction ability of the suggested models. The proposed methods were successfully applied to the analysis of pharmaceutical tablets containing AML and ATV. The methods indicated the ability of the mentioned models to solve the highly overlapped spectra of the quinary mixture, yet using inexpensive and easy to handle instruments like the UV-VIS spectrophotometer.

  2. Single Amino Acid Substitutions in the Chemotactic Sequence of Urokinase Receptor Modulate Cell Migration and Invasion

    PubMed Central

    Franco, Paola; Pavone, Vincenzo; Mugione, Pietro; Di Carluccio, Gioconda; Masucci, Maria Teresa; Arra, Claudio; Pirozzi, Giuseppe; Stoppelli, Maria Patrizia; Carriero, Maria Vincenza

    2012-01-01

    The receptor for urokinase-type plasminogen activator (uPAR) plays an important role in controlling cell migration. uPAR binds urokinase and vitronectin extracellular ligands, and signals in complex with transmembrane receptors such as Formyl-peptide Receptors (FPR)s and integrins. Previous work from this laboratory has shown that synthetic peptides, corresponding to the uPAR88–92 chemotactic sequence, when carrying the S90P or S90E substitutions, up- or down-regulate cell migration, respectively. To gain mechanistic insights into these opposite cell responses, the functional consequences of S90P and S90E mutations in full-length uPAR were evaluated. First, (HEK)-293 embryonic kidney cells expressing uPARS90P exhibit enhanced FPR activation, increased random and directional cell migration, long-lasting Akt phosphorylation, and increased adhesion to vitronectin, as well as uPAR/vitronectin receptor association. In contrast, the S90E substitution prevents agonist-triggered FPR activation and internalization, decreases binding and adhesion to vitronectin, and inhibits uPAR/vitronectin receptor association. Also, 293/uPARS90P cells appear quite elongated and their cytoskeleton well organized, whereas 293/uPARS90E cells assume a large flattened morphology, with random orientation of actin filaments. Interestingly, when HT1080 cells co-express wild type uPAR with uPAR S90E, the latter behaves as a dominant-negative, impairing uPAR-mediated signaling and reducing cell wound repair as well as lung metastasis in nude mice. In contrast, signaling, wound repair and in vivo lung metastasis of HT1080 cells bearing wild type uPAR are enhanced when they co-express uPARS90P. In conclusion, our findings indicate that Ser90 is a critical residue for uPAR signaling and that the S90P and S90E exert opposite effects on uPAR activities. These findings may be accommodated in a molecular model, in which uPARS90E and uPARS90P are forced into inactive and active forms, respectively

  3. The nucleotide sequence of HLA-B{sup *}2704 reveals a new amino acid substitution in exon 4 which is also present in HLA-B{sup *}2706

    SciTech Connect

    Rudwaleit, M.; Bowness, P.; Wordsworth, P.

    1996-12-31

    The HLA-B27 subtype HLA-B{sup *}2704 is virtually absent in Caucasians but common in Orientals, where it is associated with ankylosing spondylitis. The amino acid sequence of HLA-B{sup *}2704 has been established by peptide mapping and was shown to differ by two amino acids from HLA-B{sup *}2705, HLA-B{sup *}2704 is characterized by a serine for aspartic acid substitution at position 77 and glutamic acid for valine at position 152. To date, however, no nucleotide sequence confirming these changes at the DNA level has been published. 13 refs., 2 figs.

  4. JRC GMO-Amplicons: a collection of nucleic acid sequences related to genetically modified organisms.

    PubMed

    Petrillo, Mauro; Angers-Loustau, Alexandre; Henriksson, Peter; Bonfini, Laura; Patak, Alex; Kreysa, Joachim

    2015-01-01

    The DNA target sequence is the key element in designing detection methods for genetically modified organisms (GMOs). Unfortunately this information is frequently lacking, especially for unauthorized GMOs. In addition, patent sequences are generally poorly annotated, buried in complex and extensive documentation and hard to link to the corresponding GM event. Here, we present the JRC GMO-Amplicons, a database of amplicons collected by screening public nucleotide sequence databanks by in silico determination of PCR amplification with reference methods for GMO analysis. The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF) provides these methods in the GMOMETHODS database to support enforcement of EU legislation and GM food/feed control. The JRC GMO-Amplicons database is composed of more than 240 000 amplicons, which can be easily accessed and screened through a web interface. To our knowledge, this is the first attempt at pooling and collecting publicly available sequences related to GMOs in food and feed. The JRC GMO-Amplicons supports control laboratories in the design and assessment of GMO methods, providing inter-alia in silico prediction of primers specificity and GM targets coverage. The new tool can assist the laboratories in the analysis of complex issues, such as the detection and identification of unauthorized GMOs. Notably, the JRC GMO-Amplicons database allows the retrieval and characterization of GMO-related sequences included in patents documentation. Finally, it can help annotating poorly described GM sequences and identifying new relevant GMO-related sequences in public databases. The JRC GMO-Amplicons is freely accessible through a web-based portal that is hosted on the EU-RL GMFF website. Database URL: http://gmo-crl.jrc.ec.europa.eu/jrcgmoamplicons/. PMID:26424080

  5. JRC GMO-Amplicons: a collection of nucleic acid sequences related to genetically modified organisms

    PubMed Central

    Petrillo, Mauro; Angers-Loustau, Alexandre; Henriksson, Peter; Bonfini, Laura; Patak, Alex; Kreysa, Joachim

    2015-01-01

    The DNA target sequence is the key element in designing detection methods for genetically modified organisms (GMOs). Unfortunately this information is frequently lacking, especially for unauthorized GMOs. In addition, patent sequences are generally poorly annotated, buried in complex and extensive documentation and hard to link to the corresponding GM event. Here, we present the JRC GMO-Amplicons, a database of amplicons collected by screening public nucleotide sequence databanks by in silico determination of PCR amplification with reference methods for GMO analysis. The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF) provides these methods in the GMOMETHODS database to support enforcement of EU legislation and GM food/feed control. The JRC GMO-Amplicons database is composed of more than 240 000 amplicons, which can be easily accessed and screened through a web interface. To our knowledge, this is the first attempt at pooling and collecting publicly available sequences related to GMOs in food and feed. The JRC GMO-Amplicons supports control laboratories in the design and assessment of GMO methods, providing inter-alia in silico prediction of primers specificity and GM targets coverage. The new tool can assist the laboratories in the analysis of complex issues, such as the detection and identification of unauthorized GMOs. Notably, the JRC GMO-Amplicons database allows the retrieval and characterization of GMO-related sequences included in patents documentation. Finally, it can help annotating poorly described GM sequences and identifying new relevant GMO-related sequences in public databases. The JRC GMO-Amplicons is freely accessible through a web-based portal that is hosted on the EU-RL GMFF website. Database URL: http://gmo-crl.jrc.ec.europa.eu/jrcgmoamplicons/ PMID:26424080

  6. The Pietra Grande thrust (Brenta Dolomites, Italy): looking for co-seismic indicators along a main fault in carbonate sequences

    NASA Astrophysics Data System (ADS)

    Viganò, Alfio; Tumiati, Simone; Martin, Silvana; Rigo, Manuel

    2013-04-01

    At present, pseudotachylytes (i.e. solidified frictional melts) are the only unambiguous geological record of seismic faulting. Even if pseudotachylytes are frequently observed along faults within crystalline rocks they are discovered along carbonate faults in very few cases only, suggesting that other chemico-physical processes than melting could occur (e.g. thermal decomposition). In order to investigate possible co-seismic indicators we study the Pietra Grande thrust, a carbonate fault in the Brenta Dolomites (Trentino, NE Italy), to analyse field structure, microtextures and composition of rocks from the principal slip plane, the fault core and the damage zone. The Pietra Grande thrust is developed within limestones and dolomitic limestones of Late Triassic-Early Jurassic age (Calcari di Zu and Monte Zugna Formations). The thrust, interpreted as a north-vergent décollement deeply connected with the major Cima Tosa thrust, is a sub-horizontal fault plane gently dipping to the North that mainly separates the massive Monte Zugna Fm. limestones (upper side) from the stratified Calcari di Zu Fm. limestones with intercalated marls (lower side). On the western face of the Pietra Grande klippe the thrust is continuously well-exposed for about 1 km. The main fault plane shows reddish infillings, which form veins with thicknesses between few millimetres to several decimetres. These red veins lie parallel to the thrust plane or in same cases inject lateral fractures and minor high-angle faults departing from the main fault plane. Veins have carbonate composition and show textures characterized by fine-grained reddish matrix with embedded carbonate clasts of different size (from few millimetres to centimetres). In some portions carbonate boulders (dimension of some decimetres) are embedded in the red matrix, while clast content generally significantly decreases at the vein borders (chilled margins). Red veins are typically associated with cohesive cataclasites and

  7. A molecular mechanism realizing sequence-specific recognition of nucleic acids by TDP-43

    PubMed Central

    Furukawa, Yoshiaki; Suzuki, Yoh; Fukuoka, Mami; Nagasawa, Kenichi; Nakagome, Kenta; Shimizu, Hideaki; Mukaiyama, Atsushi; Akiyama, Shuji

    2016-01-01

    TAR DNA-binding protein 43 (TDP-43) is a DNA/RNA-binding protein containing two consecutive RNA recognition motifs (RRM1 and RRM2) in tandem. Functional abnormality of TDP-43 has been proposed to cause neurodegeneration, but it remains obscure how the physiological functions of this protein are regulated. Here, we show distinct roles of RRM1 and RRM2 in the sequence-specific substrate recognition of TDP-43. RRM1 was found to bind a wide spectrum of ssDNA sequences, while no binding was observed between RRM2 and ssDNA. When two RRMs are fused in tandem as in native TDP-43, the fused construct almost exclusively binds ssDNA with a TG-repeat sequence. In contrast, such sequence-specificity was not observed in a simple mixture of RRM1 and RRM2. We thus propose that the spatial arrangement of multiple RRMs in DNA/RNA binding proteins provides steric effects on the substrate-binding site and thereby controls the specificity of its substrate nucleotide sequences. PMID:26838063

  8. Application of combined mass spectrometry and partial amino acid sequence to the identification of gel-separated proteins.

    PubMed

    Patterson, S D; Thomas, D; Bradshaw, R A

    1996-05-01

    The combined use of peptide mass information with amino acid sequence information derived by chemical sequencing or mass spectrometry (MS)-based approaches provides a powerful means of protein identification. We have used a two-part strategy to identify proteins from nerve growth factor (NGF)-stimulated rat adrenal pheochromocytoma cell line PC-12 cell lysates that associate with the adaptor protein Shc (Shc homologous and collagen protein). Initial experiments with metabolically radiolabeled cell extracts separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a number of proteins that coimmunoprecipitated with anti-Shc antibody compared with control (unstimulated) cell extracts. The experiment was scaled up and cell lysate from NGF-stimulated PC-12 cells was applied to a glutathione-S-transferase (GST)-Shc affinity column, eluted, separated by SDS-PAGE and blotted to Immobilon-CD. The blotted proteins were proteolytically digested in situ, and the masses obtained from the extracted peptides were used in a peptide-mass search program in an attempt to identify the protein. Even if a strong candidate was found using this search, an additional step was performed to confirm the identification. The mixtures were fractionated by reversed-phase high-performance liquid chromatography (RP-HPLC) and subjected to chemical sequencing to obtain (partial) sequence information, or post-source decay (PSD-) matrix-assisted laser-desorption ionization (MALDI)-MS to obtain sequence-specific fragment ions. This data was used in a peptide-sequence tag search to confirm the identity of the proteins. This combined approach allowed identification of four proteins of M(r) 43,000 to 200,000. In one case the identified protein clearly did not correspond to the radiolabeled band, but to a protein contaminant from the column. The advantages and pitfalls of the approach are discussed. PMID:8783013

  9. Peptide mapping and amino acid sequencing of two catechol 1,2-dioxygenases (CD I1 and CD I2) from Acinetobacter lwoffii K24.

    PubMed

    Kim, S I; Ha, K S

    1997-10-31

    The partial amino acid sequences of two catechol 1,2-dioxygenases (CD I1 and CD I2) from Acinetobacter lwoffii K24 have been determined by analysis of peptides after cleavages with endopeptidase Lys-C, endopeptidase Glu-C, trypsin, and chemicals (cyanogen bromide and BNPS-skatole). They include 248 amino acid sequences (4 fragments) of CD I1 and 211 amino acid sequences (5 fragments) of CD I2. Two enzymes have more than 50% sequence homology with type I catechol 1,2-dioxygenases and less than 30% sequence homology with type II catechol 1,2-dioxygenases. Two enzymes have similar hydropathy profiles in the N-terminal region, suggesting that they have similar secondary structures. PMID:9387151

  10. The Role of HIV-1 gp41 Glycoprotein in Infectious Tropism Inferred from Physico-Chemical Properties of its Amino Acid Sequence

    NASA Astrophysics Data System (ADS)

    Figueroa, E.; Villarreal, C.; Huerta, L.; Cocho, G.

    2006-09-01

    We performed a statistical analysis of the amino acid sequence of the gp41 ectodomain of the Human Immunodeficiency Virus type 1. We found strong correlations between physicochemical properties of highly variable residues and the viral infectious tropism.

  11. Characterization of fatty acid-producing wastewater microbial communities using next generation sequencing technologies

    EPA Science Inventory

    While wastewater represents a viable source of bacterial biodiesel production, very little is known on the composition of these microbial communities. We studied the taxonomic diversity and succession of microbial communities in bioreactors accumulating fatty acids using 454-pyro...

  12. Complete Genome Sequence of Amino Acid-Utilizing Eubacterium acidaminophilum al-2 (DSM 3953)

    PubMed Central

    Poehlein, Anja; Andreesen, Jan R.

    2014-01-01

    Eubacterium acidaminophilum is a strictly anaerobic, Gram-positive, rod-shaped bacterium which belongs to cluster XI of the Clostridia. It ferments amino acids by a Stickland reaction. The genome harbors a chromosome (2.25 Mb) and a megaplasmid (0.8 Mb). It contains several gene clusters coding for selenocysteine-containing, glycine-derived, and amino acid-degrading reductases. PMID:24926057

  13. Nucleic acid detection kits

    DOEpatents

    Hall, Jeff G.; Lyamichev, Victor I.; Mast, Andrea L.; Brow, Mary Ann; Kwiatkowski, Robert W.; Vavra, Stephanie H.

    2005-03-29

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of nucleic acid from various viruses in a sample.

  14. Effects of the amino acid sequence on thermal conduction through β-sheet crystals of natural silk protein.

    PubMed

    Zhang, Lin; Bai, Zhitong; Ban, Heng; Liu, Ling

    2015-11-21

    Recent experiments have discovered very different thermal conductivities between the spider silk and the silkworm silk. Decoding the molecular mechanisms underpinning the distinct thermal properties may guide the rational design of synthetic silk materials and other biomaterials for multifunctionality and tunable properties. However, such an understanding is lacking, mainly due to the complex structure and phonon physics associated with the silk materials. Here, using non-equilibrium molecular dynamics, we demonstrate that the amino acid sequence plays a key role in the thermal conduction process through β-sheets, essential building blocks of natural silks and a variety of other biomaterials. Three representative β-sheet types, i.e. poly-A, poly-(GA), and poly-G, are shown to have distinct structural features and phonon dynamics leading to different thermal conductivities. A fundamental understanding of the sequence effects may stimulate the design and engineering of polymers and biopolymers for desired thermal properties. PMID:26455593

  15. Ribonuclease "XlaI," an activity from Xenopus laevis oocytes that excises intervening sequences from yeast transfer ribonucleic acid precursors.

    PubMed Central

    Otsuka, A; de Paolis, A; Tocchini-Valentini, G P

    1981-01-01

    A ribonuclease (RNase) activity, RNase "XlaI," responsible for the excision of intervening sequences from two yeast transfer ribonucleic acid (tRNA) precursors, pre-tRNA(Tyr) and pre-tRNA(3Leu), has been purified 54-fold from nuclear extracts of Xenopus laevis oocytes. The RNase preparation is essentially free of contaminating RNase. A quantitative assay for RNase XlaI was developed, and the reaction products were characterized. RNase XlaI cleavage sites in the yeast tRNA precursors were identical to those made by yeast extracts (including 3'-phosphate and 5'-hydroxyl termini). Cleavage of pre-tRNA(3Leu) by RNase XlaI and subsequent ligation of the half-tRNA molecules do not require removal of the 5' leader or 3' trailer sequences. Images PMID:6765601

  16. Amino acid sequences of two novel long-chain neurotoxins from the venom of the sea snake Laticauda colubrina.

    PubMed

    Kim, H S; Tamiya, N

    1982-11-01

    From the venom of a population of the sea snake Laticauda colubrina from the Solomon Islands, a neurotoxic component, Laticauda colubrina a (toxin Lc a), was isolated in 16.6% (A280) yield. Similarly, from the venom of a population of L. colubrina from the Philippines, a neurotoxic component, Laticauda colubrina b (toxin Lc b), was obtained in 10.0% (A280) yield. The LD50 values of these toxins were 0.12 microgram/g body wt. on intramuscular injection in mice. Toxins Lc a and Lc b were each composed of molecules containing 69 amino acid residues with eight half-cystine residues. The complete amino acid sequences of these two toxins were elucidated. Toxins Lc a and Lc b are different from each other at five positions of their sequences, namely at positions 31 (Phe/Ser), 32 (Leu/Ile), 33 (Lys/Arg), 50 (Pro/Arg) and 53 (Asp/His) (residues in parentheses give the residues in toxins Lc a and Lc b respectively). Toxins Lc a and Lc b have a novel structure in that they have only four disulphide bridges, although the whole amino acid sequences are homologous to those of other known long-chain neurotoxins. It is remarkable that toxins Lc a and Lc b are not coexistent at the detection error of 6% of the other toxin. Populations of Laticauda colubrina from the Solomon Islands and from the Philippines have either toxin Lc a or toxin Lc b and not both of them. PMID:7159381

  17. CSTX-9, a toxic peptide from the spider Cupiennius salei: amino acid sequence, disulphide bridge pattern and comparison with other spider toxins containing the cystine knot structure.

    PubMed

    Schalle, J; Kämpfer, U; Schürch, S; Kuhn-Nentwig, L; Haeberli, S; Nentwig, W

    2001-09-01

    CSTX-9 (68 residues, 7530.9 Da) is one of the most abundant toxic polypeptides in the venom of the wandering spider Cupiennius salei. The amino acid sequence was determined by Edman degradation using reduced and alkylated CSTX-9 and peptides generated by cleavages with endoproteinase Asp-N and trypsin, respectively. Sequence comparison with CSTX-1, the most abundant and the most toxic polypeptide in the crude spider venom, revealed a high degree of similarity (53% identity). By means of limited proteolysis with immobilised trypsin and RP-HPLC, the cystine-containing peptides of CSTX-9 were isolated and the disulphide bridges were assigned by amino acid analysis, Edman degradation and nanospray tandem mass spectrometry. The four disulphide bonds present in CSTX-9 are arranged in the following pattern: 1-4, 2-5, 3-8 and 6-7 (Cys6-Cys21, Cys13-Cys30, Cys20-Cys48, Cys32-Cys46). Sequence comparison of CSTX-1 with CSTX-9 clearly indicates the same disulphide bridge pattern, which is also found in other spider polypeptide toxins, e.g. agatoxins (omega-AGA-IVA, omega-AGA-IVB, mu-AGA-I and mu-AGA-VI) from Agelenopsis aperta, SNX-325 from Segestria florentina and curtatoxins (CT-I, CT-II and CT-III) from Hololena curta. CSTX-1/CSTX-9 belong to the family of ion channel toxins containing the inhibitor cystine knot structural motif. CSTX-9, lacking the lysine-rich C-terminal tail of CSTX-1, exhibits a ninefold lower toxicity to Drosophila melanogaster than CSTX-1. This is in accordance with previous observations of CSTX-2a and CSTX-2b, two truncated forms of CSTX-1 which, like CSTX-9, also lack the C-terminal lysine-rich tail. PMID:11693532

  18. The sequence of rat leukosialin (W3/13 antigen) reveals a molecule with O-linked glycosylation of one third of its extracellular amino acids.

    PubMed Central

    Killeen, N; Barclay, A N; Willis, A C; Williams, A F

    1987-01-01

    Leukosialin is one of the major glycoproteins of thymocytes and T lymphocytes and is notable for a very high content of O-linked carbohydrate structures. The full protein sequence for rat leukosialin as translated from cDNA clones is now reported. The molecule contains 371 amino acids with 224 residues outside the cell, one transmembrane sequence and 124 cytoplasmic residues. Data from the peptide sequence and carbohydrate composition suggest that one in three of the extracellular amino acids may be O-glycosylated with no N-linked glycosylation sites. The cDNA sequence contained a CpG rich region in the 3' coding sequence and a large 3' non-coding region which included tandem repeats of the sequence GGAT. Images Fig. 4. PMID:2965006

  19. Amorphous/nanocrystalline silicon biosensor for the specific identification of unamplified nucleic acid sequences using gold nanoparticle probes

    NASA Astrophysics Data System (ADS)

    Martins, Rodrigo; Baptista, Pedro; Raniero, Leandro; Doria, Gonçalo; Silva, Leonardo; Franco, Ricardo; Fortunato, Elvira

    2007-01-01

    Amorphous/nanocrystalline silicon pi 'ii'n devices fabricated on micromachined glass substrates are integrated with oligonucleotide-derivatized gold nanoparticles for a colorimetric detection method. The method enables the specific detection and quantification of unamplified nucleic acid sequences (DNA and RNA) without the need to functionalize the glass surface, allowing for resolution of single nucleotide differences between DNA and RNA sequences—single nucleotide polymorphism and mutation detection. The detector's substrate is glass and the sample is directly applied on the back side of the biosensor, ensuring a direct optical coupling of the assays with a concomitant maximum photon capture and the possibility to reuse the sensor.

  20. An Interpretation of the Ancestral Codon from Miller’s Amino Acids and Nucleotide Correlations in Modern Coding Sequences

    PubMed Central

    Carels, Nicolas; de Leon, Miguel Ponce

    2015-01-01

    Purine bias, which is usually referred to as an “ancestral codon”, is known to result in short-range correlations between nucleotides in coding sequences, and it is common in all species. We demonstrate that RWY is a more appropriate pattern than the classical RNY, and purine bias (Rrr) is the product of a network of nucleotide compensations induced by functional constraints on the physicochemical properties of proteins. Through deductions from universal correlation properties, we also demonstrate that amino acids from Miller’s spark discharge experiment are compatible with functional primeval proteins at the dawn of living cell radiation on earth. These amino acids match the hydropathy and secondary structures of modern proteins. PMID:25922573

  1. Rapid Nucleic Acid Sequencing Methods--Alternative Approaches to Facilitating Learning.

    ERIC Educational Resources Information Center

    Bryce, Charles F. A.

    1982-01-01

    Because advanced students had difficulty in interpreting cleavage patterns obtained by gel electrophoresis related to rapid sequencing techniques for DNA and RNA, several formats were developed to aid in understanding this topic. Formats included print, print plus scrambled print, interactive computer-based instruction, and high-resolution…

  2. Draft Genome Sequence of Ustilago trichophora RK089, a Promising Malic Acid Producer

    PubMed Central

    Zambanini, Thiemo; Buescher, Joerg M.; Meurer, Guido; Blank, Lars M.

    2016-01-01

    The basidiomycetous smut fungus Ustilago trichophora RK089 produces malate from glycerol. De novo genome sequencing revealed a 20.7-Mbp genome (301 gap-closed contigs, 246 scaffolds). A comparison to the genome of Ustilago maydis 521 revealed all essential genes for malate production from glycerol contributing to metabolic engineering for improving malate production. PMID:27469969

  3. Draft Genome Sequence of Ustilago trichophora RK089, a Promising Malic Acid Producer.

    PubMed

    Zambanini, Thiemo; Buescher, Joerg M; Meurer, Guido; Wierckx, Nick; Blank, Lars M

    2016-01-01

    The basidiomycetous smut fungus Ustilago trichophora RK089 produces malate from glycerol. De novo genome sequencing revealed a 20.7-Mbp genome (301 gap-closed contigs, 246 scaffolds). A comparison to the genome of Ustilago maydis 521 revealed all essential genes for malate production from glycerol contributing to metabolic engineering for improving malate production. PMID:27469969

  4. 37 CFR 1.824 - Form and format for nucleotide and/or amino acid sequence submissions in computer readable form.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... nucleotide and/or amino acid sequence submissions in computer readable form. 1.824 Section 1.824 Patents... submissions in computer readable form. (a) The computer readable form required by § 1.821(e) shall meet the following requirements: (1) The computer readable form shall contain a single “Sequence Listing” as either...

  5. 37 CFR 1.824 - Form and format for nucleotide and/or amino acid sequence submissions in computer readable form.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... nucleotide and/or amino acid sequence submissions in computer readable form. 1.824 Section 1.824 Patents... submissions in computer readable form. (a) The computer readable form required by § 1.821(e) shall meet the following requirements: (1) The computer readable form shall contain a single “Sequence Listing” as either...

  6. 37 CFR 1.824 - Form and format for nucleotide and/or amino acid sequence submissions in computer readable form.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... nucleotide and/or amino acid sequence submissions in computer readable form. 1.824 Section 1.824 Patents... submissions in computer readable form. (a) The computer readable form required by § 1.821(e) shall meet the following requirements: (1) The computer readable form shall contain a single “Sequence Listing” as either...

  7. 37 CFR 1.824 - Form and format for nucleotide and/or amino acid sequence submissions in computer readable form.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... nucleotide and/or amino acid sequence submissions in computer readable form. 1.824 Section 1.824 Patents... submissions in computer readable form. (a) The computer readable form required by § 1.821(e) shall meet the following requirements: (1) The computer readable form shall contain a single “Sequence Listing” as either...

  8. 37 CFR 1.824 - Form and format for nucleotide and/or amino acid sequence submissions in computer readable form.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... nucleotide and/or amino acid sequence submissions in computer readable form. 1.824 Section 1.824 Patents... submissions in computer readable form. (a) The computer readable form required by § 1.821(e) shall meet the following requirements: (1) The computer readable form shall contain a single “Sequence Listing” as either...

  9. Mutation-selection models of coding sequence evolution with site-heterogeneous amino acid fitness profiles.

    PubMed

    Rodrigue, Nicolas; Philippe, Hervé; Lartillot, Nicolas

    2010-03-01

    Modeling the interplay between mutation and selection at the molecular level is key to evolutionary studies. To this end, codon-based evolutionary models have been proposed as pertinent means of studying long-range evolutionary patterns and are widely used. However, these approaches have not yet consolidated results from amino acid level phylogenetic studies showing that selection acting on proteins displays strong site-specific effects, which translate into heterogeneous amino acid propensities across the columns of alignments; related codon-level studies have instead focused on either modeling a single selective context for all codon columns, or a separate selective context for each codon column, with the former strategy deemed too simplistic and the latter deemed overparameterized. Here, we integrate recent developments in nonparametric statistical approaches to propose a probabilistic model that accounts for the heterogeneity of amino acid fitness profiles across the coding positions of a gene. We apply the model to a dozen real protein-coding gene alignments and find it to produce biologically plausible inferences, for instance, as pertaining to site-specific amino acid constraints, as well as distributions of scaled selection coefficients. In their account of mutational features as well as the heterogeneous regimes of selection at the amino acid level, the modeling approaches studied here can form a backdrop for several extensions, accounting for other selective features, for variable population size, or for subtleties of mutational features, all with parameterizations couched within population-genetic theory. PMID:20176949

  10. Terminal sequence studies of high-molecular-weight ribonucleic acid. The 3′-termini of rabbit globin messenger ribonucleic acid

    PubMed Central

    Hunt, John A.

    1973-01-01

    Haemoglobin mRNA isolated from EDTA-treated polyribosomes has an apparent molecular weight of 120000–180000 estimated by condensation with 3H-labelled isoniazid after periodate oxidation. Analysis of the ribonuclease digests of isoniazid-labelled RNA by paper electrophoresis and column chromatography enables the amount of contaminating 18S, 7S, 5S and 4S RNA to be estimated, and a corrected molecular weight of globin mRNA as the acid is 161000 or 500 nucleotides in length. This molecule contains two groups of 3′-terminal sequences in equal yield; G-Y-A6 and G-Y-A7 in the ratio 3:2, and G-N9–16-Y-A2 and G-N9–16-Y-N3 in the ratio 3:2. The significance of these sequences is discussed in relation to the poly(A) content of globin mRNA, the specificity of the sequences, and possible function in processing and biosynthesis of mRNA. PMID:4737318

  11. From Amino Acid to Glucosinolate Biosynthesis: Protein Sequence Changes in the Evolution of Methylthioalkylmalate Synthase in Arabidopsis[W][OA

    PubMed Central

    de Kraker, Jan-Willem; Gershenzon, Jonathan

    2011-01-01

    Methylthioalkylmalate synthase (MAM) catalyzes the committed step in the side chain elongation of Met, yielding important precursors for glucosinolate biosynthesis in Arabidopsis thaliana and other Brassicaceae species. MAM is believed to have evolved from isopropylmalate synthase (IPMS), an enzyme involved in Leu biosynthesis, based on phylogenetic analyses and an overlap of catalytic abilities. Here, we investigated the changes in protein structure that have occurred during the recruitment of IPMS from amino acid to glucosinolate metabolism. The major sequence difference between IPMS and MAM is the absence of 120 amino acids at the C-terminal end of MAM that constitute a regulatory domain for Leu-mediated feedback inhibition. Truncation of this domain in Arabidopsis IPMS2 results in loss of Leu feedback inhibition and quaternary structure, two features common to MAM enzymes, plus an 8.4-fold increase in the kcat/Km for a MAM substrate. Additional exchange of two amino acids in the active site resulted in a MAM-like enzyme that had little residual IPMS activity. Hence, combination of the loss of the regulatory domain and a few additional amino acid exchanges can explain the evolution of MAM from IPMS during its recruitment from primary to secondary metabolism. PMID:21205930

  12. Templated synthesis of peptide nucleic acids via sequence-selective base-filling reactions.

    PubMed

    Heemstra, Jennifer M; Liu, David R

    2009-08-19

    The templated synthesis of nucleic acids has previously been achieved through the backbone ligation of preformed nucleotide monomers or oligomers. In contrast, here we demonstrate templated nucleic acid synthesis using a base-filling approach in which individual bases are added to abasic sites of a peptide nucleic acid (PNA). Because nucleobase substrates in this approach are not self-reactive, a base-filling approach may reduce the formation of nontemplated reaction products. Using either reductive amination or amine acylation chemistries, we observed efficient and selective addition of each of the four nucleobases to an abasic site in the middle of the PNA strand. We also describe the addition of single nucleobases to the end of a PNA strand through base filling, as well as the tandem addition of two bases to the middle of the PNA strand. These findings represent an experimental foundation for nonenzymatic information transfer through base filling. PMID:19722647

  13. Templated Synthesis of Peptide Nucleic Acids via Sequence-Selective Base-Filling Reactions

    PubMed Central

    2009-01-01

    The templated synthesis of nucleic acids has previously been achieved through the backbone ligation of preformed nucleotide monomers or oligomers. In contrast, here we demonstrate templated nucleic acid synthesis using a base-filling approach in which individual bases are added to abasic sites of a peptide nucleic acid (PNA). Because nucleobase substrates in this approach are not self-reactive, a base-filling approach may reduce the formation of nontemplated reaction products. Using either reductive amination or amine acylation chemistries, we observed efficient and selective addition of each of the four nucleobases to an abasic site in the middle of the PNA strand. We also describe the addition of single nucleobases to the end of a PNA strand through base filling, as well as the tandem addition of two bases to the middle of the PNA strand. These findings represent an experimental foundation for nonenzymatic information transfer through base filling. PMID:19722647

  14. Complete Genome Sequence of a thermotolerant sporogenic lactic acid bacterium, Bacillus coagulans strain 36D1

    SciTech Connect

    Xie, Gary; Dalin, Eileen; Tice, Hope; Chertkov, Olga; Land, Miriam L

    2011-01-01

    Bacillus coagulans is a ubiquitous soil bacterium that grows at 50-55 C and pH 5.0 and fer-ments various sugars that constitute plant biomass to L (+)-lactic acid. The ability of this sporogenic lactic acid bacterium to grow at 50-55 C and pH 5.0 makes this organism an attractive microbial biocatalyst for production of optically pure lactic acid at industrial scale not only from glucose derived from cellulose but also from xylose, a major constituent of hemi-cellulose. This bacterium is also considered as a potential probiotic. Complete genome squence of a representative strain, B. coagulans strain 36D1, is presented and discussed.

  15. Complete Genome Sequence of a thermotolerant sporogenic lactic acid bacterium, Bacillus coagulans strain 36D1

    SciTech Connect

    Rhee, Mun Su; Moritz, Brelan E.; Xie, Gary; Glavina Del Rio, Tijana; Dalin, Eileen; Tice, Hope; Bruce, David; Goodwin, Lynne A.; Chertkov, Olga; Brettin, Thomas S; Han, Cliff; Detter, J. Chris; Pitluck, Sam; Land, Miriam L; Patel, Milind; Ou, Mark; Harbrucker, Roberta; Ingram, Lonnie O.; Shanmugam, Keelnathan T.

    2011-01-01

    Bacillus coagulans is a ubiquitous soil bacterium that grows at 50-55 C and pH 5.0 and fer- ments various sugars that constitute plant biomass to L (+)-lactic acid. The ability of this spo- rogenic lactic acid bacterium to grow at 50-55 C and pH 5.0 makes this organism an attrac- tive microbial biocatalyst for production of optically pure lactic acid at industrial scale not only from glucose derived from cellulose but also from xylose, a major constituent of hemi- cellulose. This bacterium is also considered as a potential probiotic. Complete genome se- quence of a representative strain, B. coagulans strain 36D1, is presented and discussed.

  16. Molecular characterization of the body site-specific human epidermal cytokeratin 9: cDNA cloning, amino acid sequence, and tissue specificity of gene expression.

    PubMed

    Langbein, L; Heid, H W; Moll, I; Franke, W W

    1993-12-01

    Differentiation of human plantar and palmar epidermis is characterized by the suprabasal synthesis of a major special intermediate-sized filament (IF) protein, the type I (acidic) cytokeratin 9 (CK 9). Using partial amino acid (aa) sequence information obtained by direct Edman sequencing of peptides resulting from proteolytic digestion of purified CK 9, we synthesized several redundant primers by 'back-translation'. Amplification by polymerase chain reaction (PCR) of cDNAs obtained by reverse transcription of mRNAs from human foot sole epidermis, including 5'-primer extension, resulted in multiple overlapping cDNA clones, from which the complete cDNA (2353 bp) could be constructed. This cDNA encoded the CK 9 polypeptide with a calculated molecular weight of 61,987 and an isoelectric point at about pH 5.0. The aa sequence deduced from cDNA was verified in several parts by comparison with the peptide sequences and showed the typical structure of type I CKs, with a head (153 aa), and alpha-helical coiled-coil-forming rod (306 aa), and a tail (163 aa) domain. The protein displayed the highest homology to human CK 10, not only in the highly conserved rod domain but also in large parts of the head and the tail domains. On the other hand, the aa sequence revealed some remarkable differences from CK 10 and other CKs, even in the most conserved segments of the rod domain. The nuclease digestion pattern seen on Southern blot analysis of human genomic DNA indicated the existence of a unique CK 9 gene. Using CK 9-specific riboprobes for hybridization on Northern blots of RNAs from various epithelia, a mRNA of about 2.4 kb in length could be identified only in foot sole epidermis, and a weaker cross-hybridization signal was seen in RNA from bovine heel pad epidermis at about 2.0 kb. A large number of tissues and cell cultures were examined by PCR of mRNA-derived cDNAs, using CK 9-specific primers. But even with this very sensitive signal amplification, only palmar

  17. Diverse Bacterial PKS Sequences Derived From Okadaic Acid-Producing Dinoflagellates

    PubMed Central

    Perez, Roberto; Liu, Li; Lopez, Jose; An, Tianying; Rein, Kathleen S.

    2008-01-01

    Okadaic acid (OA) and the related dinophysistoxins are isolated from dinoflagellates of the genus Prorocentrum and Dinophysis. Bacteria of the Roseobacter group have been associated with okadaic acid producing dinoflagellates and have been previously implicated in OA production. Analysis of 16S rRNA libraries reveals that Roseobacter are the most abundant bacteria associated with OA producing dinoflagellates of the genus Prorocentrum and are not found in association with non-toxic dinoflagellates. While some polyketide synthase (PKS) genes form a highly supported Prorocentrum clade, most appear to be bacterial, but unrelated to Roseobacter or Alpha-Proteobacterial PKSs or those derived from other Alveolates Karenia brevis or Crytosporidium parvum. PMID:18728765

  18. Complete Genome Sequence of Moraxella osloensis Strain KMC41, a Producer of 4-Methyl-3-Hexenoic Acid, a Major Malodor Compound in Laundry.

    PubMed

    Goto, Takatsugu; Hirakawa, Hideki; Morita, Yuji; Tomida, Junko; Sato, Jun; Matsumura, Yuta; Mitani, Asako; Niwano, Yu; Takeuchi, Kohei; Kubota, Hiromi; Kawamura, Yoshiaki

    2016-01-01

    We report the complete genome sequence of Moraxella osloensis strain KMC41, isolated from laundry with malodor. The KMC41 genome comprises a 2,445,556-bp chromosome and three plasmids. A fatty acid desaturase and at least four β-oxidation-related genes putatively associated with 4-methyl-3-hexenoic acid generation were detected in the KMC41 chromosome. PMID:27445387

  19. Complete Genome Sequence of Moraxella osloensis Strain KMC41, a Producer of 4-Methyl-3-Hexenoic Acid, a Major Malodor Compound in Laundry

    PubMed Central

    Hirakawa, Hideki; Morita, Yuji; Tomida, Junko; Sato, Jun; Matsumura, Yuta; Mitani, Asako; Niwano, Yu; Takeuchi, Kohei; Kubota, Hiromi; Kawamura, Yoshiaki

    2016-01-01

    We report the complete genome sequence of Moraxella osloensis strain KMC41, isolated from laundry with malodor. The KMC41 genome comprises a 2,445,556-bp chromosome and three plasmids. A fatty acid desaturase and at least four β-oxidation-related genes putatively associated with 4-methyl-3-hexenoic acid generation were detected in the KMC41 chromosome. PMID:27445387

  20. cDNA cloning and structural characterization of a lectin from the mussel Crenomytilus grayanus with a unique amino acid sequence and antibacterial activity.

    PubMed

    Kovalchuk, Svetlana N; Chikalovets, Irina V; Chernikov, Oleg V; Molchanova, Valentina I; Li, Wei; Rasskazov, Valery A; Lukyanov, Pavel A

    2013-10-01

    An amino acid sequence of GalNAc/Gal-specific lectin from the mussel Crenomytilus grayanus (CGL) was determined by cDNA sequencing. CGL consists of 150 amino acid residues, contains three tandem repeats with high sequence similarities to each other (up to 73%) and does not belong to any known lectins family. According to circular dichroism results CGL is a β/α-protein with the predominance of β-structure. CGL was predicted to adopt a ß-trefoil fold. The lectin exhibits antibacterial activity and might be involved in the recognition and clearance of bacterial pathogens in the shellfish. PMID:23886951

  1. Snake venom toxins. The amino acid sequence of toxin Vi2, a homologue of pancreatic trypsin inhibitor, from Dendroaspis polylepis polylepis (black mamba) venom.

    PubMed

    Strydom, D J

    1977-04-25

    The amino acid sequence of venom component Vi2, a protein of low toxicity from Dendroaspis polylepis polylepis venom was determined by automatic sequence analysis in combination with sequence studies on tryptic peptides. This protein, the most retarded fraction of this venom on a cation-exchange resin, is a homologue of bovine pancreatic trypsin inhibitor consisting of a single chain of 57 amino acid residues containing six half-cystine residues. The active site lysyl residue of bovine trypsin inhibitor is conserved in Vi2 although large differences are found in the rest of the molecule. PMID:857902

  2. Isolation and complete amino acid sequence of two fibrinolytic proteinases from the toxic Saturnid caterpillar Lonomia achelous.

    PubMed

    Amarant, T; Burkhart, W; LeVine, H; Arocha-Pinango, C L; Parikh, I

    1991-08-30

    The major toxic and fibrinolytic activity of the saliva and hemolymph of the larval form of Lonomia achelous was purified to homogeneity by a combination of metal chelate and affinity chromatography. Two apparent isozymes, Achelase I (213 amino acids, pIcalc = 10.55) and Achelase II (214 amino acids, pIcalc = 8.51), were sequenced by automated Edman degradation, and their C-termini confirmed by Fourier-transform mass spectrometry. The calculated molecular weights (22,473 and 22,727) correspond well to Mr estimates of 24,000 by SDS-PAGE. No carbohydrate was detected during sequencing. The enzymes degraded all three chains of fibrin, alpha greater than beta much greater than gamma, yielding a fragmentation pattern indistinguishable from that produced by trypsin. Chromogenic peptides S-2222 (Factor Xa and trypsin), S-2251 (plasmin), S-2302 (kallikrein) and S-2444 (urokinase) were substrates while S-2288 (broad range of serine proteinases including thrombin) was not hydrolyzed. Among a range of inhibitors Hg+2, aminophenylmercuriacetate, leupeptin, antipain and E-64 but not N-ethylmaleimide or iodoacetate abolished the activity of the purified isozymes against S-2444. Phenylmethylsulfonyl fluoride, soybean trypsin inhibitor and aprotinin were less effective. The presence of the classic catalytic triad (histidine-41, aspartate-86 and serine-189) suggests that Achelases I and II may be serine proteinases, but with a potentially free cysteine-185 which could react with thiol proteinase-directed reagents. PMID:1911844

  3. Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2232 from Karijini National Park, Australia

    PubMed Central

    Walker, Robert; Watkin, Elizabeth; Tian, Rui; Bräu, Lambert; O’Hara, Graham; Goodwin, Lynne; Han, James; Reddy, Tatiparthi; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavromatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

    2013-01-01

    Burkholderia sp. strain WSM2232 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod that was trapped in 2001 from acidic soil collected from Karijini National Park (Australia) using Gastrolobium capitatum as a host. WSM2232 was effective in nitrogen fixation with G. capitatum but subsequently lost symbiotic competence during long-term storage. Here we describe the features of Burkholderia sp. strain WSM2232, together with genome sequence information and its annotation. The 7,208,311 bp standard-draft genome is arranged into 72 scaffolds of 72 contigs containing 6,322 protein-coding genes and 61 RNA-only encoding genes. The loss of symbiotic capability can now be attributed to the loss of nodulation and nitrogen fixation genes from the genome. This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project. PMID:25197442

  4. [Influence of dietotherapy enriched with conjugated linoleic acid on anthropometrical indicators and body composite structure in patients with an overweight].

    PubMed

    Bogdanov, A R; Derbeneva, S A

    2013-01-01

    The clinical efficiency of dietotherapy enriched with conjugated linoleic acid (3 g per day) in patients, suffering from the excess mass of a body and obesity of I and II degrees has been investigated. Dynamics of anthropometrical indicators and body composite structure before and after the treatment of patients from the main (n = 20) and control groups were estimated. Studied parameters were fixed in day 0, 14 and 28. Statistically significant reduction of the weight, body mass index (BMI), anthropometrical parameters (volumes of a waist, hips, a shoulder), indicators of body composite structure (fatty weight, area of splanchnic fat, muscular weight) has been revealed as a result of the carried-out course of treatment, and they were more expressed in patients of the main group. In particular, the waist size authentically decreased by 6.5% in patients from the main group, while in patients from the group of comparison the decrease reached only 4.2%. Average change of waist volume was--7.0 cm in the main group and--4.5 cm in the control group. The grasp of hips in the main group decreased by 4.2%, in the group of comparison for 2.8%. Absolute change of hips volume in the main group 1.67 fold exceeded that in the group of comparison (p < 0.005). The shin grasp in the main group decreased by 9.7%, while in the control group remained invariable. BMI decreased by 7.2% and 5%, respectively. Thus, the reduction of body fatty weight was 10% in patients from the main group and 4% in patients from the group of comparison. The body total liquid decreased by 1.4 l in the main group, and only by 0.3 l in the group of comparison. The received intergroup distinctions are statistically reliable (p < 0.05). PMID:24340933

  5. Cloning and nucleotide sequencing of a novel 7 beta-(4-carboxybutanamido)cephalosporanic acid acylase gene of Bacillus laterosporus and its expression in Escherichia coli and Bacillus subtilis.

    PubMed

    Aramori, I; Fukagawa, M; Tsumura, M; Iwami, M; Ono, H; Kojo, H; Kohsaka, M; Ueda, Y; Imanaka, H

    1991-12-01

    A strain of Bacillus species which produced an enzyme named glutaryl 7-ACA acylase which converts 7 beta-(4-carboxybutanamido)cephalosporanic acid (glutaryl 7-ACA) to 7-amino cephalosporanic acid (7-ACA) was isolated from soil. The gene for the glutaryl 7-ACA acylase was cloned with pHSG298 in Escherichia coli JM109, and the nucleotide sequence was determined by the M13 dideoxy chain termination method. The DNA sequence revealed only one large open reading frame composed of 1,902 bp corresponding to 634 amino acid residues. The deduced amino acid sequence contained a potential signal sequence in its amino-terminal region. Expression of the gene for glutaryl 7-ACA acylase was performed in both E. coli and Bacillus subtilis. The enzyme preparations purified from either recombinant strain of E. coli or B. subtilis were shown to be identical with each other as regards the profile of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and were composed of a single peptide with the molecular size of 70 kDa. Determination of the amino-terminal sequence of the two enzyme preparations revealed that both amino-terminal sequences (the first nine amino acids) were identical and completely coincided with residues 28 to 36 of the open reading frame. Extracellular excretion of the enzyme was observed in a recombinant strain of B. subtilis. PMID:1744041

  6. Nucleic acid-binding molecules with high affinity and base sequence specificity: intercalating agents covalently linked to oligodeoxynucleotides.

    PubMed Central

    Asseline, U; Delarue, M; Lancelot, G; Toulmé, F; Thuong, N T; Montenay-Garestier, T; Hélène, C

    1984-01-01

    Oligodeoxyribonucleotides covalently linked to an intercalating agent via a polymethylene linker were synthesized. Oligothymidylates attached to an acridine dye (Acr) through the 3'-phosphate group [(Tp)n(CH2) mAcr ] specifically interact with the complementary sequence. The interaction is strongly stabilized by the intercalating agent. By using absorption and fluorescence spectroscopies, it is shown that complex formation between (Tp)n(CH2) mAcr and poly(rA) involves the formation of n A X T base pairs, where n is the number of thymines in the oligonucleotide. The acridine ring intercalates between A X T base pairs. Fluorescence excitation spectra reveal the existence of two environments for the acridine ring, whose relative contributions depend on the linker length (m). The binding of (Tp)4(CH2) mAcr to poly(rA) is analyzed in terms of site binding and cooperative interactions between oligonucleotides along the polynucleotide lattice. Thermodynamic parameters show that the covalent attachment of the acridine ring strongly stabilizes the binding of the oligonucleotide to its complementary sequence. The stabilization depends on the linker length; the compound with m = 5 gives a more stable complex than that with m = 3. These results open the way to the synthesis of a family of molecules exhibiting both high-affinity and high-specificity for a nucleic acid base sequence. PMID:6587350

  7. Paleoclimate and weathering of the Tokaj (NE Hungary) loess-paleosol sequence: a comparison of geochemical weathering indices and paleoclimate parameters

    NASA Astrophysics Data System (ADS)

    Schatz, A.-K.; Scholten, T.; Kühn, P.

    2014-02-01

    The Tokaj loess-paleosol sequence in NE Hungary is one of the key sites for detailed paleoclimate reconstructions of the Quaternary in SE Europe. In this study, the geochemical composition of samples from the upper part of the sequence (45-21 ka) was analyzed and a variety of commonly used weathering indices and element ratios were applied to estimate weathering intensity. Further, similarities and differences between these weathering indices and their sensitivity to changes in paleoclimatic conditions were assessed. Results indicate that all of them accurately track changes in weathering intensity and are, with minor exceptions, very similar to each other. Based on different transfer functions for major and trace element concentrations (XRF), magnetic susceptibility (MS) and δ13C data, we calculated mean annual paleotemperature and mean annual paleoprecipitation for the time intervals of paleosol formation (45-27 ka) and dust deposition (27-21 ka). Results differ depending on the respective transfer function and method but largely agree with previously published paleoclimate data of the region. XRF- and δ13C-based results converge to a MAT of 8-10°C (paleosol) and 8-9°C (loess) and show a~MAP range of 685-879 mm a-1 (paleosol) and 572-700 mm a-1 (loess). MS-based results are most reliable with MATs of 8.4°C (paleosol) and 6.7°C (loess) and MAPs of 325-441 mm a-1 (paleosol) and 224 mm a-1 (loess).

  8. Advanced stability indicating chemometric methods for quantitation of amlodipine and atorvastatin in their quinary mixture with acidic degradation products.

    PubMed

    Darwish, Hany W; Hassan, Said A; Salem, Maissa Y; El-Zeany, Badr A

    2016-02-01

    Two advanced, accurate and precise chemometric methods are developed for the simultaneous determination of amlodipine besylate (AML) and atorvastatin calcium (ATV) in the presence of their acidic degradation products in tablet dosage forms. The first method was Partial Least Squares (PLS-1) and the second was Artificial Neural Networks (ANN). PLS was compared to ANN models with and without variable selection procedure (genetic algorithm (GA)). For proper analysis, a 5-factor 5-level experimental design was established resulting in 25 mixtures containing different ratios of the interfering species. Fifteen mixtures were used as calibration set and the other ten mixtures were used as validation set to validate the prediction ability of the suggested models. The proposed methods were successfully applied to the analysis of pharmaceutical tablets containing AML and ATV. The methods indicated the ability of the mentioned models to solve the highly overlapped spectra of the quinary mixture, yet using inexpensive and easy to handle instruments like the UV-VIS spectrophotometer. PMID:26513228

  9. Amino acid sequence and posttranslational modifications of human factor VII sub a from plasma and transfected baby hamster kidney cells

    SciTech Connect

    Thim, L.; Bjoern, S.; Christensen, M.; Nicolaisen, E.M.; Lund-Hansen, T.; Pedersen, A.H.; Hedner, U. )

    1988-10-04

    Blood coagulation factor VII is a vitamin K dependent glycoprotein which in its activated form, factor VII{sub a}, participates in the coagulation process by activating factor X and/or factor IX in the presence of Ca{sup 2+} and tissue factor. Three types of potential posttranslational modifications exist in the human factor VII{sub a} molecule, namely, 10 {gamma}-carboxylated, N-terminally located glutamic acid residues, 1 {beta}-hydroxylated aspartic acid residue, and 2 N-glycosylated asparagine residues. In the present study, the amino acid sequence and posttranslational modifications of recombinant factor VII{sub a} as purified from the culture medium of a transfected baby hamster kidney cell line have been compared to human plasma factor VII{sub a}. By use of HPLC, amino acid analysis, peptide mapping, and automated Edman degradation, the protein backbone of recombinant factor VII{sub a} was found to be identical with human factor VII{sub a}. Asparagine residues 145 and 322 were found to be fully N-glycosylated in human plasma factor VII{sub a}. In the recombinant factor VII{sub a}, asparagine residue 322 was fully glycosylated whereas asparagine residue 145 was only partially (approximately 66%) glycosylated. Besides minor differences in the sialic acid and fucose contents, the overall carbohydrate compositions were nearly identical in recombinant factor VII{sub a} and human plasma factor VII{sub a}. These results show that factor VII{sub a} as produced in the transfected baby hamster kidney cells is very similar to human plasma factor VII{sub a} and that this cell line thus might represent an alternative source for human factor VII{sub a}.

  10. Complete genome sequence of probiotic Bacillus coagulans HM-08: A potential lactic acid producer.

    PubMed

    Yao, Guoqiang; Gao, Pengfei; Zhang, Wenyi

    2016-06-20

    Bacillus coagulans HM-08 is a commercialized probiotic strain in China. Its genome contains a 3.62Mb circular chromosome with an average GC content of 46.3%. In silico analysis revealed the presence of one xyl operon as well as several other genes that are correlated to xylose utilization. The genetic information provided here may help to expand its future biotechnology potential in lactic acid production. PMID:27130497

  11. Complete Genome Sequence of the Amino Acid-Fermenting Clostridium propionicum X2 (DSM 1682)

    PubMed Central

    Poehlein, Anja; Schlien, Katja; Chowdhury, Nilanjan Pal; Gottschalk, Gerhard; Buckel, Wolfgang

    2016-01-01

    Clostridium propionicum is a strict anaerobic, Gram positive, rod-shaped bacterium that belongs to the clostridial cluster XIVb. The genome consists of one replicon (3.1 Mb) and harbors 2,936 predicted protein-encoding genes. The genome encodes all enzymes required for fermentation of the amino acids α-alanine, β-alanine, serine, threonine, and methionine. PMID:27081148

  12. Purification, characterization, and complete amino acid sequence of a trypsin inhibitor from amaranth (Amaranthus hypochondriacus) seeds.

    PubMed Central

    Valdes-Rodriguez, S; Segura-Nieto, M; Chagolla-Lopez, A; Verver y Vargas-Cortina, A; Martinez-Gallardo, N; Blanco-Labra, A

    1993-01-01

    A protein proteinase inhibitor was purified from a seed extract of amaranth (Amaranthus hypochondriacus) by precipitation with (NH4)2SO4, gel-filtration chromatography, ion-exchange chromatography, and reverse-phase high-performance liquid chromatography. It is a 69-amino acid protein with a high content of valine, arginine, and glutamic acid, but lacking in methionine. The inhibitor has a relative molecular weight of 7400 and an isoelectric point of 7.5. It is a serine proteinase inhibitor that recognizes chymotrypsin, trypsin, and trypsin-like proteinase activities extracted from larvae of the insect Prostephanus truncatus. This inhibitor belongs to the potato-I inhibitor family, showing the closest homology (59.5%) with the Lycopersicum peruvianum trypsin inhibitor, and (51%) with the proteinase inhibitor 5 extracted from the seeds of Cucurbita maxima. The position of the lysine-aspartic acid residues present in the active site of the amaranth inhibitor are found in almost the same relative position as in the inhibitor from C. maxima. PMID:8290633

  13. Naphthenic acids in coastal sediments after the Hebei Spirit oil spill: a potential indicator for oil contamination.

    PubMed

    Wan, Yi; Wang, Beili; Khim, Jong Seong; Hong, Seongjin; Shim, Won Joon; Hu, Jianying

    2014-04-01

    Naphthenic acids (NAs) as toxic components in most petroleum sources are suspected to be one of the major pollutants in the aquatic environment following oil spills, and the polarity and persistence of NAs make it a potential indicator for oil contamination. However, the contamination and potential effects of pollutants in oil spill affected areas remain unknown. To investigate NAs in oil spill affected areas, a sensitive method was first established for analysis of NAs, together with oxy-NAs in sediment samples by UPLC-QTOF-MS. Then the method was applied to determine the NA mixtures in crude oil, weathered oil, and sediments from the spilled sites after the Hebei Spirit oil spill, Taean, South Korea (Dec. 2007). Concentrations of NAs, O3-NAs, and O4-NAs were found to be 7.8-130, 3.6-44, and 0.8-20 mg kg(-1) dw in sediments from the Taean area, respectively, which were much greater than those measured in the reference sites of Manlipo and Anmyundo beaches. Concentrations of NAs were 50-100 times greater than those (0.077-2.5 mg kg(-1) dw) of PAHs in the same sediment samples, thus the ecological risk of NAs in oil spill affected areas deserves more attention. The sedimentary profiles of oil-derived NAs and background NAs centered around compounds with 21-35 and 12-21 carbons, respectively, indicating that the crude-derived NA mixtures originating from the 2007 oil spill were persistent. Acyclic NAsn=5-20 were easily degraded compared to cyclic NAsn=21-41 during the oil weathering processes, and the ratio of oxy-NAsn=21-41 relative to NAsn=21-41 could be a novel index to estimate the degree of oil weathering in sediments. Altogether, the persistent oil-derived NAsn=21-41 could be used as a potential indicator for oil-specific contamination, as such compounds would not be much affected by the properties of coastal sediments possibly due to the high sorption of the negatively charged compounds (NAs) in sediment. PMID:24579908

  14. Protein Requirements Are Elevated in Endurance Athletes after Exercise as Determined by the Indicator Amino Acid Oxidation Method

    PubMed Central

    Kato, Hiroyuki; Suzuki, Katsuya; Bannai, Makoto; Moore, Daniel R.

    2016-01-01

    A higher protein intake has been recommended for endurance athletes compared with healthy non-exercising individuals based primarily on nitrogen balance methodology. The aim of this study was to determine the estimated average protein requirement and recommended protein intake in endurance athletes during an acute 3-d controlled training period using the indicator amino acid oxidation method. After 2-d of controlled diet (1.4 g protein/kg/d) and training (10 and 5km/d, respectively), six male endurance-trained adults (28±4 y of age; Body weight, 64.5±10.0 kg; VO2peak, 60.3±6.7 ml·kg-1·min-1; means±SD) performed an acute bout of endurance exercise (20 km treadmill run) prior to consuming test diets providing variable amounts of protein (0.2–2.8 g·kg-1·d-1) and sufficient energy. Protein was provided as a crystalline amino acid mixture based on the composition of egg protein with [1-13C]phenylalanine provided to determine whole body phenylalanine flux, 13CO2 excretion, and phenylalanine oxidation. The estimated average protein requirement was determined as the breakpoint after biphasic linear regression analysis with a recommended protein intake defined as the upper 95% confidence interval. Phenylalanine flux (68.8±8.5 μmol·kg-1·h-1) was not affected by protein intake. 13CO2 excretion displayed a robust bi-phase linear relationship (R2 = 0.86) that resulted in an estimated average requirement and a recommended protein intake of 1.65 and 1.83 g protein·kg-1·d-1, respectively, which was similar to values based on phenylalanine oxidation (1.53 and 1.70 g·kg-1·d-1, respectively). We report a recommended protein intake that is greater than the RDA (0.8 g·kg-1·d-1) and current recommendations for endurance athletes (1.2–1.4 g·kg-1·d-1). Our results suggest that the metabolic demand for protein in endurance-trained adults on a higher volume training day is greater than their sedentary peers and current recommendations for athletes based primarily on

  15. Assessment of protein requirement in octogenarian women with use of the indicator amino acid oxidation technique123

    PubMed Central

    Tang, Minghua; McCabe, George P; Elango, Rajavel; Pencharz, Paul B; Ball, Ronald O; Campbell, Wayne W

    2014-01-01

    Background: Data on the protein requirements of elderly adults are limited, because it is impractical to conduct repeated nitrogen balance protocols in these vulnerable humans. Objective: This study was designed to determine the dietary protein requirement of elderly women by using the recently developed minimally invasive indicator amino acid oxidation (IAAO) technique. Design: Six white women aged 80–87 y [mean ± SEM: 82 ± 1 y and body mass index (in kg/m2) 26 ± 2] completed a 3-d protocol 7 times. Each woman consumed an adaptation diet for 2 d and on day 3 consumed a complete test diet with a crystalline amino acid mixture containing 1 of 7 protein intakes (0.1, 0.3, 0.6, 0.9, 1.2, 1.5, or 1.8 g · kg−1 · d−1) tested randomly. A group-based protein requirement was assessed by using a nonlinear mixed model of protein intake and l-[1-13C]phenylalanine oxidation. The breakpoint, at which there was no further decline in the rate of appearance of 13C in the breath, was used as an index of the mean protein requirement. Results: The mean protein requirement (95% CI) was 0.85 (0.60, 1.09) g · kg−1 · d−1. This requirement is 29% higher than the current Estimated Average Requirement (EAR) for adults of 0.66 g · kg−1 · d−1 based on the nitrogen balance technique, although the 95% CI includes the current EAR. The corresponding adequate protein allowance of 1.15 (0.77, 1.54) g · kg−1 · d−1 is 44% higher, although the 95% CI includes the Recommended Dietary Allowance (RDA) of 0.80 g · kg−1 · d−1. Conclusions: Notwithstanding uncertainty about the validity of the use of the IAAO technique to assess protein requirements, the results of this study with octogenarian women suggest that the current EAR and RDA for elderly women may be underestimated. The limitations of this short-term, noninvasive method underscore the need for new research that uses alternative experimental designs and measuring physiologic, morphologic, and health

  16. Inferences from protein and nucleic acid sequences - Early molecular evolution, divergence of kingdoms and rates of change

    NASA Technical Reports Server (NTRS)

    Dayhoff, M. O.; Barker, W. C.; Mclaughlin, P. J.

    1974-01-01

    Description of new sensitive, objective methods for establishing the probable common ancestry of very distantly related sequences and the quantitative evolutionary change which has taken place. These methods are applied to four families of proteins and nucleic acids and evolutionary trees will be derived where possible. Of the three families containing duplications of genetic material, two are nucleic acids: transfer RNA and 5S ribosomal RNA. Both of these structures are functional in the synthesis of coded proteins, and prototypes must have been present in the cell at the inception of the fundamental coding process that all living things share. There are many types of tRNA which recognize the various nucleotide triplets and the 20 amino acids. These types are thought to have arisen as a result of many gene duplications. Relationships among these types are discussed. The 5S ribosomal RNA, presently functional in both eukaryotes and prokaryotes, is very likely descended from an early form incorporating almost a complete duplication of genetic material. The amount of evolution in the various lines can again be compared. The other two families containing duplications are proteins; ferredoxin and cytochrome c.

  17. Species specific amino acid sequence-protein local structure relationships: An analysis in the light of a structural alphabet.

    PubMed

    de Brevern, Alexandre G; Joseph, Agnel Praveen

    2011-05-01

    Protein structure analysis and prediction methods are based on non-redundant data extracted from the available protein structures, regardless of the species from which the protein originates. Hence, these datasets represent the global knowledge on protein folds, which constitutes a generic distribution of amino acid sequence-protein structure (AAS-PS) relationships. In this study, we try to elucidate whether the AAS-PS relationship could possess specificities depending on the specie. For this purpose, we have chosen three different species: Saccharomyces cerevisiae, Plasmodium falciparum and Arabidopsis thaliana. We analyzed the AAS-PS behaviors of the proteins from these three species and compared it to the "expected" distribution of a classical non-redundant databank. With the classical secondary structure description, only slight differences in amino acid preferences could be observed. With a more precise description of local protein structures (Protein Blocks), significant changes could be highlighted. S. cerevisiae's AAS-PS relationship is close to the general distribution, while striking differences are observed in the case of A. thaliana. P. falciparum is the most distant one. This study presents some interesting view-points on AAS-PS relationship. Certain species exhibit unique preferences for amino acids to be associated with protein local structural elements. Thus, AAS-PS relationships are species dependent. These results can give useful insights for improving prediction methodologies which take the species specific information into account. PMID:21333657

  18. Amino acid sequence alignment of bacterial and mammalian pancreatic serine proteases based on topological equivalences.

    PubMed

    James, M N; Delbaere, L T; Brayer, G D

    1978-06-01

    The three-dimensional structures of the bacterial serine proteases SGPA, SGPB, and alpha-lytic protease have been compared with those of the pancreatic enzymes alpha-chymotrypsin and elastase. This comparison shows that approximately 60% (55-64%) of the alpha-carbon atom positions of the bacterial serine proteases are topologically equivalent to the alpha-carbon atom positions of the pancreatic enzymes. The corresponding value for a comparison of the bacterial enzymes among themselves is approximately 84%. The results of these topological comparisons have been used to deduce an experimentally sound sequence alignment for these several enzymes. This alignment shows that there is extensive tertiary structural homology among the bacteria and pancreatic enzymes without significant primary sequence identity (less than 21%). The acquisition of a zymogen function by the pancreatic enzymes is accompanied by two major changes to the bacterial enzymes' architecture: an insertion of 9 residues to increase the length of the N-terminal loop, and one of 12 residues to a loop near the activation salt bridge. In addition, in these two enzyme families, the methionine loop (residues 164-182) adopts very different comformations which are associated with their altered substrate specificities. PMID:96920

  19. Genotyping-by-sequencing approach indicates geographic distance as the main factor affecting genetic structure and gene flow in Brazilian populations of Grapholita molesta (Lepidoptera, Tortricidae).

    PubMed

    Silva-Brandão, Karina Lucas; Silva, Oscar Arnaldo Batista Neto E; Brandão, Marcelo Mendes; Omoto, Celso; Sperling, Felix A H

    2015-06-01

    The oriental fruit moth Grapholita molesta is one of the major pests of stone and pome fruit species in Brazil. Here, we applied 1226 SNPs obtained by genotyping-by-sequencing to test whether host species associations or other factors such as geographic distance structured populations of this pest. Populations from the main areas of occurrence of G. molesta were sampled principally from peach and apple orchards. Three main clusters were recovered by neighbor-joining analysis, all defined by geographic proximity between sampling localities. Overall genetic structure inferred by a nonhierarchical amova resulted in a significant ΦST value = 0.19109. Here, we demonstrate for the first time that SNPs gathered by genotyping-by-sequencing can be used to infer genetic structure of a pest insect in Brazil; moreover, our results indicate that those markers are very informative even over a restricted geographic scale. We also demonstrate that host plant association has little effect on genetic structure among Brazilian populations of G. molesta; on the other hand, reduced gene flow promoted by geographic isolation has a stronger impact on population differentiation. PMID:26029261

  20. Genotyping-by-sequencing approach indicates geographic distance as the main factor affecting genetic structure and gene flow in Brazilian populations of Grapholita molesta (Lepidoptera, Tortricidae)

    PubMed Central

    Silva-Brandão, Karina Lucas; Silva, Oscar Arnaldo Batista Neto e; Brandão, Marcelo Mendes; Omoto, Celso; Sperling, Felix A H

    2015-01-01

    The oriental fruit moth Grapholita molesta is one of the major pests of stone and pome fruit species in Brazil. Here, we applied 1226 SNPs obtained by genotyping-by-sequencing to test whether host species associations or other factors such as geographic distance structured populations of this pest. Populations from the main areas of occurrence of G. molesta were sampled principally from peach and apple orchards. Three main clusters were recovered by neighbor-joining analysis, all defined by geographic proximity between sampling localities. Overall genetic structure inferred by a nonhierarchical amova resulted in a significant ΦST value = 0.19109. Here, we demonstrate for the first time that SNPs gathered by genotyping-by-sequencing can be used to infer genetic structure of a pest insect in Brazil; moreover, our results indicate that those markers are very informative even over a restricted geographic scale. We also demonstrate that host plant association has little effect on genetic structure among Brazilian populations of G. molesta; on the other hand, reduced gene flow promoted by geographic isolation has a stronger impact on population differentiation. PMID:26029261

  1. DNA sequence of the control region of phage D108: the N-terminal amino acid sequences of repressor and transposase are similar both in phage D108 and in its relative, phage Mu.

    PubMed Central

    Mizuuchi, M; Weisberg, R A; Mizuuchi, K

    1986-01-01

    We have determined the DNA sequence of the control region of phage D108 up to position 1419 at the left end of the phage genome. Open reading frames for the repressor gene, ner gene, and the 5' part of the A gene (which codes for transposase) are found in the sequence. The genetic organization of this region of phage D108 is quite similar to that of phage Mu in spite of considerable divergence, both in the nucleotide sequence and in the amino acid sequences of the regulatory proteins of the two phages. The N-terminal amino acid sequences of the transposases of the two phages also share only limited homology. On the other hand, a significant amino acid sequence homology was found within each phage between the N-terminal parts of the repressor and transposase. We propose that the N-terminal domains of the repressor and transposase of each phage interact functionally in the process of making the decision between the lytic and the lysogenic mode of growth. PMID:3012481

  2. Polyriboadenylate Sequences at the 3′-Termini of Ribonucleic Acid Obtained from Mammalian Leukemia and Sarcoma Viruses

    PubMed Central

    Phillips, Leo A.; Park, James J.; Hollis, Vincent W.

    1974-01-01

    The location of poly(A) sequences in the RNA of mammalian RNA-tumor viruses was determined by enzymatic analyses. The 56-64S viral genomic RNAs, the 20-40S viral subunit RNAs, and the 4-5S poly(A) sequences excised from these viral RNAs were subjected to either hydrolysis with a 3′-OH specific exoribonuclease from Ehrlich ascites tumor cells or phosphorolysis from the 3′-termini with polynucleotide phosphorylase from Micrococcus luteus. Purified adenosine-labeled poly(A) fragments, excised from genomic viral RNAs by RNase A and T1 digestion, were hydrolyzed with the 3′-OH specific exoribonuclease for various periods of time. Poly(U) filter binding studies of the residual poly(A) indicated that 97% of the poly(A) fragments were hydrolyzed. Adenosine-labeled genomic and subunit viral RNAs and excised poly(A) fragments were phosphorolyzed from their 3′-termini for various periods of time with polynucleotide phosphorylase. The degree of phosphorolysis was monitored by poly(U) filter binding studies, and CCl3COOH insolubility and solubility determinations. There was an initial preferential rate of phosphorolysis of the poly(A) sequences of genomic and subunit viral RNAs as compared to the total adenosine-labeled viral RNAs. The data from these two different enzymatic mechanisms of action indicated conclusively that the poly(A) sequences were located at the 3′-termini of genomic and subunit viral RNAs. PMID:4373712

  3. Isolation and amino acid sequences of opossum vasoactive intestinal polypeptide and cholecystokinin octapeptide.

    PubMed Central

    Eng, J; Yu, J; Rattan, S; Yalow, R S

    1992-01-01

    Evolutionary history suggests that the marsupials entered South America from North America about 75 million years ago and subsequently dispersed into Australia before the separation between South America and Antarctica-Australia. A question of interest is whether marsupial peptides resemble the corresponding peptides of Old or New World mammals. Previous studies had shown that "little" gastrin of the North American marsupial, the opossum, is identical in length to that of the New World mammals, the guinea pig and chinchilla. In this report, we demonstrate that opossum cholecystokinin octapeptide, like that of the Australian marsupials, the Eastern quoll and the Tamar wallaby, is identical to the cholecystokinin octapeptide of Old World mammals and differs from that of the guinea pig and chinchilla. However, opossum vasoactive intestinal polypeptide differs from the usual Old World mammalian vasoactive intestinal polypeptide in five sites: [sequence; see text]. PMID:1542675

  4. Evolution of early life inferred from protein and ribonucleic acid sequences

    NASA Technical Reports Server (NTRS)

    Dayhoff, M. O.; Schwartz, R. M.

    1978-01-01

    The chemical structures of ferredoxin, 5S ribosomal RNA, and c-type cytochrome sequences have been employed to construct a phylogenetic tree which connects all major photosynthesizing organisms: the three types of bacteria, blue-green algae, and chloroplasts. Anaerobic and aerobic bacteria, eukaryotic cytoplasmic components and mitochondria are also included in the phylogenetic tree. Anaerobic nonphotosynthesizing bacteria similar to Clostridium were the earliest organisms, arising more than 3.2 billion years ago. Bacterial photosynthesis evolved nearly 3.0 billion years ago, while oxygen-evolving photosynthesis, originating in the blue-green algal line, came into being about 2.0 billion years ago. The phylogenetic tree supports the symbiotic theory of the origin of eukaryotes.

  5. Amino acid sequence and molecular modelling of glycoprotein IIb-IIIa and fibronectin receptor iso-antagonists from Trimeresurus elegans venom.

    PubMed Central

    Scaloni, A; Di Martino, E; Miraglia, N; Pelagalli, A; Della Morte, R; Staiano, N; Pucci, P

    1996-01-01

    Low-molecular-mass Arg-Gly-Asp (RGD)-containing polypeptides were isolated from the venom of Trimeresurus elegans by a simple two-step procedure consisting of membrane filtration and reverse-phase HPLC. A combination of electrospray MS, fast-atom bombardment MS and Edman degradation allowed us to ascertain the presence in the venom of different isoforms and to determine their primary structures. The amino acid sequences resembled the structure of elegantin, the only disintegrin previously reported from the T. elegans venom [Williams, Rucinski, Holt and Niewiarowski (1990) Biochim. Biophys, Acta 1039, 81-89]. MS analyses indicated the occurrence of differential proteolytic processing at both the N-terminus and the C-termins of the polypeptide chains. The amino acid sequence alignment of the elegantin isoforms with known components of the disintegrin family demonstrated the complete conservation of the 12 cysteine residues involved in disulphide bridges. Molecular modelling of elegantins predicted an overall folding of these molecules quite similar to that reported for the kistrin solution structure. The newly identified polypeptide isoforms strongly inhibited ADP-induced aggregation in both human and canine platelet-rich plasma but showed a different species-dependent specificity. These molecules were also able to inhibit B16-BL6 murine melanoma cell adhesion to immobilized fibronectin. The comparison of the structures and biological activities of elegantin isoforms and kistrin allowed us to highlight some structural features that, in addition to the RGD locus might be involved in the interaction of these snake-venom polypeptides with the integrin receptors on the platelet and cell surface. PMID:8920980

  6. Complete amino acid sequence and characterization of the reaction mechanism of a glucosamine-induced novel alcohol dehydrogenase from Agrobacterium radiobacter (tumefaciens).

    PubMed

    Iwamoto, Ryoko; Kubota, Humie; Hosoki, Tomoko; Ikehara, Kenji; Tanaka, Mieko

    2002-02-15

    A glucosamine-induced novel alcohol dehydrogenase has been isolated from Agrobacterium radiobacter (tumefaciens) and its fundamental properties have been characterized. The enzyme catalyzes NAD-dependent dehydrogenation of aliphatic alcohols and amino alcohols. In this work, the complete amino acid sequence of the alcohol dehydrogenase was determined by PCR method using genomic DNA of A. radiobacter as template. The enzyme comprises 336 amino acids and has a molecular mass of 36 kDa. The primary structure of the enzyme demonstrates a high homology to structures of alcohol dehydrogenases from Shinorhizobium meliloti (83% identity, 90% positive) and Pseudomonas aeruginosa (65% identity, 76% positive). The two Zn(2+) ion binding sites, both the active site and another site that contributed to stabilization of the enzyme, are conserved in those enzymes. Sequences analysis of the NAD-dependent dehydrogenase family using a hypothetical phylogenetic tree indicates that these three enzymes form a new group distinct from other members of the Zn-containing long-chain alcohol dehydrogenase family. The physicochemical properties of alcohol dehydrogenase from A. radiobacter were characterized as follows. (1) Stereospecificity of the hydride transfer from ethanol to NADH was categorized as pro-R type by NMR spectra of NADH formed in the enzymatic reaction using ethanol-D(6) was used as substrate. (2) Optimal pH for all alcohols with no amino group examined was pH 8.5 (of the C(2)-C(6) alcohols, n-amyl alcohol demonstrated the highest activity). Conversely, glucosaminitol was optimally dehydrogenated at pH 10.0. (3) The rate-determining step of the dehydrogenase for ethanol is deprotonation of the enzyme-NAD-Zn-OHCH(2)CH(3) complex to enzyme-NAD-Zn-O(-)CH(2)CH(3) complex and that for glucosaminitol is H(2)O addition to enzyme-Zn-NADH complex. PMID:11831851

  7. A Possible Mechanism of Zika Virus Associated Microcephaly: Imperative Role of Retinoic Acid Response Element (RARE) Consensus Sequence Repeats in the Viral Genome.

    PubMed

    Kumar, Ashutosh; Singh, Himanshu N; Pareek, Vikas; Raza, Khursheed; Dantham, Subrahamanyam; Kumar, Pavan; Mochan, Sankat; Faiq, Muneeb A

    2016-01-01

    Owing to the reports of microcephaly as a consistent outcome in the fetuses of pregnant women infected with ZIKV in Brazil, Zika virus (ZIKV)-microcephaly etiomechanistic relationship has recently been implicated. Researchers, however, are still struggling to establish an embryological basis for this interesting causal handcuff. The present study reveals robust evidence in favor of a plausible ZIKV-microcephaly cause-effect liaison. The rationale is based on: (1) sequence homology between ZIKV genome and the response element of an early neural tube developmental marker "retinoic acid" in human DNA and (2) comprehensive similarities between the details of brain defects in ZIKV-microcephaly and retinoic acid embryopathy. Retinoic acid is considered as the earliest factor for regulating anteroposterior axis of neural tube and positioning of structures in developing brain through retinoic acid response elements (RARE) consensus sequence (5'-AGGTCA-3') in promoter regions of retinoic acid-dependent genes. We screened genomic sequences of already reported virulent ZIKV strains (including those linked to microcephaly) and other viruses available in National Institute of Health genetic sequence database (GenBank) for the RARE consensus repeats and obtained results strongly bolstering our hypothesis that ZIKV strains associated with microcephaly may act through precipitation of dysregulation in retinoic acid-dependent genes by introducing extra stretches of RARE consensus sequence repeats in the genome of developing brain cells. Additional support to our hypothesis comes from our findings that screening of other viruses for RARE consensus sequence repeats is positive only for those known to display neurotropism and cause fetal brain defects (for which maternal-fetal transmission during developing stage may be required). The numbers of RARE sequence repeats appeared to match with the virulence of screened positive viruses. Although, bioinformatic evidence and embryological

  8. Analyses of mitochondrial amino acid sequence datasets support the proposal that specimens of Hypodontus macropi from three species of macropodid hosts represent distinct species

    PubMed Central

    2013-01-01

    Background Hypodontus macropi is a common intestinal nematode of a range of kangaroos and wallabies (macropodid marsupials). Based on previous multilocus enzyme electrophoresis (MEE) and nuclear ribosomal DNA sequence data sets, H. macropi has been proposed to be complex of species. To test this proposal using independent molecular data, we sequenced the whole mitochondrial (mt) genomes of individuals of H. macropi from three different species of hosts (Macropus robustus robustus, Thylogale billardierii and Macropus [Wallabia] bicolor) as well as that of Macropicola ocydromi (a related nematode), and undertook a comparative analysis of the amino acid sequence datasets derived from these genomes. Results The mt genomes sequenced by next-generation (454) technology from H. macropi from the three host species varied from 13,634 bp to 13,699 bp in size. Pairwise comparisons of the amino acid sequences predicted from these three mt genomes revealed differences of 5.8% to 18%. Phylogenetic analysis of the amino acid sequence data sets using Bayesian Inference (BI) showed that H. macropi from the three different host species formed distinct, well-supported clades. In addition, sliding window analysis of the mt genomes defined variable regions for future population genetic studies of H. macropi in different macropodid hosts and geographical regions around Australia. Conclusions The present analyses of inferred mt protein sequence datasets clearly supported the hypothesis that H. macropi from M. robustus robustus, M. bicolor and T. billardierii represent distinct species. PMID:24261823

  9. Using Triple Helix Forming Peptide Nucleic Acids for Sequence-selective Recognition of Double-stranded RNA

    PubMed Central

    Hnedzko, Dziyana; Cheruiyot, Samwel K.; Rozners, Eriks

    2014-01-01

    Non-coding RNAs play important roles in regulation of gene expression. Specific recognition and inhibition of these biologically important RNAs that form complex double-helical structures will be highly useful for fundamental studies in biology and practical applications in medicine. This protocol describes a strategy developed in our laboratory for sequence-selective recognition of double-stranded RNA (dsRNA) using triple helix forming peptide nucleic acids (PNAs) that bind in the major grove of RNA helix. The strategy developed uses chemically modified nucleobases, such as 2-aminopyridine (M) that enables strong triple helical binding at physiologically relevant conditions, and 2-pyrimidinone (P) and 3-oxo-2,3-dihydropyridazine (E) that enable recognition of isolated pyrimidines in the purine rich strand of the RNA duplex. Detailed protocols for preparation of modified PNA monomers, solid-phase synthesis and HPLC purification of PNA oligomers, and measuring dsRNA binding affinity using isothermal titration calorimetry are included. PMID:25199637

  10. Nucleic acid sequences encoding D1 and D1/D2 domains of human coxsackievirus and adenovirus receptor (CAR)

    DOEpatents

    Freimuth, Paul I.

    2010-04-06

    The invention provides recombinant human CAR (coxsackievirus and adenovirus receptor) polypeptides which bind adenovirus. Specifically, polypeptides corresponding to adenovirus binding domain D1 and the entire extracellular domain of human CAR protein comprising D1 and D2 are provided. In another aspect, the invention provides nucleic acid sequences encoding these domains and expression vectors for producing the domains and bacterial cells containing such vectors. The invention also includes an isolated fusion protein comprised of the D1 polypeptide fused to a polypeptide which facilitates folding of D1 when expressed in bacteria. The functional D1 domain finds application in a therapeutic method for treating a patient infected with a CAR D1-binding virus, and also in a method for identifying an antiviral compound which interferes with viral attachment. The invention also provides a method for specifically targeting a cell for infection by a virus which binds to D1.

  11. Prediction of Residue Status to Be Protected or Not Protected From Hy-drogen Exchange Using Amino Acid Sequence Only.

    PubMed

    Nikita V, Dovidchenko; Oxana V, Galzitskaya

    2008-01-01

    We have outlined here some structural aspects of local flexibility. Important functional properties are related to flexible segments. We try to predict regions that have been shown to exhibit the highest probability of being folded in the equilibrium intermediate or native state and will be protected from hydrogen exchange using amino acid sequence only. Our approach FoldUnfold for the prediction of unstructured regions has been applied to seven different proteins. For 80% of the residues considered in this paper we can predict correctly their status: will they be protected or not from hydrogen exchange. An additional goal of our study is to assess whether properties inferred using the bioinformatics approach are easily applicable to predict behavior of proteins in solution. PMID:18949078

  12. Prediction of Residue Status to Be Protected or Not Protected From Hy-drogen Exchange Using Amino Acid Sequence Only

    PubMed Central

    Dovidchenko, Nikita V; Galzitskaya, Oxana V

    2008-01-01

    We have outlined here some structural aspects of local flexibility. Important functional properties are related to flexible segments. We try to predict regions that have been shown to exhibit the highest probability of being folded in the equilibrium intermediate or native state and will be protected from hydrogen exchange using amino acid sequence only. Our approach FoldUnfold for the prediction of unstructured regions has been applied to seven different proteins. For 80% of the residues considered in this paper we can predict correctly their status: will they be protected or not from hydrogen exchange. An additional goal of our study is to assess whether properties inferred using the bioinformatics approach are easily applicable to predict behavior of proteins in solution. PMID:18949078

  13. Ferulic acid release and 4-vinylguaiacol formation during brewing and fermentation: indications for feruloyl esterase activity in Saccharomyces cerevisiae.

    PubMed

    Coghe, Stefan; Benoot, Koen; Delvaux, Filip; Vanderhaegen, Bart; Delvaux, Freddy R

    2004-02-11

    The release of ferulic acid and the subsequent thermal or enzymatic decarboxylation to 4-vinylguaiacol are inherent to the beer production process. Phenolic, medicinal, or clove-like flavors originating from 4-vinylguaiacol frequently occur in beer made with wheat or wheat malt. To evaluate the release of ferulic acid and the transformation to 4-vinylguaiacol, beer was brewed with different proportions of barley malt, wheat, and wheat malt. Ferulic acid as well as 4-vinylguaiacol levels were determined by HPLC at several stages of the beer production process. During brewing, ferulic acid was released at the initial mashing phase, whereas moderate levels of 4-vinylguaiacol were formed by wort boiling. Higher levels of the phenolic flavor compound were produced during fermentations with brewery yeast strains of the Pof(+) phenotype. In beer made with barley malt, ferulic acid was mainly released during the brewing process. Conversely, 60-90% of ferulic acid in wheat or wheat malt beer was hydrolyzed during fermentation, causing higher 4-vinylguaiacol levels in these beers. As cereal enzymes are most likely inactivated during wort boiling, the additional release of ferulic acid during fermentation suggests the activity of feruloyl esterases produced by brewer's yeast. PMID:14759156

  14. Primary structure of a histidine-rich proteolytic fragment of human ceruloplasmin. II. Amino acid sequence of the tryptic peptides.

    PubMed

    Kingston, I B; Kingston, B L; Putnam, F W

    1980-04-10

    Amino acid sequence studies of tryptic peptides isolated from a histidine-rich fragment (Cp F5) of human ceruloplasmin are described. Nineteen tryptic peptides were isolated from unmodified Cp F5 and five tryptic peptides were isolated from citraconylated Cp F5. These peptides, together with the cyanogen bromide fragments reported previously, allowed the assembly of the complete sequence of Cp F5. The fragment has 159 residues and a molecular weight of 18,650; it lacks carbohydrate, is rich in histidine, and contains 1 free cysteine that may be part of a copper-binding site. Human ceruloplasmin is a single polypeptide chain with a molecular weight of about 130,000 that is readily cleaved to large fragments by proteolytic enzymes; the relationships of Cp F5 to intact ceruloplasmin and to structural subunits earlier proposed is described. Cp F5 probably is an intact globular domain that is attached to the COOH-terminal end of ceruloplasmin by a labile interdomain peptide bond. PMID:6987230

  15. Ambient temperature detection of PCR amplicons with a novel sequence-specific nucleic acid lateral flow biosensor.

    PubMed

    Ang, Geik Yong; Yu, Choo Yee; Yean, Chan Yean

    2012-01-01

    In the field of diagnostics, molecular amplification targeting unique genetic signature sequences has been widely used for rapid identification of infectious agents, which significantly aids physicians in determining the choice of treatment as well as providing important epidemiological data for surveillance and disease control assessment. We report the development of a rapid nucleic acid lateral flow biosensor (NALFB) in a dry-reagent strip format for the sequence-specific detection of single-stranded polymerase chain reaction (PCR) amplicons at ambient temperature (22-25°C). The NALFB was developed in combination with a linear-after-the-exponential PCR assay and the applicability of this biosensor was demonstrated through detection of the cholera toxin gene from diarrheal-causing toxigenic Vibrio cholerae. Amplification using the advanced asymmetric PCR boosts the production of fluorescein-labeled single-stranded amplicons, allowing capture probes immobilized on the NALFB to hybridize specifically with complementary targets in situ on the strip. Subsequent visual formation of red lines is achieved through the binding of conjugated gold nanoparticles to the fluorescein label of the captured amplicons. The visual detection limit observed with synthetic target DNA was 0.3 ng and 1 pg with pure genomic DNA. Evaluation of the NALFB with 164 strains of V. cholerae and non-V. cholerae bacteria recorded 100% for both sensitivity and specificity. The whole procedure of the low-cost NALFB, which is performed at ambient temperature, eliminates the need for preheated buffers or additional equipment, greatly simplifying the protocol for sequence-specific PCR amplicon analysis. PMID:22705404

  16. Compartmentation of malic acid in mesophyll cells of Kalanchoee daigremontiana: indications of a intracellular cytosolic vesicle transport mechanism

    SciTech Connect

    Balsamo, R.A.; Uribe, E.G.

    1987-04-01

    Leaf tissue was harvested over a 24hr period at one to three hour intervals. The malic acid levels in the tissue were assayed spectrophotometrically and the percent cell volume occupied by cytosolic vesicles in replicate samples was determined. The total volume of the cytosolic vesicles fluctuated throughout the photoperiod concommitantly with malic acid concentrations present in the tissue. An intact leaf tissue section (10.2cm/sup 2/) was radiolabeled with /sup 14/CO/sub 2/ seven hours into the dark period for thirty minutes. Two dimensional thin layer chromatography and electrophoresis of the tissue determined that 96% of the label was incorporated into malic acid. A freeze substitution procedure was initiated followed by microautoradiography (Fisher 1971) which allowed for the tracing of intracellular malic acid migration and compartmentation within the mesophyll cells. The results and interpretation of this experiment will be presented.

  17. Primary structure of a histidine-rich proteolytic fragment of human ceruloplasmin. I. Amino acid sequence of the cyanogen bromide peptides.

    PubMed

    Kingston, I B; Kingston, B L; Putnam, F W

    1980-04-10

    A histidine-rich fragment, Cp F5, with a molecular weight of 18,650 was isolated from human ceruloplasmin. It consists of 159 amino acids and contains a possible copper-binding site. The sequence of the first 18 NH2-terminal residues of Cp F5 was determined by automated Edman degradation. Cp F5 was cleaved by cyanogen bromide to produce nine fragments of from 2 to 63 residues. The amino acid sequence of all of the cyanogen bromide fragments was investigated using automated and manual Edman degradation, the fragments being digested with trypsin, chymotrypsin, thermolysin, staphylococcal protease, and pepsin as appropriate. The results, in conjunction with the data on the tryptic peptides reported in the accompanying paper (Kingston, I.B., Kingston, B.L., and Putnam, F.L. (1980) J. Biol. Chem. 255, 2886-2896), establish the complete amino acid sequence of Cp F5. PMID:6987229

  18. EGENES: Transcriptome-Based Plant Database of Genes with Metabolic Pathway Information and Expressed Sequence Tag Indices in KEGG1[C][W][OA

    PubMed Central

    Masoudi-Nejad, Ali; Goto, Susumu; Jauregui, Ruy; Ito, Masumi; Kawashima, Shuichi; Moriya, Yuki; Endo, Takashi R.; Kanehisa, Minoru

    2007-01-01

    EGENES is a knowledge-based database for efficient analysis of plant expressed sequence tags (ESTs) that was recently added to the KEGG suite of databases. It links plant genomic information with higher order functional information in a single database. It also provides gene indices for each genome. The genomic information in EGENES is a collection of EST contigs constructed from assembly of ESTs. Due to the extremely large genomes of plant species, the bulk collection of data such as ESTs is a quick way to capture a complete repertoire of genes expressed in an organism. Using ESTs for reconstructing metabolic pathways is a new expansion in KEGG and provides researchers with a new resource for species in which only EST sequences are available. Functional annotation in EGENES is a process of linking a set of genes/transcripts in each genome with a network of interacting molecules in the cell. EGENES is a multispecies, integrated resource consisting of genomic, chemical, and network information containing a complete set of building blocks (genes and molecules) and wiring diagrams (biological pathways) to represent cellular functions. Using EGENES, genome-based pathway annotation and EST-based annotation can now be compared and mutually validated. The ultimate goals of EGENES will be to: bring new plant species into KEGG by clustering and annotating ESTs; abstract knowledge and principles from large-scale plant EST data; and improve computational prediction of systems of higher complexity. EGENES will be updated at least once a year. EGENES is publicly available and is accessible by the following link or by KEGG's navigation system (http://www.genome.jp/kegg-bin/create_kegg_menu?category=plants_egenes). PMID:17468225

  19. Compound-specific isotopes of fatty acids as indicators of trophic interactions in the East China Sea ecosystem

    NASA Astrophysics Data System (ADS)

    Wu, Ying; Wang, Na; Zhang, Jing; Wan, Ruijing; Dai, Fangqun; Jin, Xianshi

    2016-09-01

    The composition and compound-specific isotopes of fatty acids were studied within food webs in the East China Sea. Lipid-normalized stable carbon isotopes of total organic carbon had a good correlation with trophic level. Variations in fatty acid compositions among diff erent species were observed but were unclear. Diff erent dietary structures could be traced from molecular isotopes of selected fatty acids in the Shiba shrimp ( Matapenaeus joyneri), the coastal mud shrimp ( Solenocera crassicornis) and the northern Maoxia shrimp ( Acetes chinensis). Both M. joyneri and S. crassicornis are mainly benthos feeders, while A. chinensis is a pelagic species, although they have a similar fatty acid composition. There was a good correlation for isotopes of arachidonic acid (C20:4n6; ARA) and docosahexaenoic acid (C22:6n3; DHA) among pelagic species from higher trophic levels. The isotopic compositions of DHA in benthic species were more negative than those of pelagic species at the same trophic level. The fact that the diet of benthic species contains more degraded items, the carbon isotopes of which are derived from a large biochemical fraction, may be the reason for this variation. A comparative study of benthic and pelagic species demonstrated the diff erent carbon sources in potential food items and the presence of a more complex system at the water-sediment interface.

  20. Compound-specific isotopes of fatty acids as indicators of trophic interactions in the East China Sea ecosystem

    NASA Astrophysics Data System (ADS)

    Wu, Ying; Wang, Na; Zhang, Jing; Wan, Ruijing; Dai, Fangqun; Jin, Xianshi

    2016-01-01

    The composition and compound-specific isotopes of fatty acids were studied within food webs in the East China Sea. Lipid-normalized stable carbon isotopes of total organic carbon had a good correlation with trophic level. Variations in fatty acid compositions among diff erent species were observed but were unclear. Diff erent dietary structures could be traced from molecular isotopes of selected fatty acids in the Shiba shrimp (Matapenaeus joyneri), the coastal mud shrimp (Solenocera crassicornis) and the northern Maoxia shrimp (Acetes chinensis). Both M. joyneri and S. crassicornis are mainly benthos feeders, while A. chinensis is a pelagic species, although they have a similar fatty acid composition. There was a good correlation for isotopes of arachidonic acid (C20:4n6; ARA) and docosahexaenoic acid (C22:6n3; DHA) among pelagic species from higher trophic levels. The isotopic compositions of DHA in benthic species were more negative than those of pelagic species at the same trophic level. The fact that the diet of benthic species contains more degraded items, the carbon isotopes of which are derived from a large biochemical fraction, may be the reason for this variation. A comparative study of benthic and pelagic species demonstrated the diff erent carbon sources in potential food items and the presence of a more complex system at the water-sediment interface.

  1. Significant Association between Sulfate-Reducing Bacteria and Uranium-Reducing Microbial Communities as Revealed by a Combined Massively Parallel Sequencing-Indicator Species Approach

    SciTech Connect

    Cardenas, Erick; Leigh, Mary Beth; Marsh, Terence; Tiedje, James M.; Wu, Wei-min; Luo, Jian; Ginder-Vogel, Matthew; Kitanidis, Peter K.; Criddle, Craig; Carley, Jack M; Carroll, Sue L; Gentry, Terry J; Watson, David B; Gu, Baohua; Jardine, Philip M; Zhou, Jizhong

    2010-10-01

    Massively parallel sequencing has provided a more affordable and high-throughput method to study microbial communities, although it has mostly been used in an exploratory fashion. We combined pyrosequencing with a strict indicator species statistical analysis to test if bacteria specifically responded to ethanol injection that successfully promoted dissimilatory uranium(VI) reduction in the subsurface of a uranium contamination plume at the Oak Ridge Field Research Center in Tennessee. Remediation was achieved with a hydraulic flow control consisting of an inner loop, where ethanol was injected, and an outer loop for flow-field protection. This strategy reduced uranium concentrations in groundwater to levels below 0.126 M and created geochemical gradients in electron donors from the inner-loop injection well toward the outer loop and downgradient flow path. Our analysis with 15 sediment samples from the entire test area found significant indicator species that showed a high degree of adaptation to the three different hydrochemical-created conditions. Castellaniella and Rhodanobacter characterized areas with low pH, heavy metals, and low bioactivity, while sulfate-, Fe(III)-, and U(VI)-reducing bacteria (Desulfovibrio, Anaeromyxobacter, and Desulfosporosinus) were indicators of areas where U(VI) reduction occurred. The abundance of these bacteria, as well as the Fe(III) and U(VI) reducer Geobacter, correlated with the hydraulic connectivity to the substrate injection site, suggesting that the selected populations were a direct response to electron donor addition by the groundwater flow path. A false-discovery-rate approach was implemented to discard false-positive results by chance, given the large amount of data compared.

  2. Significant association between sulfate-reducing bacteria and uranium-reducing microbial communities as revealed by a combined massively parallel sequencing-indicator species approach.

    PubMed

    Cardenas, Erick; Wu, Wei-Min; Leigh, Mary Beth; Carley, Jack; Carroll, Sue; Gentry, Terry; Luo, Jian; Watson, David; Gu, Baohua; Ginder-Vogel, Matthew; Kitanidis, Peter K; Jardine, Philip M; Zhou, Jizhong; Criddle, Craig S; Marsh, Terence L; Tiedje, James M

    2010-10-01

    Massively parallel sequencing has provided a more affordable and high-throughput method to study microbial communities, although it has mostly been used in an exploratory fashion. We combined pyrosequencing with a strict indicator species statistical analysis to test if bacteria specifically responded to ethanol injection that successfully promoted dissimilatory uranium(VI) reduction in the subsurface of a uranium contamination plume at the Oak Ridge Field Research Center in Tennessee. Remediation was achieved with a hydraulic flow control consisting of an inner loop, where ethanol was injected, and an outer loop for flow-field protection. This strategy reduced uranium concentrations in groundwater to levels below 0.126 μM and created geochemical gradients in electron donors from the inner-loop injection well toward the outer loop and downgradient flow path. Our analysis with 15 sediment samples from the entire test area found significant indicator species that showed a high degree of adaptation to the three different hydrochemical-created conditions. Castellaniella and Rhodanobacter characterized areas with low pH, heavy metals, and low bioactivity, while sulfate-, Fe(III)-, and U(VI)-reducing bacteria (Desulfovibrio, Anaeromyxobacter, and Desulfosporosinus) were indicators of areas where U(VI) reduction occurred. The abundance of these bacteria, as well as the Fe(III) and U(VI) reducer Geobacter, correlated with the hydraulic connectivity to the substrate injection site, suggesting that the selected populations were a direct response to electron donor addition by the groundwater flow path. A false-discovery-rate approach was implemented to discard false-positive results by chance, given the large amount of data compared. PMID:20729318

  3. A combined massively parallel sequencing indicator species approach revealed significant association between sulfate-reducing bacteria and uranium-reducing microbial communities

    SciTech Connect

    Cardenas, Erick; Wu, Wei-min; Leigh, Mary Beth; Carley, Jack M; Carroll, Sue L; Gentry, Terry; Luo, Jian; Watson, David B; Gu, Baohua; Ginder-Vogel, Matthew A.; Kitanidis, Peter K.; Jardine, Philip; Kelly, Shelly D; Zhou, Jizhong; Criddle, Craig; Marsh, Terence; Tiedje, James

    2010-08-01

    Massively parallel sequencing has provided a more affordable and high throughput method to study microbial communities, although it has been mostly used in an exploratory fashion. We combined pyrosequencing with a strict indicator species statistical analysis to test if bacteria specifically responded to ethanol injection that successfully promoted dissimilatory uranium (VI) reduction in the subsurface of a uranium contamination plume at the Oak Ridge Field Research Center in Tennessee, USA. Remediation was achieved with a hydraulic flow control consisting of an inner loop, where ethanol was injected, and an outer loop for flow field protection. This strategy reduced uranium concentrations in groundwater to levels below 0.126 {micro}M, and created geochemical gradients in electron donors from the inner loop injection well towards the outer loop and down-gradient flow path. Our analysis with 15 sediment samples from the entire test area found significant indicator species that showed a high degree of adaptation to the three different hydrochemical created conditions. Castellaniella, and Rhodanobacter characterized areas with low pH, heavy metals, and low bioactivity; while sulfate-, Fe(III)-, and U(VI)-reducing bacteria (Desulfovibrio, Anaeromyxobacter, and Desulfosporosinus) were indicators of areas where U(VI) reduction occurred. Abundance of these bacteria as well as the Fe(III)- and U(VI)-reducer Geobacter correlated with the hydraulic connectivity to the substrate injection site, suggesting that the selected populations were a direct response to the electron donor addition and by the groundwater flow path. A false discovery rate approach was implemented to discard false positives by chance given the large amount of data compared.

  4. Protective immunogenicity of two synthetic peptides selected from the amino acid sequence of Bordetella pertussis toxin subunit S1.

    PubMed Central

    Askelöf, P; Rodmalm, K; Wrangsell, G; Larsson, U; Svenson, S B; Cowell, J L; Undén, A; Bartfai, T

    1990-01-01

    Two peptides, corresponding to amino acids 1-17 and 169-186 of the amino acid sequence of pertussis toxin (PT) subunit S1, were synthesized and coupled to the diphtheria toxin cross-reactive mutant protein CRM 197 and evaluated for immunogenicity and protective capacity against PT challenge in vivo. The peptide-CRM conjugates induced high antibody titers against native toxin in mice (BALB/c, C57/Black, and outbred NMRI) as measured by ELISA. Upon PT challenge (0.5 microgram of toxin) of the NMRI mice, the CRM conjugates of peptides 1-17 and 169-186 fully protected the mice from PT-induced leukocytosis. Immunization with the corresponding bovine serum albumin conjugates of these two peptides also fully protected mice. Rabbit antiserum to the peptide 1-17-CRM conjugate was highly efficient in inhibiting the ADP-ribosylating activity of PT but did not neutralize the clustering effect of PT on Chinese hamster ovary cells. In contrast, the rabbit antiserum raised against the peptide 169-186-CRM conjugate neutralized the clustering effect of PT on Chinese hamster ovary cells but did not inhibit the enzymatic activity of PT. Peptide 169-186-CRM conjugates mimic the immunoglobulin binding properties of PT and also cause clustering of Chinese hamster ovary cells. The CRM conjugates of these two peptides constitute a synthetic pertussis vaccine candidate with the ability to provide a chemically well-defined, safe, and efficient pertussis vaccine. Images PMID:2304902

  5. Hybridization properties of long nucleic acid probes for detection of variable target sequences, and development of a hybridization prediction algorithm

    PubMed Central

    Öhrmalm, Christina; Jobs, Magnus; Eriksson, Ronnie; Golbob, Sultan; Elfaitouri, Amal; Benachenhou, Farid; Strømme, Maria; Blomberg, Jonas

    2010-01-01

    One of the main problems in nucleic acid-based techniques for detection of infectious agents, such as influenza viruses, is that of nucleic acid sequence variation. DNA probes, 70-nt long, some including the nucleotide analog deoxyribose-Inosine (dInosine), were analyzed for hybridization tolerance to different amounts and distributions of mismatching bases, e.g. synonymous mutations, in target DNA. Microsphere-linked 70-mer probes were hybridized in 3M TMAC buffer to biotinylated single-stranded (ss) DNA for subsequent analysis in a Luminex® system. When mismatches interrupted contiguous matching stretches of 6 nt or longer, it had a strong impact on hybridization. Contiguous matching stretches are more important than the same number of matching nucleotides separated by mismatches into several regions. dInosine, but not 5-nitroindole, substitutions at mismatching positions stabilized hybridization remarkably well, comparable to N (4-fold) wobbles in the same positions. In contrast to shorter probes, 70-nt probes with judiciously placed dInosine substitutions and/or wobble positions were remarkably mismatch tolerant, with preserved specificity. An algorithm, NucZip, was constructed to model the nucleation and zipping phases of hybridization, integrating both local and distant binding contributions. It predicted hybridization more exactly than previous algorithms, and has the potential to guide the design of variation-tolerant yet specific probes. PMID:20864443

  6. Nucleic acid amplification in vitro: detection of sequences with low copy numbers and application to diagnosis of human immunodeficiency virus type 1 infection.

    PubMed Central

    Guatelli, J C; Gingeras, T R; Richman, D D

    1989-01-01

    The enzymatic amplification of specific nucleic acid sequences in vitro has revolutionized the use of nucleic acid hybridization assays for viral detection. With this method, the copy number of a pathogen-specific sequence is increased several orders of magnitude before detection is attempted. The sensitivity and specificity of detection are thus markedly improved. Mullis and Faloona devised the first method of sequence amplification in vitro, the polymerase chain reaction (K.B. Mullis and F.A. Faloona, Methods Enzymol. 155:355-350, 1987). By this method, synthetic oligonucleotide primers direct repeated, target-specific, deoxyribonucleic acid-synthetic reactions, resulting in an exponential increase in the amount of the specific target sequence. The application of sequence amplification to viral detection was initially performed with human immunodeficiency virus type 1 and human T-cell lymphoma virus type I. In principle, however, this approach can be applied to the detection of any deoxyribonucleic or ribonucleic acid virus; the only requirement is that sufficient nucleotide sequence data exist to allow the synthesis of target-specific oligonucleotide primers. The use of target amplification in vitro will permit a variety of studies of viral pathogenesis which have not been feasible because of the low copy number of the viral nucleic acids in infected material. This approach is particularly applicable to the study of human retroviral infections, which are chronic and persistent and are characterized by low titers of virus in tissues. In addition, target amplification in vitro will facilitate the development of new methods of sequence detection, which will be useful for rapid viral diagnosis in the clinical laboratory. PMID:2650862

  7. The outer capsid protein VP4 of equine rotavirus strain H-2 represents a unique VP4 type by amino acid sequence analysis.

    PubMed

    Hardy, M E; Gorziglia, M; Woode, G N

    1993-03-01

    The nucleotide and deduced amino acid sequence of G serotype 3 equine rotavirus strain H-2 was determined. A predicted 776-amino-acid H-2 VP4 shows less than or equal to 85.3% identity to other rotavirus VP4 types sequenced to date and thus represents a new P serotype. A PCR-generated probe derived from a cDNA clone of H-2 gene 4 hybridized to gene 4 of several tissue-culture-adapted equine rotavirus isolates, demonstrating that the gene 4 allele present in the H-2 strain is present in the equine rotavirus population. PMID:8382410

  8. Some considerations regarding the use of amino acid racemization in human dentine as an indicator of age at death.

    PubMed

    Carolan, V A; Gardner, M L; Lucy, D; Pollard, A M

    1997-01-01

    An HPLC method is described for simultaneously obtaining the enantiomeric ratio of three amino acids (aspartic acid, serine, and glutamic acid) from dental collagen, with a view to using this information for estimating age at death. Results are reported from a sample of twenty three known age modern teeth, six known age 19th C. AD teeth, and two unknown age Romano-British teeth. It was found (as expected) that all three D/L ratios changed significantly with chronological age. Standard calibration techniques were used to estimate ages for the six 19th C. AD specimens from regression equations estimated from the modern specimens, and also to predict (for the first time) the error associated with such estimates. Errors using aspartic acid were found to be similar to those obtained by other methods of age estimation from dental evidence, serine, and glutamic acid providing much poorer age estimates. Additionally, a systematic difference in the age-enantiomeric ratio relationship was observed between modern and older dental samples. It is concluded that there is some fundamental difference in the observed enantiomeric ratios between modern teeth and older samples, possibly as a result of the chemical alteration of the dental proteins. PMID:8988569

  9. Complete Genome Sequence of the d-Amino Acid Catabolism Bacterium Phaeobacter sp. Strain JL2886, Isolated from Deep Seawater of the South China Sea.

    PubMed

    Fu, Yingnan; Wang, Rui; Zhang, Zilian; Jiao, Nianzhi

    2016-01-01

    Phaeobacter sp. strain JL2886, isolated from deep seawater of the South China Sea, can catabolize d-amino acids. Here, we report the complete genome sequence of Phaeobacter sp. JL2886. It comprises ~4.06 Mbp, with a G+C content of 61.52%. A total of 3,913 protein-coding genes and 10 genes related to d-amino acid catabolism were obtained. PMID:27587825

  10. Complete Genome Sequence of the d-Amino Acid Catabolism Bacterium Phaeobacter sp. Strain JL2886, Isolated from Deep Seawater of the South China Sea

    PubMed Central

    Fu, Yingnan; Wang, Rui

    2016-01-01

    Phaeobacter sp. strain JL2886, isolated from deep seawater of the South China Sea, can catabolize d-amino acids. Here, we report the complete genome sequence of Phaeobacter sp. JL2886. It comprises ~4.06 Mbp, with a G+C content of 61.52%. A total of 3,913 protein-coding genes and 10 genes related to d-amino acid catabolism were obtained. PMID:27587825

  11. The highly conserved amino acid sequence motif Tyr-Gly-Asp-Thr-Asp-Ser in alpha-like DNA polymerases is required by phage phi 29 DNA polymerase for protein-primed initiation and polymerization.

    PubMed Central

    Bernad, A; Lázaro, J M; Salas, M; Blanco, L

    1990-01-01

    The alpha-like DNA polymerases from bacteriophage phi 29 and other viruses, prokaryotes and eukaryotes contain an amino acid consensus sequence that has been proposed to form part of the dNTP binding site. We have used site-directed mutants to study five of the six highly conserved consecutive amino acids corresponding to the most conserved C-terminal segment (Tyr-Gly-Asp-Thr-Asp-Ser). Our results indicate that in phi 29 DNA polymerase this consensus sequence, although irrelevant for the 3'----5' exonuclease activity, is essential for initiation and elongation. Based on these results and on its homology with known or putative metal-binding amino acid sequences, we propose that in phi 29 DNA polymerase the Tyr-Gly-Asp-Thr-Asp-Ser consensus motif is part of the dNTP binding site, involved in the synthetic activities of the polymerase (i.e., initiation and polymerization), and that it is involved particularly in the metal binding associated with the dNTP site. Images PMID:2191296

  12. Prediction of posttranslational modification sites from amino acid sequences with kernel methods.

    PubMed

    Xu, Yan; Wang, Xiaobo; Wang, Yongcui; Tian, Yingjie; Shao, Xiaojian; Wu, Ling-Yun; Deng, Naiyang

    2014-03-01

    Post-translational modification (PTM) is the chemical modification of a protein after its translation and one of the later steps in protein biosynthesis for many proteins. It plays an important role which modifies the end product of gene expression and contributes to biological processes and diseased conditions. However, the experimental methods for identifying PTM sites are both costly and time-consuming. Hence computational methods are highly desired. In this work, a novel encoding method PSPM (position-specific propensity matrices) is developed. Then a support vector machine (SVM) with the kernel matrix computed by PSPM is applied to predict the PTM sites. The experimental results indicate that the performance of new method is better or comparable with the existing methods. Therefore, the new method is a useful computational resource for the identification of PTM sites. A unified standalone software PTMPred is developed. It can be used to predict all types of PTM sites if the user provides the training datasets. The software can be freely downloaded from http://www.aporc.org/doc/wiki/PTMPred. PMID:24291233

  13. Thioacidolysis Marker Compound for Ferulic Acid Incorporation into Angiosperm Lignins (and an Indicator for Cinnamoyl-coenzyme-A Reductase Deficiency

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A molecular marker compound, derived from lignin by the thioacidolysis degradative method, for structures produced when ferulic acid is incorporated into lignification in angiosperms (poplar, Arabidopsis, tobacco) has been structurally identified as 1,2,2-trithioethyl ethylguaiacol [1-(4-hydroxy-3-m...

  14. Spectrophotometric Determination of the Dissociation Constant of an Acid-Base Indicator Using a Mathematical Deconvolution Technique

    ERIC Educational Resources Information Center

    Alter, Krystyn P.; Molloy, John L.; Niemeyer, Emily D.

    2005-01-01

    A laboratory experiment reinforces the concept of acid-base equilibria while introducing a common application of spectrophotometry and can easily be completed within a standard four-hour laboratory period. It provides students with an opportunity to use advanced data analysis techniques like data smoothing and spectral deconvolution to…

  15. Amino acid sequence and properties of vasopressin-associated elephant neurophysin.

    PubMed

    Huang, H B; Wellner, D; Naudé, R; Oelofsen, W; Oosthuizen, M M; Breslow, E

    1994-09-01

    The primary structure of an elephant neurophysin, homologous to vasopressin-associated neurophysins, is reported. The protein contains a Tyr for Asn substitution at position 75, a position in direct contact with residues 77 and 78 of the monomer-monomer interface. This Tyr residue therefore serves as a potential reporter of the path involved in the long-range linkage between peptide binding and dimerization in this system. NMR studies of the protein in unliganded and liganded states demonstrated normal dimerization properties and the expected increase in dimerization associated with binding peptide. In keeping with an elevated pKa of 11.1 assigned to Tyr-75 by UV spectrophotometric titration, the NMR signals from the 3,5 and 2,6 ring protons of Tyr-75 were shifted 0.3 and 0.2 ppm upfield, respectively, relative to their positions in small peptides, indicating significant shielding and/or hydrogen bonding. The Tyr-75 ring proton signals narrowed slightly, with no discernible change in chemical shift, on conversion from dimer to monomer in the unliganded state. Ring protons of Tyr-49, distant from the monomer-monomer interface, but adjacent to the peptide-binding site, were markedly perturbed by dimerization, in accord with their behavior in bovine neurophysins. The results suggest that the secondary and tertiary structure of the region 75-78 is largely unchanged by dimerization, and argue against an important role for this region in dimerization-mediated conformational changes that alter the binding site in the unliganded state.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7822104

  16. Microbial profiling of South African acid mine water samples using next generation sequencing platform.

    PubMed

    Kamika, I; Azizi, S; Tekere, M

    2016-07-01

    This study monitored changes in bacterial and fungal structure in a mine water in a monthly basis over 4 months. Over the 4-month study period, mine water samples contained more bacteria (91.06 %) compared to fungi (8.94 %). For bacteria, mine water samples were dominated by Proteobacteria (39.14 to 65.06 %) followed by Firmicutes (26.34 to 28.9 %) in summer, and Cyanobacteria (27.05 %) in winter. In the collected samples, 18 % of bacteria could not be assigned to a phylum and remained unclassified suggesting hitherto vast untapped microbial diversity especially during winter. The fungal domain was the sole eukaryotic microorganism found in the mine water samples with unclassified fungi (68.2 to 91 %) as the predominant group, followed by Basidiomycota (6.9 to 27.8 %). The time of collection, which was linked to the weather, had higher impact on bacterial community than fungal community. The bacterial operational taxonomic units (OTUs) ranged from 865 to 4052 over the 4-month sampling period, while fungal OTUs varied from 73 to 249. The diversity indices suggested that the bacterial community inhabiting the mine water samples were more diverse than the fungal community. The canonical correspondence analysis (CCA) results highlighted that the bacterial community variance had the strongest relationship with water temperature, conductivity, pH, and dissolved oxygen (DO) content, as compared to fungi and water characteristics, had the greatest contribution to both bacterial and fungal community variance. The results provided the relationships between microbial community and environmental variables in the studied mining sites. PMID:26980100

  17. Predicting Secretory Proteins of Malaria Parasite by Incorporating Sequence Evolution Information into Pseudo Amino Acid Composition via Grey System Model

    PubMed Central

    Lin, Wei-Zhong; Fang, Jian-An; Xiao, Xuan; Chou, Kuo-Chen

    2012-01-01

    The malaria disease has become a cause of poverty and a major hindrance to economic development. The culprit of the disease is the parasite, which secretes an array of proteins within the host erythrocyte to facilitate its own survival. Accordingly, the secretory proteins of malaria parasite have become a logical target for drug design against malaria. Unfortunately, with the increasing resistance to the drugs thus developed, the situation has become more complicated. To cope with the drug resistance problem, one strategy is to timely identify the secreted proteins by malaria parasite, which can serve as potential drug targets. However, it is both expensive and time-consuming to identify the secretory proteins of malaria parasite by experiments alone. To expedite the process for developing effective drugs against malaria, a computational predictor called “iSMP-Grey” was developed that can be used to identify the secretory proteins of malaria parasite based on the protein sequence information alone. During the prediction process a protein sample was formulated with a 60D (dimensional) feature vector formed by incorporating the sequence evolution information into the general form of PseAAC (pseudo amino acid composition) via a grey system model, which is particularly useful for solving complicated problems that are lack of sufficient information or need to process uncertain information. It was observed by the jackknife test that iSMP-Grey achieved an overall success rate of 94.8%, remarkably higher than those by the existing predictors in this area. As a user-friendly web-server, iSMP-Grey is freely accessible to the public at http://www.jci-bioinfo.cn/iSMP-Grey. Moreover, for the convenience of most experimental scientists, a step-by-step guide is provided on how to use the web-server to get the desired results without the need to follow the complicated mathematical equations involved in this paper. PMID:23189138

  18. Next-generation re-sequencing of genes involved in increased platelet reactivity in diabetic patients on acetylsalicylic acid.

    PubMed

    Postula, Marek; Janicki, Piotr K; Eyileten, Ceren; Rosiak, Marek; Kaplon-Cieslicka, Agnieszka; Sugino, Shigekazu; Wilimski, Radosław; Kosior, Dariusz A; Opolski, Grzegorz; Filipiak, Krzysztof J; Mirowska-Guzel, Dagmara

    2016-06-01

    The objective of this study was to investigate whether rare missense genetic variants in several genes related to platelet functions and acetylsalicylic acid (ASA) response are associated with the platelet reactivity in patients with diabetes type 2 (T2D) on ASA therapy. Fifty eight exons and corresponding introns of eight selected genes, including PTGS1, PTGS2, TXBAS1, PTGIS, ADRA2A, ADRA2B, TXBA2R, and P2RY1 were re-sequenced in 230 DNA samples from T2D patients by using a pooled PCR amplification and next-generation sequencing by Illumina HiSeq2000. The observed non-synonymous variants were confirmed by individual genotyping of 384 DNA samples comprising of the individuals from the original discovery pools and additional verification cohort of 154 ASA-treated T2DM patients. The association between investigated phenotypes (ASA induced changes in platelets reactivity by PFA-100, VerifyNow and serum thromboxane B2 level [sTxB2]), and accumulation of rare missense variants (genetic burden) in investigated genes was tested using statistical collapsing tests. We identified a total of 35 exonic variants, including 3 common missense variants, 15 rare missense variants, and 17 synonymous variants in 8 investigated genes. The rare missense variants exhibited statistically significant difference in the accumulation pattern between a group of patients with increased and normal platelet reactivity based on PFA-100 assay. Our study suggests that genetic burden of the rare functional variants in eight genes may contribute to differences in the platelet reactivity measured with the PFA-100 assay in the T2DM patients treated with ASA. PMID:26599574

  19. Purification and amino acid sequence of a highly insecticidal toxin from the venom of the brazilian spider Phoneutria nigriventer which inhibits NMDA-evoked currents in rat hippocampal neurones.

    PubMed

    de Figueiredo, S G; de Lima, M E; Nascimento Cordeiro, M; Diniz, C R; Patten, D; Halliwell, R F; Gilroy, J; Richardson, M

    2001-01-01

    A new insecticidal toxin Tx4(5-5) was isolated from the fraction PhTx4 of the venom of the spider Phoneutria nigriventer by reverse phase high performance liquid chromatography (HPLC) and anion exchange HPLC. The complete amino acid sequence determined by automated Edman degradation showed that Tx4(5-5) is a single chain polypeptide composed of 47 amino acid residues, including 10 cysteines, with a calculated molecular mass of 5175 Da. Tx4(5-5) shows 64% of sequence identity with Tx4(6-1), another insecticidal toxin from the same venom. Tx4(5-5) was highly toxic to house fly (Musca domestica), cockroach (Periplaneta americana) and cricket (Acheta domesticus ), producing neurotoxic effects (knock-down, trembling with uncoordinated movements) at doses as low as 50 ng/g (house fly), 250 ng/g (cockroach) and 150 ng/g (cricket). In contrast, intracerebroventricular injections (30 microg) into mice induced no behavioural effects. Preliminary electrophysiological studies carried out on whole-cell voltage-clamped rat hippocampal neurones indicated that Tx4(5-5) (at 1 microM) reversibly inhibited the N-methyl-D-aspartate-subtype of ionotropic glutamate receptor, while having little or no effect on kainate-, alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid- or gamma-aminobutyric acid-activated currents. PMID:10978749

  20. Peptides Composed of Alternating L- and D-Amino Acids Inhibit Amyloidogenesis in Three Distinct Amyloid Systems Independent of Sequence.

    PubMed

    Kellock, Jackson; Hopping, Gene; Caughey, Byron; Daggett, Valerie

    2016-06-01

    There is now substantial evidence that soluble oligomers are primary toxic agents in amyloid diseases. The development of an antibody recognizing the toxic soluble oligomeric forms of different and unrelated amyloid species suggests a common conformational intermediate during amyloidogenesis. We previously observed a common occurrence of a novel secondary structure element, which we call α-sheet, in molecular dynamics (MD) simulations of various amyloidogenic proteins, and we hypothesized that the toxic conformer is composed of α-sheet structure. As such, α-sheet may represent a conformational signature of the misfolded intermediates of amyloidogenesis and a potential unique binding target for peptide inhibitors. Recently, we reported the design and characterization of a novel hairpin peptide (α1 or AP90) that adopts stable α-sheet structure and inhibits the aggregation of the β-Amyloid Peptide Aβ42 and transthyretin. AP90 is a 23-residue hairpin peptide featuring alternating D- and L-amino acids with favorable conformational propensities for α-sheet formation, and a designed turn. For this study, we reverse engineered AP90 to identify which of its design features is most responsible for conferring α-sheet stability and inhibitory activity. We present experimental characterization (CD and FTIR) of seven peptides designed to accomplish this. In addition, we measured their ability to inhibit aggregation in three unrelated amyloid species: Aβ42, transthyretin, and human islet amylin polypeptide. We found that a hairpin peptide featuring alternating L- and D-amino acids, independent of sequence, is sufficient for conferring α-sheet structure and inhibition of aggregation. Additionally, we show a correlation between α-sheet structural stability and inhibitory activity. PMID:27012425

  1. The delta EEG (sleep)-inducing peptide (DSIP). XI. Amino-acid analysis, sequence, synthesis and activity of the nonapeptide.

    PubMed

    Schoenenberger, G A; Maier, P F; Tobler, H J; Wilson, K; Monnier, M

    1978-09-01

    A peptide which induces slow-wave EEG (sleep) after intraventricular infusion into the brain has been isolated from the extracorporeal dialysate of cerebral venous blood in rabbits submitted to hypnogenic electrical stimulation of the intralaminar thalamic area. It was shown by amino-acid analysis and sequence determination to be Trp-Ala-Gly-Gly-Asp-Ala-Ser-Gly-Glu and named "Delta Sleep-Inducing Peptide" (DSIP). This compound was synthesized as well as 5 possible metabolic products (1--8, 2--9, 2--8, 1--4 and 5--9), 2 nonapeptide analogues (with one and two amino-acids exchanged) and a related tripeptide (Trp-Ser-Glu). All 9 synthetic peptides were infused intraventricularly in rabbits (6 nmol/kg in 0.05 ml of CSF-like solution over 3.5 min) and tested under double-blind conditions. A total of 61 rabbits including controls were used. The EEG from the frontal neocortex and the limbic archicortex were subjected to direct fast-Fourier transformation and analyzed by an 1108 computer system. A highly specific delta and spindle EEG-enhancing effect of the synthetic DSIP could be demonstrated. The mean increase of EEG delta activity reached 35% in the neocortex and limbic cortex as compared to control animals receiving CSF-like solution or any of the other 8 peptides. The final chemical characterization of the synthetic DSIP revealed that only the pure alpha-aspartyl peptide is highly active in contrast to its beta-Asp isomer. A neurohumoral modulating and programming activity was suggested. PMID:568769

  2. Species specific identification of spore-producing microbes using the gene sequence of small acid-soluble spore coat proteins for amplification based diagnostics

    DOEpatents

    McKinney, Nancy

    2002-01-01

    PCR (polymerase chain reaction) primers for the detection of certain Bacillus species, such as Bacillus anthracis. The primers specifically amplify only DNA found in the target species and can distinguish closely related species. Species-specific PCR primers for Bacillus anthracis, Bacillus globigii and Clostridium perfringens are disclosed. The primers are directed to unique sequences within sasp (small acid soluble protein) genes.

  3. Draft Genome Sequences of Salmonella enterica subsp. enterica Serovar Berta ATCC 8392 and a Nalidixic Acid-Resistant Isolate of This Strain

    PubMed Central

    Cooper, Ashley; Koziol, Adam G.; Carrillo, Catherine D.

    2016-01-01

    Salmonella enterica subspecies enterica serovar Berta has been isolated in multiple animal species and has been implicated in human disease. Here, we report a 4.7-Mbp draft genome sequence of S. enterica serovar Berta (ATCC strain 8392) and a nalidixic acid-resistant isolate derived from this strain. PMID:27103707

  4. COMPARISON OF PHYLOGENETIC RELATIONSHIPS BASED ON PHOSPHOLIPID FATTY ACID PROFILES AND RIBOSOMAL RNA SEQUENCE SIMILARITIES AMONG DISSIMILATORY SULFATE-REDUCING BACTERIA

    EPA Science Inventory

    Twenty-five isolates of dissimilatory sulfate-reducing bacteria were clustered based on similarity analysis of their phospholipid ester-linked fatty acids (PLFA). f these, twenty-three showed the phylogenetic relationships based on the sequence similarity of their 16S rRNA direct...

  5. Single-spore elemental analyses indicate that dipicolinic acid-deficient Bacillus subtilis spores fail to accumulate calcium.

    PubMed

    Hintze, Paul E; Nicholson, Wayne L

    2010-06-01

    Dipicolinic acid (pyridine-2,6-carboxylic acid; DPA) is a major component of bacterial spores and has been shown to be an important determinant of spore resistance. In the core of dormant Bacillus subtilis spores, DPA is associated with divalent calcium in a 1:1 chelate (Ca-DPA). Spores excrete Ca-DPA during germination, but it is unknown whether Ca and DPA are imported separately or together into the developing spore. Elemental analysis by scanning electron microscopy-energy-dispersive X-ray spectroscopy (SEM-EDS) of wild-type spores and mutant spores lacking the ability to synthesize DPA showed that DPA-less spores also lacked calcium, suggesting that the two compounds may be co-imported. PMID:20396869

  6. Effect of tocopherol and acetylsalicylic acid on the biochemical indices of blood in dioxin-exposed rats.

    PubMed

    Rosińczuk, Joanna; Całkosiński, Ireneusz

    2015-07-01

    New sources of dioxins and increased dioxin concentrations in the environment, coupled with their increased bioavailability along the food chain and accumulation in adipose tissues, contribute to various adverse long-term biological effects. The purpose of the study was to determine whether tocopherol protects the CNS by decreasing the pro-inflammatory influence of free radicals generated by TCDD; whether acetylsalicylic acid inhibits the production of inflammatory mediators; and whether the combined administration of tocopherol and acetylsalicylic acid to TCDD-exposed rats has a potential CNS-protective effect. The study included 117 rats divided into 8 groups: 75 female and 12 male Buffalo rats aged 8-10 weeks, weighing 140-160 g; as well as 30 female rats aged 6 weeks and weighing 120 g, which were the offspring of females from each study group. In the experiment, the following substances were used: 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), dosed at 5 μg/kg BW and 12.5 μg/kg BW, diluted in a 1% DMSO solution at the concentration of 1 μg/ml; α-tocopherol acetate, dosed at 30 mg/kg BW, in 0.2 ml of oil solution; and acetylsalicylic acid, 50mg/kg BW, suspended in 0.5 ml of starch solution, administered orally using a feeding tube. Pleurisy was induced by an injection of 0.15 ml of 1% carrageenin solution. The use of tocopherol reduces the adverse effects of the inflammatory reaction induced by TCDD. Administering tocopherol improves protein metabolism by reducing protein catabolism, and raises γ-globulin fraction levels. Combined acetylsalicylic acid and tocopherol suppress catabolic processes accompanying inflammation. PMID:26056971

  7. Elemental stoichiometry indicates predominant influence of potassium and phosphorus limitation on arbuscular mycorrhizal symbiosis in acidic soil at high altitude.

    PubMed

    Khan, Mohammad Haneef; Meghvansi, Mukesh K; Gupta, Rajeev; Veer, Vijay

    2015-09-15

    The functioning of high-altitude agro-ecosystems is constrained by the harsh environmental conditions, such as low temperatures, acidic soil, and low nutrient supply. It is therefore imperative to investigate the site-specific ecological stoichiometry with respect to AM symbiosis in order to maximize the arbuscular mycorrhizal (AM) benefits for the plants in such ecosystems. Here, we assess the elemental stoichiometry of four Capsicum genotypes grown on acidic soil at high altitude in Arunachal Pradesh, India. Further, we try to identify the predominant resource limitations influencing the symbioses of different Capsicum genotypes with the AM fungi. Foliar and soil elemental stoichiometric relations of Capsicum genotypes were evaluated with arbuscular mycorrhizal (AM) colonization and occurrence under field conditions. AM fungal diversity in rhizosphere, was estimated through PCR-DGGE profiling. Results demonstrated that the symbiotic interaction of various Capsicum genotypes with the AM fungi in acidic soil was not prominent in the study site as evident from the low range of root colonization (21-43.67%). In addition, despite the rich availability of carbon in plant leaves as well as in soil, the carbon-for-phosphorus trade between AMF and plants appeared to be limited. Our results provide strong evidences of predominant influence of the potassium-limitation, in addition to phosphorus-limitation, on AM symbiosis with Capsicum in acidic soil at high altitude. We also conclude that the potassium should be considered in addition to carbon, nitrogen, and phosphorus in further studies investigating the stoichiometric relationships with the AMF symbioses in high altitude agro-ecosystems. PMID:26555273

  8. Seasonal variation of Fatty acids and stable carbon isotopes in sponges as indicators for nutrition: biomarkers in sponges identified.

    PubMed

    Koopmans, Marieke; van Rijswijk, Pieter; Boschker, Henricus T S; Marco, Houtekamer; Martens, Dirk; Wijffels, Rene H

    2015-02-01

    To get a better understanding of sponge feeding biology and efficiencies, the fatty acid (FA) composition and (13)C natural abundance of sponges and of suspended particulate matter (SPM) from surrounding seawater was studied in different seasons at three locations. Haliclona oculata and Haliclona xena from the Oosterschelde, the Netherlands, Halichondria panicea and H. xena from Lake Veere, the Netherlands, and Aplysina aerophoba and Dysidea avara from the Mediterranean, Spain, were studied. Several FA biomarkers for different algal groups, bacteria and sponge biomass were identified in all sponges. The FA concentration variation in sponges was related to changes in fatty acid concentration in SPM. Stable carbon isotopic ratios (δ(13)C) in sponge specific FAs showed very limited seasonal variation at all sites. Algal FAs in sponges were mainly acquired from the SPM through active filtration in all seasons. At the two sites in the Netherlands only in May (spring), the sponge specific FAs had similar δ(13)C ratios as algal FAs, suggesting that sponges were mainly growing during spring and probably summer. During autumn and winter, they were still actively filtering, but the food collected during this period had little effect on sponge δ(13)C values suggesting limited incorporation of filtered material into the sponge body. The sponge A. aerophoba relied mostly on the symbiotic bacteria. In conclusion, fatty acid composition in combination with stable carbon isotope analysis can be used to analyze the food source of sponges. PMID:25107690

  9. Purification and N-terminal amino acid sequence of a chondroitin sulphate/dermatan sulphate proteoglycan isolated from intima/media preparations of human aorta.

    PubMed

    Stöcker, G; Meyer, H E; Wagener, C; Greiling, H

    1991-03-01

    A proteoglycan (PG) was purified to homogeneity from intima/media preparations of human aorta specimens by the following chromatographic steps: Sepharose Q anion exchange, Sepharose CL-4B size exclusion, hydroxyapatite, MonoQ anion exchange and TSK G 4000 SW size exclusion. The purity of the preparation was established by SDS/PAGE using direct staining by silver or Dimethylmethylene Blue, as well as by Western blots of biotin-labelled samples. The electrophoretic mobility of the native PG was less than that of a 200,000-Mr standard protein. After treatment with chondroitin sulphate lyase ABC, a core protein of Mr 15,000 was revealed. The Mr of the glycosaminoglycan (GAG) peptides was less than 24,000, by comparison with a keratan sulphate peptide. The composition of the GAG chains was determined by differential digestion of the PG by chondroitin sulphate lyases AC/ABC or chondroitin sulphate lyase AC alone followed by anion-exchange chromatography of the resulting disaccharides. The GAG chains are composed of approximately one-third of dermatan sulphate and two-thirds chondroitin sulphate disaccharide units. The sequence of the 20 N-terminal amino acids is identical with the sequence previously reported for PG I isolated from human developing bone [Fisher, Termine & Young (1989) J. Biol. Chem. 264, 4571-4576]. The assignment of glycosylation sites to the serine residues in positions 5 and 10 was confirmed. The findings indicate that the chondroitin sulphate/dermatan sulphate PG is a major PG in intima/media preparations of human aorta and represents a biglycan-type PG. PMID:1848758

  10. RoboOligo: software for mass spectrometry data to support manual and de novo sequencing of post-transcriptionally modified ribonucleic acids

    PubMed Central

    Sample, Paul J.; Gaston, Kirk W.; Alfonzo, Juan D.; Limbach, Patrick A.

    2015-01-01

    Ribosomal ribonucleic acid (RNA), transfer RNA and other biological or synthetic RNA polymers can contain nucleotides that have been modified by the addition of chemical groups. Traditional Sanger sequencing methods cannot establish the chemical nature and sequence of these modified-nucleotide containing oligomers. Mass spectrometry (MS) has become the conventional approach for determining the nucleotide composition, modification status and sequence of modified RNAs. Modified RNAs are analyzed by MS using collision-induced dissociation tandem mass spectrometry (CID MS/MS), which produces a complex dataset of oligomeric fragments that must be interpreted to identify and place modified nucleosides within the RNA sequence. Here we report the development of RoboOligo, an interactive software program for the robust analysis of data generated by CID MS/MS of RNA oligomers. There are three main functions of RoboOligo: (i) automated de novo sequencing via the local search paradigm. (ii) Manual sequencing with real-time spectrum labeling and cumulative intensity scoring. (iii) A hybrid approach, coined ‘variable sequencing’, which combines the user intuition of manual sequencing with the high-throughput sampling of automated de novo sequencing. PMID:25820423

  11. Nucleic acid sequence-based amplification assays for rapid detection of West Nile and St. Louis encephalitis viruses.

    PubMed

    Lanciotti, R S; Kerst, A J

    2001-12-01

    The development and application of nucleic acid sequence-based amplification (NASBA) assays for the detection of West Nile (WN) and St. Louis encephalitis (SLE) viruses are reported. Two unique detection formats were developed for the NASBA assays: a postamplification detection step with a virus-specific internal capture probe and electrochemiluminescence (NASBA-ECL assay) and a real-time assay with 6-carboxyfluorescein-labeled virus-specific molecular beacon probes (NASBA-beacon assay). The sensitivities and specificities of these NASBA assays were compared to those of a newly described standard reverse transcription (RT)-PCR and TaqMan assays for SLE virus and to a previously published TaqMan assay for WN virus. The NASBA assays demonstrated exceptional sensitivities and specificities compared to those of virus isolation, the TaqMan assays, and standard RT-PCR, with the NASBA-beacon assay yielding results in less than 1 h. These assays should be of utility in the diagnostic laboratory to complement existing diagnostic testing methodologies and as a tool in conducting flavivirus surveillance in the United States. PMID:11724870

  12. Purification and complete amino acid sequence of a new type of sweet protein taste-modifying activity, curculin.

    PubMed

    Yamashita, H; Theerasilp, S; Aiuchi, T; Nakaya, K; Nakamura, Y; Kurihara, Y

    1990-09-15

    A new taste-modifying protein named curculin was extracted with 0.5 M NaCl from the fruits of Curculigo latifolia and purified by ammonium sulfate fractionation, CM-Sepharose ion-exchange chromatography, and gel filtration. Purified curculin thus obtained gave a single band having a Mr of 12,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the presence of 8 M urea. The molecular weight determined by low-angle laser light scattering was 27,800. These results suggest that native curculin is a dimer of a 12,000-Da polypeptide. The complete amino acid sequence of curculin was determined by automatic Edman degradation. Curculin consists of 114 residues. Curculin itself elicits a sweet taste. After curculin, water elicits a sweet taste, and sour substances induce a stronger sense of sweetness. No protein with both sweet-tasting and taste-modifying activities has ever been found. There are five sets of tripeptides common to miraculin (a taste-modifying protein), six sets of tripeptides common to thaumatin (a sweet protein), and two sets of tripeptides common to monellin (a sweet protein). Anti-miraculin serum was not immunologically reactive with curculin. The mechanism of the taste-modifying action of curculin is discussed. PMID:2394746

  13. Sequencing around 5-Hydroxyconiferyl Alcohol-Derived Units in Caffeic Acid O-Methyltransferase-Deficient Poplar Lignins1[OA

    PubMed Central

    Lu, Fachuang; Marita, Jane M.; Lapierre, Catherine; Jouanin, Lise; Morreel, Kris; Boerjan, Wout; Ralph, John

    2010-01-01

    Caffeic acid O-methyltransferase (COMT) is a bifunctional enzyme that methylates the 5- and 3-hydroxyl positions on the aromatic ring of monolignol precursors, with a preference for 5-hydroxyconiferaldehyde, on the way to producing sinapyl alcohol. Lignins in COMT-deficient plants contain benzodioxane substructures due to the incorporation of 5-hydroxyconiferyl alcohol (5-OH-CA), as a monomer, into the lignin polymer. The derivatization followed by reductive cleavage method can be used to detect and determine benzodioxane structures because of their total survival under this degradation method. Moreover, partial sequencing information for 5-OH-CA incorporation into lignin can be derived from detection or isolation and structural analysis of the resulting benzodioxane products. Results from a modified derivatization followed by reductive cleavage analysis of COMT-deficient lignins provide evidence that 5-OH-CA cross couples (at its β-position) with syringyl and guaiacyl units (at their O-4-positions) in the growing lignin polymer and then either coniferyl or sinapyl alcohol, or another 5-hydroxyconiferyl monomer, adds to the resulting 5-hydroxyguaiacyl terminus, producing the benzodioxane. This new terminus may also become etherified by coupling with further monolignols, incorporating the 5-OH-CA integrally into the lignin structure. PMID:20427467

  14. Sequencing around 5-hydroxyconiferyl alcohol-derived units in caffeic acid O-methyltransferase-deficient poplar lignins.

    PubMed

    Lu, Fachuang; Marita, Jane M; Lapierre, Catherine; Jouanin, Lise; Morreel, Kris; Boerjan, Wout; Ralph, John

    2010-06-01

    Caffeic acid O-methyltransferase (COMT) is a bifunctional enzyme that methylates the 5- and 3-hydroxyl positions on the aromatic ring of monolignol precursors, with a preference for 5-hydroxyconiferaldehyde, on the way to producing sinapyl alcohol. Lignins in COMT-deficient plants contain benzodioxane substructures due to the incorporation of 5-hydroxyconiferyl alcohol (5-OH-CA), as a monomer, into the lignin polymer. The derivatization followed by reductive cleavage method can be used to detect and determine benzodioxane structures because of their total survival under this degradation method. Moreover, partial sequencing information for 5-OH-CA incorporation into lignin can be derived from detection or isolation and structural analysis of the resulting benzodioxane products. Results from a modified derivatization followed by reductive cleavage analysis of COMT-deficient lignins provide evidence that 5-OH-CA cross couples (at its beta-position) with syringyl and guaiacyl units (at their O-4-positions) in the growing lignin polymer and then either coniferyl or sinapyl alcohol, or another 5-hydroxyconiferyl monomer, adds to the resulting 5-hydroxyguaiacyl terminus, producing the benzodioxane. This new terminus may also become etherified by coupling with further monolignols, incorporating the 5-OH-CA integrally into the lignin structure. PMID:20427467

  15. Transposition of a plasmid deoxyribonucleic acid sequence that mediates ampicillin resistance: independence from host rec functions and orientation of insertion.

    PubMed Central

    Rubens, C; Heffron, F; Falkow, S

    1976-01-01

    Insertion of the transposable deoxyribonucleic acid sequence that specifies the TEM beta-lactamase (TnA) occurred in at least 19 sites on the 5.5 x 10(6)-dalton plasmid RSF1010. There was no significant difference in the frequency of transposition or in the distribution of TnA insertion sites for recombinant plasmids isolated from recombination-proficient (rec+) or recombination-deficient (rec-) bacterial host cells. The site and orientation of TnA insertions were determined by both heteroduplex analysis and enzymatic digestion with restriction endonucleases. Insertion in the gene encoding for sulfonamide resistance occurred without circular permutation in one or the other of two distinct orientations. Insertions in orientation P were strongly polar on distal gene expression, whereas insertions in orientation M were mutagenic but not polar. In addition, we have observed that TnA elements from different R plasmids show fine structural heterogeneity, and that TnA insertion at a site adjacent to the origin of replication causes an increase in plasmid copy number. Images PMID:789346

  16. Detection of Aspergillus fumigatus in a rat model of invasive pulmonary aspergillosis by real-time nucleic acid sequence-based amplification.

    PubMed

    Zhao, Yanan; Park, Steven; Warn, Peter; Shrief, Raghdaa; Harrison, Elizabeth; Perlin, David S

    2010-04-01

    Rapid and sensitive detection of Aspergillus from clinical samples may facilitate the early diagnosis of invasive pulmonary aspergillosis (IPA). A real-time nucleic acid sequence-based amplification (NASBA) method was investigated by use of an inhalational rat model of IPA. Immunosuppressed male Sprague-Dawley rats were exposed to Aspergillus fumigatus spores for an hour in an aerosol chamber. Bronchoalveolar lavage (BAL) fluid, lung tissues, and whole blood were collected from five infected rats at 1, 24, 48, 72, and 96 h postinfection and five uninfected rats at the end of the experiment. Total nucleic acid (TNA) was extracted on an easyMAG instrument. A primer-molecular beacon set targeting 28S rRNA was designed to detect Aspergillus spp. The results were compared to those of quantitative PCR (qPCR) (18S rDNA) and quantitative culture. The analytical sensitivity of the real-time NASBA assay was <1 CFU/assay. A linear range of detection was demonstrated over 5 log units of conidia (10 to 10(5) spores). Both NASBA and qPCR showed a progressive increase in lung tissue burdens, while the CFU counts were stable over time. The fungal burdens in BAL fluid were more variable and not indicative of a progressive infection. The results of both real-time assays correlated well for both sample types (r = 0.869 and P < 0.0001 for lung tissue, r = 0.887 and P < 0.0001 for BAL fluid). For all whole-blood specimens, NASBA identified Aspergillus-positive samples in the group from which samples were collected at 72 h postinfection (three of five samples) and the group from which samples were collected at 96 h postinfection (five of five samples), but no positive results were obtained by culture or PCR. Real-time NASBA is highly sensitive and useful for the detection of Aspergillus in an experimental model of IPA. PMID:20129972

  17. The phylogeny of acetic acid bacteria based on the partial sequences of 16S ribosomal RNA: the elevation of the subgenus Gluconoacetobacter to the generic level.

    PubMed

    Yamada, Y; Hoshino, K; Ishikawa, T

    1997-08-01

    Thirty-six strains of acetic acid bacteria classified in the genera Acetobacter, Gluconobacter, and Acidomonas were examined for their partial base sequences in positions 1220 through 1375, 156 bases, of 16S rRNA. The strains of the Q10-equipped Gluconobacter species examined were divided into two subgroups, which included the type strains of Gluconobacter oxydans, the type species of the genus Gluconobacter, and of a second species, Gluconobacter cerinus, respectively. The base differences numbered four between the two type strains. The strains of the Q9-equipped species examined classified in the type subgenus Acetobacter of the genus Acetobacter were not very distant phylogenetically from those of the genus Gluconobacter. The calculated number of base differences was 9-6 between the type strains of G. oxydans and G. cerinus and the type strains of Acetobacter aceti and Acetobacter pasteurianus. In contrast, the strains of the Q10-equipped species examined classified in the subgenus Gluconoacetobacter of the genus Acetobacter were very distant phylogenetically from those of the Acetobacter and Gluconobacter species mentioned above. The number of base differences was calculated to be 14-8. Furthermore, the strains of the methanol-assimilating, Q10-equipped species of the genus Acidomonas examined were located in phylogenetically isolated positions. The type strain of Acidomonas methanolica (identical to Acetobacter methanolicus), the type species of the genus Acidomonas, had 16-9 base differences. The data obtained here indicated that the members of the subgenus Gluconoacetobacter of the genus Acetobacter can be distinguished at the generic level. The new genus Gluconoacetobacter was proposed with the type species, Gluconoacetobacter liquefaciens, in recognition of the genus Acidomonas along with the genera Acetobacter and Gluconobacter in the classification of the acetic acid bacteria. PMID:9301103

  18. Amino acid sequence of the AhR1 ligand-binding domain predicts avian sensitivity to dioxin like compounds: in vivo verification in European starlings.

    PubMed

    Eng, Margaret L; Elliott, John E; Jones, Stephanie P; Williams, Tony D; Drouillard, Ken G; Kennedy, Sean W

    2014-12-01

    Research has demonstrated that the sensitivity of avian species to the embyrotoxic effects of dioxin-like compounds can be predicted by the amino acid identities at two key sites within the ligand-binding domain of the aryl hydrocarbon receptor 1 (AhR1). The domestic chicken (Gallus gallus domesticus) has been established as a highly sensitive species to the toxic effects of dioxin-like compounds. Results from genotyping and in vitro assays predict that the European starling (Sturnus vulgaris) is also highly sensitive to dioxin-like compound toxicity. The objective of the present study was to test that prediction in vivo. To do this, we used egg injections in field nesting starlings with 3,3',4,4',5-pentachlorobiphenyl (PCB-126), a dioxin-like polychlorinated biphenyl. Eggs were dosed with either the vehicle control or 1 of 5 doses (1.4, 7.1, 15.9, 32.1, and 52.9 ng PCB-126/g egg). A dose-dependent increase in embryo mortality occurred, and the median lethal dose (LD50; 95% confidence interval [CI]) was 5.61 (2.33-9.08) ng/g. Hepatic CYP1A4/5 messenger RNA (mRNA) expression in hatchlings also increased in a dose-dependent manner, with CYP1A4 being more induced than CYP1A5. No effect of dose on morphological measures was seen, and we did not observe any overt malformations. These results indicate that, other than the chicken, the European starling is the most sensitive species to the effects of PCB-126 on avian embryo mortality reported to date, which supports the prediction of relative sensitivity to dioxin-like compounds based on amino acid sequence of the AhR1. PMID:25209921

  19. PhTX-II a Basic Myotoxic Phospholipase A2 from Porthidium hyoprora Snake Venom, Pharmacological Characterization and Amino Acid Sequence by Mass Spectrometry

    PubMed Central

    Huancahuire-Vega, Salomón; Ponce-Soto, Luis Alberto; Marangoni, Sergio

    2014-01-01

    A monomeric basic PLA2 (PhTX-II) of 14149.08 Da molecular weight was purified to homogeneity from Porthidium hyoprora venom. Amino acid sequence by in tandem mass spectrometry revealed that PhTX-II belongs to Asp49 PLA2 enzyme class and displays conserved domains as the catalytic network, Ca2+-binding loop and the hydrophobic channel of access to the catalytic site, reflected in the high catalytic activity displayed by the enzyme. Moreover, PhTX-II PLA2 showed an allosteric behavior and its enzymatic activity was dependent on Ca2+. Examination of PhTX-II PLA2 by CD spectroscopy indicated a high content of alpha-helical structures, similar to the known structure of secreted phospholipase IIA group suggesting a similar folding. PhTX-II PLA2 causes neuromuscular blockade in avian neuromuscular preparations with a significant direct action on skeletal muscle function, as well as, induced local edema and myotoxicity, in mice. The treatment of PhTX-II by BPB resulted in complete loss of their catalytic activity that was accompanied by loss of their edematogenic effect. On the other hand, enzymatic activity of PhTX-II contributes to this neuromuscular blockade and local myotoxicity is dependent not only on enzymatic activity. These results show that PhTX-II is a myotoxic Asp49 PLA2 that contributes with toxic actions caused by P. hyoprora venom. PMID:25365526

  20. Nucleotide sequences and characterization of liv genes encoding components of the high-affinity branched-chain amino acid transport system in Salmonella typhimurium.

    PubMed

    Matsubara, K; Ohnishi, K; Kiritani, K

    1992-07-01

    A 7.6-kb fragment of Salmonella typhimurium LT2 containing the liv gene cluster, which specifies the high-affinity branched-chain amino acid transport system (LIV-I), has been isolated. The upstream region contains the livB and livC genes encoding the leucine-isoleucine-valine-threonine and leucine-specific binding proteins, respectively. In this study, the nucleotide sequence of the 4-kb downstream segment was determined and found to contain four reading frames, designated as livA, livE, livF, and livG, that encode putative membrane-associated proteins. The livA and livE genes encode hydrophobic proteins composed of 308 and 425 amino acid residues, respectively. The livF and livG genes encode hydrophilic proteins of 255 and 237 amino acids, respectively; both the proteins contain consensus amino acid sequences found in proteins with ATP-binding sites. These four genes linked together have a potential rho-independent transcriptional terminator adjacent to the 3'-end of livG. No promoter sequence was found in the immediate upstream region of the livAEFG cluster. The livA, livE, livF, and livG gene products were identified as proteins with apparent M(r)s of 25,500, 34,500, 28,000, and 26,000, respectively, by SDS-polyacryl-amide gel electrophoresis. The deduced amino acid sequences of these four proteins showed strong homology to those of the corresponding membrane-associated proteins required for the high-affinity branched-chain amino acid transport systems from both Escherichia coli and Pseudomonas aeruginosa. PMID:1429514