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Sample records for acid sequence level

  1. Complete Genome Sequence of Enterobacter cloacae UW5, a Rhizobacterium Capable of High Levels of Indole-3-Acetic Acid Production.

    PubMed

    Coulson, Thomas J D; Patten, Cheryl L

    2015-08-06

    We report the complete genome sequence of Enterobacter cloacae UW5, an indole-3-acetic acid-producing rhizobacterium originally isolated from the rhizosphere of grass. The 4.9-Mbp genome has a G+C content of 54% and contains 4,496 protein-coding sequences.

  2. Complete Genome Sequence of Enterobacter cloacae UW5, a Rhizobacterium Capable of High Levels of Indole-3-Acetic Acid Production

    PubMed Central

    Coulson, Thomas J. D.

    2015-01-01

    We report the complete genome sequence of Enterobacter cloacae UW5, an indole-3-acetic acid-producing rhizobacterium originally isolated from the rhizosphere of grass. The 4.9-Mbp genome has a G+C content of 54% and contains 4,496 protein-coding sequences. PMID:26251488

  3. Complete genome sequence of Lactococcus lactis IO-1, a lactic acid bacterium that utilizes xylose and produces high levels of L-lactic acid.

    PubMed

    Kato, Hiroaki; Shiwa, Yuh; Oshima, Kenshiro; Machii, Miki; Araya-Kojima, Tomoko; Zendo, Takeshi; Shimizu-Kadota, Mariko; Hattori, Masahira; Sonomoto, Kenji; Yoshikawa, Hirofumi

    2012-04-01

    We report the complete genome sequence of Lactococcus lactis IO-1 (= JCM7638). It is a nondairy lactic acid bacterium, produces nisin Z, ferments xylose, and produces predominantly L-lactic acid at high xylose concentrations. From ortholog analysis with other five L. lactis strains, IO-1 was identified as L. lactis subsp. lactis.

  4. 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.

  5. High speed nucleic acid sequencing

    SciTech Connect

    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. 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.

  7. 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

  8. The complete amino acid sequence of prochymosin.

    PubMed Central

    Foltmann, B; Pedersen, V B; Jacobsen, H; Kauffman, D; Wybrandt, G

    1977-01-01

    The total sequence of 365 amino acid residues in bovine prochymosin is presented. Alignment with the amino acid sequence of porcine pepsinogen shows that 204 amino acid residues are common to the two zymogens. Further comparison and alignment with the amino acid sequence of penicillopepsin shows that 66 residues are located at identical positions in all three proteases. The three enzymes belong to a large group of proteases with two aspartate residues in the active center. This group forms a family derived from one common ancestor. PMID:329280

  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. 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.

  13. 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.

  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. 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...

  16. 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.

  17. Los Alamos sequence analysis package for nucleic acids and proteins.

    PubMed Central

    Kanehisa, M I

    1982-01-01

    An interactive system for computer analysis of nucleic acid and protein sequences has been developed for the Los Alamos DNA Sequence Database. It provides a convenient way to search or verify various sequence features, e.g., restriction enzyme sites, protein coding frames, and properties of coded proteins. Further, the comprehensive analysis package on a large-scale database can be used for comparative studies on sequence and structural homologies in order to find unnoted information stored in nucleic acid sequences. PMID:6174934

  18. 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.

  19. The complementary deoxyribonucleic acid sequence of guinea pig endometrial prorelaxin.

    PubMed

    Lee, Y A; Bryant-Greenwood, G D; Mandel, M; Greenwood, F C

    1992-03-01

    The nucleotide sequence of the relaxin gene transcript in the endometrium of the late pregnant guinea pig has been determined. The strategy used was a combination of polymerase chain reaction (PCR) with primers designed from the mRNA sequence of porcine preprorelaxin, rapid amplification of cDNA ends-PCR, and blunt end cloning in M13 mp18. With heterologous primers, a 226-basepair (bp) segment of the guinea pig relaxin gene sequence was obtained and was used to design a guinea pig-specific primer for use with the rapid amplification of cDNA ends-PCR method. The latter allowed completion of the sequence of 336 bp, with a 96-bp overlap. The sequence obtained shows greater homology at both the nucleotide and amino acid levels with porcine and human relaxins H1 and H2 than with rat relaxin, supporting the thesis that the guinea pig is not a rodent. The transcription of the guinea pig endometrial relaxin gene during pregnancy was confirmed by Northern analysis of guinea pig endometrial tissues with a species-specific cDNA probe. The endometrial relaxin gene is transcribed during pregnancy, but not in lactation, consistent with the observed immunostaining for relaxin.

  20. 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.

  1. 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.

  2. 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

  3. 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.

  4. 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.

  5. Comparison of Levels of Human Immunodeficiency Virus Type 1 RNA in Plasma as Measured by the NucliSens Nucleic Acid Sequence-Based Amplification and Quantiplex Branched-DNA Assays

    PubMed Central

    Ginocchio, C. C.; Tetali, S.; Washburn, D.; Zhang, F.; Kaplan, M. H.

    1999-01-01

    This study compared levels of human immunodeficiency virus type 1 RNA in plasma as measured by the Quantiplex branched-DNA and NucliSens nucleic acid sequence-based amplification assays. RNA was detectable in 118 of 184 samples (64.13%) by the Quantiplex assay and in 171 of 184 samples (92.94%) by the NucliSens assay. Regression analysis indicated that a linear relationship existed between the two sets of values (P < 0.0001), although the Quantiplex and NucliSens values were significantly different (P < 0.001), with the NucliSens values being approximately 0.323 log higher. Spearman correlation analysis indicated that the overall changes in patient viral load patterns were highly correlative between the two assays: r = 0.912, P < 0.0001. The lower limits of sensitivity were determined to be approximately 100 copies/ml and 1,200 to 1,400 copies/ml for the NucliSens and Quantiplex assays, respectively. PMID:10074556

  6. Comparison of levels of human immunodeficiency virus type 1 RNA in plasma as measured by the NucliSens nucleic acid sequence-based amplification and Quantiplex branched-DNA assays.

    PubMed

    Ginocchio, C C; Tetali, S; Washburn, D; Zhang, F; Kaplan, M H

    1999-04-01

    This study compared levels of human immunodeficiency virus type 1 RNA in plasma as measured by the Quantiplex branched-DNA and NucliSens nucleic acid sequence-based amplification assays. RNA was detectable in 118 of 184 samples (64.13%) by the Quantiplex assay and in 171 of 184 samples (92.94%) by the NucliSens assay. Regression analysis indicated that a linear relationship existed between the two sets of values (P < 0.0001), although the Quantiplex and NucliSens values were significantly different (P < 0.001), with the NucliSens values being approximately 0.323 log higher. Spearman correlation analysis indicated that the overall changes in patient viral load patterns were highly correlative between the two assays: r = 0.912, P < 0.0001. The lower limits of sensitivity were determined to be approximately 100 copies/ml and 1,200 to 1,400 copies/ml for the NucliSens and Quantiplex assays, respectively.

  7. 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.

  8. Dipeptide Sequence Determination: Analyzing Phenylthiohydantoin Amino Acids by HPLC

    NASA Astrophysics Data System (ADS)

    Barton, Janice S.; Tang, Chung-Fei; Reed, Steven S.

    2000-02-01

    Amino acid composition and sequence determination, important techniques for characterizing peptides and proteins, are essential for predicting conformation and studying sequence alignment. This experiment presents improved, fundamental methods of sequence analysis for an upper-division biochemistry laboratory. Working in pairs, students use the Edman reagent to prepare phenylthiohydantoin derivatives of amino acids for determination of the sequence of an unknown dipeptide. With a single HPLC technique, students identify both the N-terminal amino acid and the composition of the dipeptide. This method yields good precision of retention times and allows use of a broad range of amino acids as components of the dipeptide. Students learn fundamental principles and techniques of sequence analysis and HPLC.

  9. Effect of domoic acid on brain amino acid levels.

    PubMed

    Durán, R; Arufe, M C; Arias, B; Alfonso, M

    1995-03-01

    The administration of Domoic Acid (Dom) in a 0.2 mg/kg i.p. dose induces changes in the levels of amino acids of neurochemical interest (Asp, Glu, Gly, Tau, Ala, GABA) in different rat brain regions (hypothalamus, hippocampus, amygdala, striatum, cortex and midbrain). The most affected amino acid is the GABA, the main inhibitory amino acid neurotransmitter, whereas glutamate, the main excitatory amino acid, is not affected. The rat brain regions that seem to be the main target of the Dom action belong to the limbic system (hippocampus, amygdala). The possible implication of the amino acids in the actions of Dom is also discussed.

  10. Amino acid sequence of mouse submaxillary gland renin.

    PubMed Central

    Misono, K S; Chang, J J; Inagami, T

    1982-01-01

    The complete amino acid sequences of the heavy chain and light chain of mouse submaxillary gland renin have been determined. The heavy chain consists of 288 amino acid residues having a Mr of 31,036 calculated from the sequence. The light chain contains 48 amino acid residues with a Mr of 5,458. The sequence of the heavy chain was determined by automated Edman degradations of the cyanogen bromide peptides and tryptic peptides generated after citraconylation, as well as other peptides generated therefrom. The sequence of the light chain was derived from sequence analyses of the peptides generated by cyanogen bromide cleavage or by digestion with Staphylococcus aureus protease. The sequences in the active site regions in renin containing two catalytically essential aspartyl residues 32 and 215 were found identical with those in pepsin, chymosin, and penicillopepsin. Comparison of the amino acid sequence of renin with that of porcine pepsin indicated a 42% sequence identity of the heavy chain with the amino-terminal and middle regions and a 46% identity of the light chain with the carboxyl-terminal region of the porcine pepsin sequence. Residues identical in renin and pepsin are distributed throughout the length of the molecules, suggesting a similarity in their overall structures. PMID:6812055

  11. Amino Acid Sequence of Human Cholinesterase

    DTIC Science & Technology

    1985-10-01

    liquid chromatography (HPLC). Activity testing of the aged, DFP-labeled cholinesterase showed that 99.8% of the active sites had been labeled, since...acids were quantitated by ninhydrin at the AAA Labs, or by derivatization with phenylisothiocyanate at the University of Michigan. The latter method

  12. Analysis of cloned cDNA and genomic sequences for phytochrome: complete amino acid sequences for two gene products expressed in etiolated Avena.

    PubMed Central

    Hershey, H P; Barker, R F; Idler, K B; Lissemore, J L; Quail, P H

    1985-01-01

    Cloned cDNA and genomic sequences have been analyzed to deduce the amino acid sequence of phytochrome from etiolated Avena. Restriction endonuclease site polymorphism between clones indicates that at least four phytochrome genes are expressed in this tissue. Sequence analysis of two complete and one partial coding region shows approximately 98% homology at both the nucleotide and amino acid levels, with the majority of amino acid changes being conservative. High sequence homology is also found in the 5'-untranslated region but significant divergence occurs in the 3'-untranslated region. The phytochrome polypeptides are 1128 amino acid residues long corresponding to a molecular mass of 125 kdaltons. The known protein sequence at the chromophore attachment site occurs only once in the polypeptide, establishing that phytochrome has a single chromophore per monomer covalently linked to Cys-321. Computer analyses of the amino acid sequences have provided predictions regarding a number of structural features of the phytochrome molecule. PMID:3001642

  13. Cystatin. Amino acid sequence and possible secondary structure.

    PubMed Central

    Schwabe, C; Anastasi, A; Crow, H; McDonald, J K; Barrett, A J

    1984-01-01

    The amino acid sequence of cystatin, the protein from chicken egg-white that is a tight-binding inhibitor of many cysteine proteinases, is reported. Cystatin is composed of 116 amino acid residues, and the Mr is calculated to be 13 143. No striking similarity to any other known sequence has been detected. The results of computer analysis of the sequence and c.d. spectrometry indicate that the secondary structure includes relatively little alpha-helix (about 20%) and that the remainder is mainly beta-structure. PMID:6712597

  14. Mouse Vk gene classification by nucleic acid sequence similarity.

    PubMed

    Strohal, R; Helmberg, A; Kroemer, G; Kofler, R

    1989-01-01

    Analyses of immunoglobulin (Ig) variable (V) region gene usage in the immune response, estimates of V gene germline complexity, and other nucleic acid hybridization-based studies depend on the extent to which such genes are related (i.e., sequence similarity) and their organization in gene families. While mouse Igh heavy chain V region (VH) gene families are relatively well-established, a corresponding systematic classification of Igk light chain V region (Vk) genes has not been reported. The present analysis, in the course of which we reviewed the known extent of the Vk germline gene repertoire and Vk gene usage in a variety of responses to foreign and self antigens, provides a classification of mouse Vk genes in gene families composed of members with greater than 80% overall nucleic acid sequence similarity. This classification differed in several aspects from that of VH genes: only some Vk gene families were as clearly separated (by greater than 25% sequence dissimilarity) as typical VH gene families; most Vk gene families were closely related and, in several instances, members from different families were very similar (greater than 80%) over large sequence portions; frequently, classification by nucleic acid sequence similarity diverged from existing classifications based on amino-terminal protein sequence similarity. Our data have implications for Vk gene analyses by nucleic acid hybridization and describe potentially important differences in sequence organization between VH and Vk genes.

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

    PubMed Central

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

    2016-01-01

    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

  16. 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.

  17. Amino acid sequences of proteins from Leptospira serovar pomona.

    PubMed

    Alves, S F; Lefebvre, R B; Probert, W

    2000-01-01

    This report describes a partial amino acid sequences from three putative outer envelope proteins from Leptospira serovar pomona. In order to obtain internal fragments for protein sequencing, enzymatic and chemical digestion was performed. The enzyme clostripain was used to digest the proteins 32 and 45 kDa. In situ digestion of 40 kDa molecular weight protein was accomplished using cyanogen bromide. The 32 kDa protein generated two fragments, one of 21 kDa and another of 10 kDa that yielded five residues. A fragment of 24 kDa that yielded nineteen residues of amino acids was obtained from 45 kDa protein. A fragment with a molecular weight of 20 kDa, yielding a twenty amino acids sequence from the 40 kDa protein.

  18. Extensive amino acid sequence homologies between animal lectins

    SciTech Connect

    Paroutaud, P.; Levi, G.; Teichberg, V.I.; Strosberg, A.D.

    1987-09-01

    The authors have established the amino acid sequence of the ..beta..-D-galactoside binding lectin from the electric eel and the sequences of several peptides from a similar lectin isolated from human placenta. These sequences were compared with the published sequences of peptides derived from the ..beta..-D-galactoside binding lectin from human lung and with sequences deduced from cDNAs assigned to the ..beta..-D-galactoside binding lectins from chicken embryo skin and human hepatomas. Significant homologies were observed. One of the highly conserved regions that contains a tryptophan residue and two glutamic acid resides is probably part of the ..beta..-D-galactoside binding site, which, on the basis of spectroscopic studies of the electric eel lectin, is expected to contain such residues. The similarity of the hydropathy profiles and the predicted secondary structure of the lectins from chicken skin and electric eel, in spite of differences in their amino acid sequences, strongly suggests that these proteins have maintained structural homologies during evolution and together with the other ..beta..-D-galactoside binding lectins were derived form a common ancestor gene.

  19. Amino acid sequence of porcine spleen cathepsin D.

    PubMed Central

    Shewale, J G; Tang, J

    1984-01-01

    The amino acid sequence of porcine spleen cathepsin D heavy chain has been determined and, hence, the complete structure of this enzyme is now known. The sequence of heavy chain was constructed by aligning the structures of peptides generated by cyanogen bromide, trypsin, and endo-proteinase Lys C cleavages. The structure of the light chain has been published previously. The cathepsin D molecule contains 339 amino acid residues in two polypeptide chains: a 97-residue light chain and a 242-residue heavy chain, with a combined Mr of 36,779 (without carbohydrate). There are two carbohydrate units linked to asparagine residues 70 and 192. The disulfide bond arrangement in cathepsin D is probably similar to that of pepsin, because the positions of six half-cystine residues are conserved. The active site aspartyl residues, corresponding to aspartic acid-32 and -215 of pepsin, are located at residues 33 and 224 in the cathepsin D molecule. The amino acid sequence around these aspartyl residues is strongly conserved. Cathepsin D shows a strong homology with other acid proteases. When the sequence of cathepsin D, renin, and pepsin are aligned, 32.7% of the residues are identical. The homology is observed throughout the length of the molecules, indicating that three-dimensional structures of all three molecules are similar. PMID:6587385

  20. Purification, characterization and partial amino acid sequence of glycogen synthase from Saccharomyces cerevisiae.

    PubMed Central

    Carabaza, A; Arino, J; Fox, J W; Villar-Palasi, C; Guinovart, J J

    1990-01-01

    Glycogen synthase from Saccharomyces cerevisiae was purified to homogeneity. The enzyme showed a subunit molecular mass of 80 kDa. The holoenzyme appears to be a tetramer. Antibodies developed against purified yeast glycogen synthase inactivated the enzyme in yeast extracts and allowed the detection of the protein in Western blots. Amino acid analysis showed that the enzyme is very rich in glutamate and/or glutamine residues. The N-terminal sequence (11 amino acid residues) was determined. In addition, selected tryptic-digest peptides were purified by reverse-phase h.p.l.c. and submitted to gas-phase sequencing. Up to eight sequences (79 amino acid residues) could be aligned with the human muscle enzyme sequence. Levels of identity range between 37 and 100%, indicating that, although human and yeast glycogen synthases probably share some conserved regions, significant differences in their primary structure should be expected. Images Fig. 1. Fig. 2. Fig. 3. PMID:2114092

  1. Active site amino acid sequence of human factor D.

    PubMed

    Davis, A E

    1980-08-01

    Factor D was isolated from human plasma by chromatography on CM-Sephadex C50, Sephadex G-75, and hydroxylapatite. Digestion of reduced, S-carboxymethylated factor D with cyanogen bromide resulted in three peptides which were isolated by chromatography on Sephadex G-75 (superfine) equilibrated in 20% formic acid. NH2-Terminal sequences were determined by automated Edman degradation with a Beckman 890C sequencer using a 0.1 M Quadrol program. The smallest peptide (CNBr III) consisted of the NH2-terminal 14 amino acids. The other two peptides had molecular weights of 17,000 (CNBr I) and 7000 (CNBr II). Overlap of the NH2-terminal sequence of factor D with the NH2-terminal sequence of CNBr I established the order of the peptides. The NH2-terminal 53 residues of factor D are somewhat more homologous with the group-specific protease of rat intestine than with other serine proteases. The NH2-terminal sequence of CNBr II revealed the active site serine of factor D. The typical serine protease active site sequence (Gly-Asp-Ser-Gly-Gly-Pro was found at residues 12-17. The region surrounding the active site serine does not appear to be more highly homologous with any one of the other serine proteases. The structural data obtained point out the similarities between factor D and the other proteases. However, complete definition of the degree of relationship between factor D and other proteases will require determination of the remainder of the primary structure.

  2. TranslatorX: multiple alignment of nucleotide sequences guided by amino acid translations.

    PubMed

    Abascal, Federico; Zardoya, Rafael; Telford, Maximilian J

    2010-07-01

    We present TranslatorX, a web server designed to align protein-coding nucleotide sequences based on their corresponding amino acid translations. Many comparisons between biological sequences (nucleic acids and proteins) involve the construction of multiple alignments. Alignments represent a statement regarding the homology between individual nucleotides or amino acids within homologous genes. As protein-coding DNA sequences evolve as triplets of nucleotides (codons) and it is known that sequence similarity degrades more rapidly at the DNA than at the amino acid level, alignments are generally more accurate when based on amino acids than on their corresponding nucleotides. TranslatorX novelties include: (i) use of all documented genetic codes and the possibility of assigning different genetic codes for each sequence; (ii) a battery of different multiple alignment programs; (iii) translation of ambiguous codons when possible; (iv) an innovative criterion to clean nucleotide alignments with GBlocks based on protein information; and (v) a rich output, including Jalview-powered graphical visualization of the alignments, codon-based alignments coloured according to the corresponding amino acids, measures of compositional bias and first, second and third codon position specific alignments. The TranslatorX server is freely available at http://translatorx.co.uk.

  3. The amino acid sequence of iguana (Iguana iguana) pancreatic ribonuclease.

    PubMed

    Zhao, W; Beintema, J J; Hofsteenge, J

    1994-01-15

    The pyrimidine-specific ribonuclease superfamily constitutes a group of homologous proteins so far found only in higher vertebrates. Four separate families are found in mammals, which have resulted from gene duplications in mammalian ancestors. To learn more about the evolutionary history of this superfamily, the primary structure and other characteristics of the pancreatic enzyme from iguana (Iguana iguana), a herbivorous lizard species belonging to the reptiles, have been determined. The polypeptide chain consists of 119 amino acid residues. The positions of insertions and deletions in the sequence are identical to those in the enzyme from snapping turtle. However, the two enzymes differ at 54% of the amino acid positions. Iguana ribonuclease contains no carbohydrate, although the enzyme possesses three recognition sites for carbohydrate attachment, and has a high number of acidic residues in a localized part of the sequence.

  4. 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

  5. Amino acid sequence and comparative antigenicity of chicken metallothionein.

    PubMed Central

    McCormick, C C; Fullmer, C S; Garvey, J S

    1988-01-01

    The complete amino acid sequence of metallothionein (MT) from chicken liver is reported. The primary structure was determined by automated sequence analysis of peptides produced by limited acid hydrolysis and by trypsin digestion. The comparative antigenicity of chicken MT was determined by radioimmunoassay using rabbit anti-rat MT polyclonal antibody. Chicken MT consists of 63 amino acids as compared to 61 found in MTs from mammals. One insertion (and two substitutions) occurs in the amino-terminal region, a region considered invariant among mammalian MTs. Eighteen of the 20 cysteines in chicken MT were aligned with cysteines from other mammalian sequences. Two cysteines near the carboxyl terminus are shifted by one residue due to the insertion of proline in that region. Overall, the chicken protein showed approximately equal to 68% sequence identity in a comparison with various mammalian MTs. The affinity of the polyclonal antibody for chicken MT was decreased by 2 orders of magnitude in comparison to that of a mammalian MT (rat MT isoforms). This reduced affinity is attributed to major substitutions in chicken MT in the regions of the principal determinants of mammalian MTs. Theoretical analysis of the primary structure predicted the secondary structure to consist of reverse turns and random coils with no stable beta or helix conformations. There is no evidence that chicken MT differs functionally from mammalian MTs. PMID:2448773

  6. 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.

  7. 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.

  8. Sequence determinants of prokaryotic gene expression level under heat stress.

    PubMed

    Xiong, Heng; Yang, Yi; Hu, Xiao-Pan; He, Yi-Ming; Ma, Bin-Guang

    2014-11-01

    Prokaryotic gene expression is environment-dependent and temperature plays an important role in shaping the gene expression profile. Revealing the regulation mechanisms of gene expression pertaining to temperature has attracted tremendous efforts in recent years particularly owning to the yielding of transcriptome and proteome data by high-throughput techniques. However, most of the previous works concentrated on the characterization of the gene expression profile of individual organism and little effort has been made to disclose the commonality among organisms, especially for the gene sequence features. In this report, we collected the transcriptome and proteome data measured under heat stress condition from recently published literature and studied the sequence determinants for the expression level of heat-responsive genes on multiple layers. Our results showed that there indeed exist commonness and consistent patterns of the sequence features among organisms for the differentially expressed genes under heat stress condition. Some features are attributed to the requirement of thermostability while some are dominated by gene function. The revealed sequence determinants of bacterial gene expression level under heat stress complement the knowledge about the regulation factors of prokaryotic gene expression responding to the change of environmental conditions. Furthermore, comparisons to thermophilic adaption have been performed to reveal the similarity and dissimilarity of the sequence determinants for the response to heat stress and for the adaption to high habitat temperature, which elucidates the complex landscape of gene expression related to the same physical factor of temperature.

  9. Nucleic and Amino Acid Sequences Support Structure-Based Viral Classification

    PubMed Central

    Sinclair, Robert M.; Ravantti, Janne J.

    2017-01-01

    ABSTRACT Viral capsids ensure viral genome integrity by protecting the enclosed nucleic acids. Interactions between the genome and capsid and between individual capsid proteins (i.e., capsid architecture) are intimate and are expected to be characterized by strong evolutionary conservation. For this reason, a capsid structure-based viral classification has been proposed as a way to bring order to the viral universe. The seeming lack of sufficient sequence similarity to reproduce this classification has made it difficult to reject structural convergence as the basis for the classification. We reinvestigate whether the structure-based classification for viral coat proteins making icosahedral virus capsids is in fact supported by previously undetected sequence similarity. Since codon choices can influence nascent protein folding cotranslationally, we searched for both amino acid and nucleotide sequence similarity. To demonstrate the sensitivity of the approach, we identify a candidate gene for the pandoravirus capsid protein. We show that the structure-based classification is strongly supported by amino acid and also nucleotide sequence similarities, suggesting that the similarities are due to common descent. The correspondence between structure-based and sequence-based analyses of the same proteins shown here allow them to be used in future analyses of the relationship between linear sequence information and macromolecular function, as well as between linear sequence and protein folds. IMPORTANCE Viral capsids protect nucleic acid genomes, which in turn encode capsid proteins. This tight coupling of protein shell and nucleic acids, together with strong functional constraints on capsid protein folding and architecture, leads to the hypothesis that capsid protein-coding nucleotide sequences may retain signatures of ancient viral evolution. We have been able to show that this is indeed the case, using the major capsid proteins of viruses forming icosahedral capsids

  10. 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.

  11. Accident sequence precursor analysis level 2/3 model development

    SciTech Connect

    Lui, C.H.; Galyean, W.J.; Brownson, D.A.

    1997-02-01

    The US Nuclear Regulatory Commission`s Accident Sequence Precursor (ASP) program currently uses simple Level 1 models to assess the conditional core damage probability for operational events occurring in commercial nuclear power plants (NPP). Since not all accident sequences leading to core damage will result in the same radiological consequences, it is necessary to develop simple Level 2/3 models that can be used to analyze the response of the NPP containment structure in the context of a core damage accident, estimate the magnitude of the resulting radioactive releases to the environment, and calculate the consequences associated with these releases. The simple Level 2/3 model development work was initiated in 1995, and several prototype models have been completed. Once developed, these simple Level 2/3 models are linked to the simple Level 1 models to provide risk perspectives for operational events. This paper describes the methods implemented for the development of these simple Level 2/3 ASP models, and the linkage process to the existing Level 1 models.

  12. 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.

  13. 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

  14. The amino acid sequence of rabbit cardiac troponin I.

    PubMed Central

    Grand, R J; Wilkinson, J M

    1976-01-01

    The complete amino acid sequence of troponin I from rabbit cardiac muscle was determined by the isolation of four unique CNBr fragments, together with overlapping tryptic peptides containing radioactive methionine residues. Overlap data for residues 35-36, 93-94 and 140-145 are incomplete, the sequence at these positions being based on homology with the sequence of the fast-skeletal-muscle protein. Cardiac troponin I is a single polypeptide chain of 206 residues with mol.wt. 23550 and an extinction coefficient, E 1%,1cm/280, of 4.37. The protein has a net positive charge of 14 and is thus somewhat more basic than troponin I from fast-skeletal muscle. Comparison of the sequences of troponin I from cardiac and fast skeletal muscle show that the cardiac protein has 26 extra residues at the N-terminus which account for the larger size of the protein. In the remainder of sequence there is a considerable degree of homology, this being greater in the C-terminal two-thirds of the molecule. The region in the cardiac protein corresponding to the peptide with inhibitory activity from the fast-skeletal-muscle protein is very similar and it seems unlikely that this is the cause of the difference in inhibitory activity between the two proteins. The region responsible for binding troponin C, however, possesses a lower degree of homology. Detailed evidence on which the sequence is based has been deposited as Supplementary Publication SUP 50072 (20 pages), at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7QB, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1976) 153, 5. PMID:1008822

  15. Amino acid sequence of a mouse immunoglobulin mu chain.

    PubMed Central

    Kehry, M; Sibley, C; Fuhrman, J; Schilling, J; Hood, L E

    1979-01-01

    The complete amino acid sequence of the mouse mu chain from the BALB/c myeloma tumor MOPC 104E is reported. The C mu region contains four consecutive homology regions of approximately 110 residues and a COOH-terminal region of 19 residues. A comparison of this mu chain from mouse with a complete mu sequence from human (Ou) and a partial mu chain sequence from dog (Moo) reveals a striking gradient of increasing homology from the NH2-terminal to the COOH-terminal portion of these mu chains, with the former being the least and the latter the most highly conserved. Four of the five sites of carbohydrate attachment appear to be at identical residue positions when the constant regions of the mouse and human mu chains are compared. The mu chain of MOPC 104E has a carbohydrate moiety attached in the second hypervariable region. This is particularly interesting in view of the fact that MOPC 104E binds alpha-(1 leads to 3)-dextran, a simple carbohydrate. The structural and functional constraints imposed by these comparative sequence analyses are discussed. PMID:111247

  16. Sequence-level mechanisms of human epigenome evolution.

    PubMed

    Prendergast, James G D; Chambers, Emily V; Semple, Colin A M

    2014-06-24

    DNA methylation and chromatin states play key roles in development and disease. However, the extent of recent evolutionary divergence in the human epigenome and the influential factors that have shaped it are poorly understood. To determine the links between genome sequence and human epigenome evolution, we examined the divergence of DNA methylation and chromatin states following segmental duplication events in the human lineage. Chromatin and DNA methylation states were found to have been generally well conserved following a duplication event, with the evolution of the epigenome largely uncoupled from the total number of genetic changes in the surrounding DNA sequence. However, the epigenome at tissue-specific, distal regulatory regions was observed to be unusually prone to diverge following duplication, with particular sequence differences, altering known sequence motifs, found to be associated with divergence in patterns of DNA methylation and chromatin. Alu elements were found to have played a particularly prominent role in shaping human epigenome evolution, and we show that human-specific AluY insertion events are strongly linked to the evolution of the DNA methylation landscape and gene expression levels, including at key neurological genes in the human brain. Studying paralogous regions within the same sample enables the study of the links between genome and epigenome evolution while controlling for biological and technical variation. We show DNA methylation and chromatin divergence between duplicated regions are linked to the divergence of particular genetic motifs, with Alu elements having played a disproportionate role in the evolution of the epigenome in the human lineage.

  17. Diverse Levels of Sequence Selectivity and Catalytic Efficiency of Protein-Tyrosine Phosphatases

    PubMed Central

    Selner, Nicholas G.; Luechapanichkul, Rinrada; Chen, Xianwen; Neel, Benjamin G.; Zhang, Zhong-Yin; Knapp, Stefan; Bell, Charles E.; Pei, Dehua

    2014-01-01

    The sequence selectivity of 14 classical protein-tyrosine phosphatases (PTPs) (PTPRA, PTPRB, PTPRC, PTPRD, PTPRO, PTP1B, SHP-1, SHP-2, HePTP, PTP-PEST, TCPTP, PTPH1, PTPD1, and PTPD2) was systematically profiled by screening their catalytic domains against combinatorial peptide libraries. All of the PTPs exhibit similar preference for pY peptides rich in acidic amino acids and disfavor positively charged sequences, but differ vastly in their degrees of preference/disfavor. Some PTPs (PTP-PEST, SHP-1, and SHP-2) are highly selective for acidic over basic (or neutral) peptides (by >105-fold), whereas others (PTPRA and PTPRD) show no to little sequence selectivity. PTPs also have diverse intrinsic catalytic efficiencies (kcat/KM values against optimal substrates), which differ by >105-fold due to different kcat and/or KM values. Moreover, PTPs show little positional preference for the acidic residues relative to the pY residue. Mutation of Arg47 of PTP1B, which is located near the pY-1 and pY-2 residues of a bound substrate, decreased the enzymatic activity by 3–18-fold toward all pY substrates containing acidic residues anywhere within the pY-6 to pY+5 region. Similarly, mutation of Arg24, which is situated near the C-terminus of a bound substrate, adversely affected the kinetic activity of all acidic substrates. A co-crystal structure of PTP1B bound with a nephrin pY1193 peptide suggests that Arg24 engages in electrostatic interactions with acidic residues at the pY+1, pY+2, and likely other positions. These results suggest that long-range electrostatic interactions between positively charged residues near the PTP active site and acidic residues on pY substrates allow a PTP to bind acidic substrates with similar affinities and the varying levels of preference for acidic sequences by different PTPs are likely caused by the different electrostatic potentials near their active sites. The implications of the varying sequence selectivity and intrinsic catalytic

  18. 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.

  19. Nucleic acid (cDNA) and amino acid sequences of alpha-type gliadins from wheat (Triticum aestivum).

    PubMed Central

    Kasarda, D D; Okita, T W; Bernardin, J E; Baecker, P A; Nimmo, C C; Lew, E J; Dietler, M D; Greene, F C

    1984-01-01

    The complete amino acid sequence for an alpha-type gliadin protein of wheat (Triticum aestivum Linnaeus) endosperm has been derived from a cloned cDNA sequence. An additional cDNA clone that corresponds to about 75% of a similar alpha-type gliadin has been sequenced and shows some important differences. About 97% of the composite sequence of A-gliadin (an alpha-type gliadin fraction) has also been obtained by direct amino acid sequencing. This sequence shows a high degree of similarity with amino acid sequences derived from both cDNA clones and is virtually identical to one of them. On the basis of sequence information, after loss of the signal sequence, the mature alpha-type gliadins may be divided into five different domains, two of which may have evolved from an ancestral gliadin gene, whereas the remaining three contain repeating sequences that may have developed independently. Images PMID:6589619

  20. Structural gene and complete amino acid sequence of Vibrio alginolyticus collagenase.

    PubMed Central

    Takeuchi, H; Shibano, Y; Morihara, K; Fukushima, J; Inami, S; Keil, B; Gilles, A M; Kawamoto, S; Okuda, K

    1992-01-01

    The DNA encoding the collagenase of Vibrio alginolyticus 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 both collagenase antigen and collagenase activity. The open reading frame from the ATG initiation codon was 2442 bp in length for the collagenase structural gene. The amino acid sequence, deduced from the nucleotide sequence, revealed that the mature collagenase consists of 739 amino acids with an Mr of 81875. The amino acid sequences of 20 polypeptide fragments were completely identical with the deduced amino acid sequences of the collagenase gene. The amino acid composition predicted from the DNA sequence was similar to the chemically determined composition of purified collagenase reported previously. The analyses of both the DNA and amino acid sequences of the collagenase gene were rigorously performed, but we could not detect any significant sequence similarity to other collagenases. Images Fig. 2. PMID:1311172

  1. 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

  2. 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

    2016-07-28

    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.

  3. 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....

  4. Multiple site-selective insertions of non-canonical amino acids into sequence-repetitive polypeptides

    PubMed Central

    Wu, I-Lin; Patterson, Melissa A.; Carpenter Desai, Holly E.; Mehl, Ryan A.; Giorgi, Gianluca

    2013-01-01

    A simple and efficient method is described for introduction of non-canonical amino acids at multiple, structurally defined sites within recombinant polypeptide sequences. E. coli MRA30, a bacterial host strain with attenuated activity for release factor 1 (RF1), is assessed for its ability to support the incorporation of a diverse range of non-canonical amino acids in response to multiple encoded amber (TAG) codons within genetic templates derived from superfolder GFP and an elastin-mimetic protein polymer. Suppression efficiency and isolated protein yield were observed to depend on the identity of the orthogonal aminoacyl-tRNA synthetase/tRNACUA pair and the non-canonical amino acid substrate. This approach afforded elastin-mimetic protein polymers containing non-canonical amino acid derivatives at up to twenty-two positions within the repeat sequence with high levels of substitution. The identity and position of the variant residues was confirmed by mass spectrometric analysis of the full-length polypeptides and proteolytic cleavage fragments resulting from thermolysin digestion. The accumulated data suggest that this multi-site suppression approach permits the preparation of protein-based materials in which novel chemical functionality can be introduced at precisely defined positions within the polypeptide sequence. PMID:23625817

  5. Nucleic acid (cDNA) and amino acid sequences of the maize endosperm protein glutelin-2.

    PubMed Central

    Prat, S; Cortadas, J; Puigdomènech, P; Palau, J

    1985-01-01

    The cDNA coding for a glutelin-2 protein from maize endosperm has been cloned and the complete amino acid sequence of the protein derived for the first time. An immature maize endosperm cDNA bank was screened for the expression of a beta-lactamase:glutelin-2 (G2) fusion polypeptide by using antibodies against the purified 28 kd G2 protein. A clone corresponding to the 28 kd G2 protein was sequenced and the primary structure of this protein was derived. Five regions can be defined in the protein sequence: an 11 residue N-terminal part, a repeated region formed by eight units of the sequence Pro-Pro-Pro-Val-His-Leu, an alternating Pro-X stretch 21 residues long, a Cys rich domain and a C-terminal part rich in Gln. The protein sequence is preceded by 19 residues which have the characteristics of the signal peptide found in secreted proteins. Unlike zeins, the main maize storage proteins, 28 kd glutelin-2 has several homologous sequences in common with other cereal storage proteins. Images PMID:3839076

  6. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 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 Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide...

  7. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2012-07-01 2012-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide...

  8. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2014-07-01 2014-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide...

  9. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 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 Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide...

  10. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2013-07-01 2013-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide...

  11. Design, synthesis, and characterization of a protein sequencing reagent yielding amino acid derivatives with enhanced detectability by mass spectrometry.

    PubMed Central

    Aebersold, R.; Bures, E. J.; Namchuk, M.; Goghari, M. H.; Shushan, B.; Covey, T. C.

    1992-01-01

    We report the design, chemical synthesis, and structural and functional characterization of a novel reagent for protein sequence analysis by the Edman degradation, yielding amino acid derivatives rapidly detectable at high sensitivity by ion-evaporation mass spectrometry. We demonstrate that the reagent 3-[4'(ethylene-N,N,N-trimethylamino)phenyl]-2-isothiocyanate is chemically stable and shows coupling and cyclization/cleavage yields comparable to phenylisothiocyanate, the standard reagent in chemical sequence analysis, under conditions typically encountered in manual or automated sequence analysis. Amino acid derivatives generated with this reagent were detectable by ion-evaporation mass spectrometry at the subfemtomole sensitivity level at a pace of one sample per minute. Furthermore, derivatives were identified by their mass, thus permitting the rapid and highly sensitive determination of the molecular nature of modified amino acids. Derivatives of amino acids with acidic, basic, polar, or hydrophobic side chains were reproducibly detectable at comparable sensitivities. The polar nature of the reagent required covalent immobilization of polypeptides prior to automated sequence analysis. This reagent, used in automated sequence analysis, has the potential for overcoming the limitations in sensitivity, speed, and the ability to characterize modified amino acid residues inherent in the chemical sequencing methods that are currently used. PMID:1304351

  12. Human liver apolipoprotein B-100 cDNA: complete nucleic acid and derived amino acid sequence.

    PubMed Central

    Law, S W; Grant, S M; Higuchi, K; Hospattankar, A; Lackner, K; Lee, N; Brewer, H B

    1986-01-01

    Human apolipoprotein B-100 (apoB-100), the ligand on low density lipoproteins that interacts with the low density lipoprotein receptor and initiates receptor-mediated endocytosis and low density lipoprotein catabolism, has been cloned, and the complete nucleic acid and derived amino acid sequences have been determined. ApoB-100 cDNAs were isolated from normal human liver cDNA libraries utilizing immunoscreening as well as filter hybridization with radiolabeled apoB-100 oligodeoxynucleotides. The apoB-100 mRNA is 14.1 kilobases long encoding a mature apoB-100 protein of 4536 amino acids with a calculated amino acid molecular weight of 512,723. ApoB-100 contains 20 potential glycosylation sites, and 12 of a total of 25 cysteine residues are located in the amino-terminal region of the apolipoprotein providing a potential globular structure of the amino terminus of the protein. ApoB-100 contains relatively few regions of amphipathic helices, but compared to other human apolipoproteins it is enriched in beta-structure. The delineation of the entire human apoB-100 sequence will now permit a detailed analysis of the conformation of the protein, the low density lipoprotein receptor binding domain(s), and the structural relationship between apoB-100 and apoB-48 and will provide the basis for the study of genetic defects in apoB-100 in patients with dyslipoproteinemias. PMID:3464946

  13. Computer selection of oligonucleotide probes from amino acid sequences for use in gene library screening.

    PubMed

    Yang, J H; Ye, J H; Wallace, D C

    1984-01-11

    We present a computer program, FINPROBE, which utilizes known amino acid sequence data to deduce minimum redundancy oligonucleotide probes for use in screening cDNA or genomic libraries or in primer extension. The user enters the amino acid sequence of interest, the desired probe length, the number of probes sought, and the constraints on oligonucleotide synthesis. The computer generates a table of possible probes listed in increasing order of redundancy and provides the location of each probe in the protein and mRNA coding sequence. Activation of a next function provides the amino acid and mRNA sequences of each probe of interest as well as the complementary sequence and the minimum dissociation temperature of the probe. A final routine prints out the amino acid sequence of the protein in parallel with the mRNA sequence listing all possible codons for each amino acid.

  14. Analysis of protein function and its prediction from amino acid sequence.

    PubMed

    Clark, Wyatt T; Radivojac, Predrag

    2011-07-01

    Understanding protein function is one of the keys to understanding life at the molecular level. It is also important in the context of human disease because many conditions arise as a consequence of alterations of protein function. The recent availability of relatively inexpensive sequencing technology has resulted in thousands of complete or partially sequenced genomes with millions of functionally uncharacterized proteins. Such a large volume of data, combined with the lack of high-throughput experimental assays to functionally annotate proteins, attributes to the growing importance of automated function prediction. Here, we study proteins annotated by Gene Ontology (GO) terms and estimate the accuracy of functional transfer from protein sequence only. We find that the transfer of GO terms by pairwise sequence alignments is only moderately accurate, showing a surprisingly small influence of sequence identity (SID) in a broad range (30-100%). We developed and evaluated a new predictor of protein function, functional annotator (FANN), from amino acid sequence. The predictor exploits a multioutput neural network framework which is well suited to simultaneously modeling dependencies between functional terms. Experiments provide evidence that FANN-GO (predictor of GO terms; available from http://www.informatics.indiana.edu/predrag) outperforms standard methods such as transfer by global or local SID as well as GOtcha, a method that incorporates the structure of GO.

  15. Advanced accident sequence precursor analysis level 2 models

    SciTech Connect

    Galyean, W.J.; Brownson, D.A.; Rempe, J.L.

    1996-03-01

    The U.S. Nuclear Regulatory Commission Accident Sequence Precursor program pursues the ultimate objective of performing risk significant evaluations on operational events (precursors) occurring in commercial nuclear power plants. To achieve this objective, the Office of Nuclear Regulatory Research is supporting the development of simple probabilistic risk assessment models for all commercial nuclear power plants (NPP) in the U.S. Presently, only simple Level 1 plant models have been developed which estimate core damage frequencies. In order to provide a true risk perspective, the consequences associated with postulated core damage accidents also need to be considered. With the objective of performing risk evaluations in an integrated and consistent manner, a linked event tree approach which propagates the front end results to back end was developed. This approach utilizes simple plant models that analyze the response of the NPP containment structure in the context of a core damage accident, estimate the magnitude and timing of a radioactive release to the environment, and calculate the consequences for a given release. Detailed models and results from previous studies, such as the NUREG-1150 study, are used to quantify these simple models. These simple models are then linked to the existing Level 1 models, and are evaluated using the SAPHIRE code. To demonstrate the approach, prototypic models have been developed for a boiling water reactor, Peach Bottom, and a pressurized water reactor, Zion.

  16. 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

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data...

  17. 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

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data...

  18. 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

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data...

  19. 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

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data...

  20. 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

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data...

  1. Gene sequence and predicted amino acid sequence of the motA protein, a membrane-associated protein required for flagellar rotation in Escherichia coli.

    PubMed Central

    Dean, G E; Macnab, R M; Stader, J; Matsumura, P; Burks, C

    1984-01-01

    The motA and motB gene products of Escherichia coli are integral membrane proteins necessary for flagellar rotation. We determined the DNA sequence of the region containing the motA gene and its promoter. Within this sequence, there is an open reading frame of 885 nucleotides, which with high probability (98% confidence level) meets criteria for a coding sequence. The 295-residue amino acid translation product had a molecular weight of 31,974, in good agreement with the value determined experimentally by gel electrophoresis. The amino acid sequence, which was quite hydrophobic, was subjected to a theoretical analysis designed to predict membrane-spanning alpha-helical segments of integral membrane proteins; four such hydrophobic helices were predicted by this treatment. Additional amphipathic helices may also be present. A remarkable feature of the sequence is the existence of two segments of high uncompensated charge density, one positive and the other negative. Possible organization of the protein in the membrane is discussed. Asymmetry in the amino acid composition of translated DNA sequences was used to distinguish between two possible initiation codons. The use of this method as a criterion for authentication of coding regions is described briefly in an Appendix. PMID:6090403

  2. Analysis of amino acid sequence variations and immunoglobulin E-binding epitopes of German cockroach tropomyosin.

    PubMed

    Jeong, Kyoung Yong; Lee, Jongweon; Lee, In-Yong; Ree, Han-Il; Hong, Chein-Soo; Yong, Tai-Soon

    2004-09-01

    The allergenicities of tropomyosins from different organisms have been reported to vary. The cDNA encoding German cockroach tropomyosin (Bla g 7) was isolated, expressed, and characterized previously. In the present study, the amino acid sequence variations in German cockroach tropomyosin were analyzed in order to investigate its influence on allergenicity. We also undertook the identification of immunodominant peptides containing immunoglobulin E (IgE) epitopes which may facilitate the development of diagnostic and immunotherapeutic strategies based on the recombinant proteins. Two-dimensional gel electrophoresis and immunoblot analysis with mouse anti-recombinant German cockroach tropomyosin serum was performed to investigate the isoforms at the protein level. Reverse transcriptase PCR (RT-PCR) was applied to examine the sequence diversity. Eleven different variants of the deduced amino acid sequences were identified by RT-PCR. German cockroach tropomyosin has only minor sequence variations that did not seem to affect its allergenicity significantly. These results support the molecular basis underlying the cross-reactivities of arthropod tropomyosins. Recombinant fragments were also generated by PCR, and IgE-binding epitopes were assessed by enzyme-linked immunosorbent assay. Sera from seven patients revealed heterogeneous IgE-binding responses. This study demonstrates multiple IgE-binding epitope regions in a single molecule, suggesting that full-length tropomyosin should be used for the development of diagnostic and therapeutic reagents.

  3. 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.

  4. 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

  5. Human retroviruses and aids, 1992. A compilation and analysis of nucleic acid and amino acid sequences

    SciTech Connect

    Myers, G.; Korber, B.; Berzofsky, J.A.; Pavlakis, G.N.; Smith, R.F.

    1992-10-01

    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) HIV and SIV Nucleotide Sequences; (H) 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. 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 below of the parts of the compendium, the user should read the individual introductions for each part.

  6. Completion of the amino acid sequence of the alpha 1 chain from type I calf skin collagen. Amino acid sequence of alpha 1(I)B8.

    PubMed Central

    Glanville, R W; Breitkreutz, D; Meitinger, M; Fietzek, P P

    1983-01-01

    The complete amino acid sequence of the 279-residue CNBr peptide CB8 from the alpha 1 chain of type I calf skin collagen is presented. It was determined by sequencing overlapping fragments of CB8 produced by Staphylococcus aureus V8 proteinase, trypsin, Endoproteinase Arg-C and hydroxylamine. Tryptic cleavages were also made specific for lysine by blocking arginine residues with cyclohexane-1,2-dione. This completes the amino acid sequence analysis of the 1054-residues-long alpha (I) chain of calf skin collagen. PMID:6354180

  7. Advanced accident sequence precursor analysis level 1 models

    SciTech Connect

    Sattison, M.B.; Thatcher, T.A.; Knudsen, J.K.; Schroeder, J.A.; Siu, N.O.

    1996-03-01

    INEL has been involved in the development of plant-specific Accident Sequence Precursor (ASP) models for the past two years. These models were developed for use with the SAPHIRE suite of PRA computer codes. They contained event tree/linked fault tree Level 1 risk models for the following initiating events: general transient, loss-of-offsite-power, steam generator tube rupture, small loss-of-coolant-accident, and anticipated transient without scram. Early in 1995 the ASP models were revised based on review comments from the NRC and an independent peer review. These models were released as Revision 1. The Office of Nuclear Regulatory Research has sponsored several projects at the INEL this fiscal year to further enhance the capabilities of the ASP models. Revision 2 models incorporates more detailed plant information into the models concerning plant response to station blackout conditions, information on battery life, and other unique features gleaned from an Office of Nuclear Reactor Regulation quick review of the Individual Plant Examination submittals. These models are currently being delivered to the NRC as they are completed. A related project is a feasibility study and model development of low power/shutdown (LP/SD) and external event extensions to the ASP models. This project will establish criteria for selection of LP/SD and external initiator operational events for analysis within the ASP program. Prototype models for each pertinent initiating event (loss of shutdown cooling, loss of inventory control, fire, flood, seismic, etc.) will be developed. A third project concerns development of enhancements to SAPHIRE. In relation to the ASP program, a new SAPHIRE module, GEM, was developed as a specific user interface for performing ASP evaluations. This module greatly simplifies the analysis process for determining the conditional core damage probability for a given combination of initiating events and equipment failures or degradations.

  8. An Integrated Sequence-Structure Database incorporating matching mRNA sequence, amino acid sequence and protein three-dimensional structure data.

    PubMed Central

    Adzhubei, I A; Adzhubei, A A; Neidle, S

    1998-01-01

    We have constructed a non-homologous database, termed the Integrated Sequence-Structure Database (ISSD) which comprises the coding sequences of genes, amino acid sequences of the corresponding proteins, their secondary structure and straight phi,psi angles assignments, and polypeptide backbone coordinates. Each protein entry in the database holds the alignment of nucleotide sequence, amino acid sequence and the PDB three-dimensional structure data. The nucleotide and amino acid sequences for each entry are selected on the basis of exact matches of the source organism and cell environment. The current version 1.0 of ISSD is available on the WWW at http://www.protein.bio.msu.su/issd/ and includes 107 non-homologous mammalian proteins, of which 80 are human proteins. The database has been used by us for the analysis of synonymous codon usage patterns in mRNA sequences showing their correlation with the three-dimensional structure features in the encoded proteins. Possible ISSD applications include optimisation of protein expression, improvement of the protein structure prediction accuracy, and analysis of evolutionary aspects of the nucleotide sequence-protein structure relationship. PMID:9399866

  9. 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.

  10. Recondensation level of repetitive sequences in the plant protoplast nucleus is limited by oxidative stress

    PubMed Central

    Ondřej, Vladan; Navrátilová, Božena; Protivánková, Iva; Piterková, Jana; Sedlářová, Michaela; Luhová, Lenka; Lebeda, Aleš

    2010-01-01

    Protoplast cultures are remarkable examples of plant cell dedifferentiation. The state of dedifferentiation is evidenced by changes in cell morphology, genome organization, as well as by the capability of protoplasts to differentiate into multiple types of cells (depending on the type of the stimulus applied). The first change in the genome structure is connected with large-scale chromatin decondensation, affecting chromocentres involving various types of these repetitive sequences. This paper describes not only the de- and recondensation of satellite DNA type I and 5S rDNA repetitive sequences, but it also compares the recondensation level of chromatin with the levels of oxidative stress which were decreased by using an antioxidant, as well as the capabilities of the antioxidative systems within protoplasts, during the first 72 h of their culture. It is demonstrated that the treatment of protoplasts with ascorbic acid not only decreased the level of oxidative stress but also positively stimulated the expression of the ascorbate peroxidase and catalase. It also led to a greater recondensation of the chromatin (when compared to the untreated protoplasts); in addition, it supported cell proliferation. It is concluded that large-scale genome relaxation is more directly connected with oxidative stress than with large changes in the expression of genes; and further, that its recondensation is related to the start of (as well as the level of) protection by the antioxidative systems. PMID:20363868

  11. 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.

  12. Synovial fluid lactic acid levels in septic arthritis.

    PubMed

    Riley, T V

    1981-01-01

    Synovial fluid lactic acid estimations were carried out on 50 samples by gas liquid chromatography. Specimens from 4 patients with bacteria arthritis, other than gonococcal, had a mean lactic acid concentration of 215 mg/dl. One patient with gonococcal arthritis had a synovial fluid lactic acid of 30 mg/dl. Forty-one patients with inflammatory arthritis and 4 patients with degenerative arthritis had mean synovial fluid lactic acid levels of 27 and 23 mg/dl respectively. The estimation of synovial fluid lactic acid is reliable in differentiating septic arthritis from inflammatory and degenerative arthritis except when the infecting organism is NEisseria gonorrhoeae.

  13. Cerebrospinal fluid ascorbic acid levels in neurological disorders.

    PubMed

    Brau, R H; García-Castiñeiras, S; Rifkinson, N

    1984-02-01

    The ascorbic acid/dehydroascorbic acid system was analyzed in the cerebrospinal fluid (CSF) of 41 patients with different neurological disorders. The chi-square test of covariance analysis revealed in this sample significant differences in the CSF levels of total ascorbic acid when patients were classified by diagnostic categories. The population analyzed contained a group of 18 patients (back pain/sciatica group) in whom no overt neurological abnormalities were disclosed upon evaluation. Taking the CSF levels of total ascorbic acid and dehydroascorbic acid in these patients as the reference (3.57 +/- 0.87 (SD)/100 ml and 0.53 +/- 0.19 mg/100 ml, respectively), it was found that head-traumatized patients showed a significant reduction in the concentration of total ascorbic acid in the CSF. CSF ascorbic acid levels were also significantly lower in patients with increased intracranial pressure (noninfected hydrocephalus group) and in patients with cerebral tumors. Although the CSF concentration of dehydroascorbic acid did not correspondingly increase over the reference values in these three groups of patients, the tendency existed for dehydroascorbic acid to represent in them a higher percentage of total ascorbic acid. After examining different alternatives, it is concluded that the hypothesis of free radical damage to the central nervous system after certain types of injury (trauma, ischemia, and tumors) may provide a satisfactory explanation of our findings. A rationale for the use of vitamin C in the management of some neurological patients is also derived from this work.

  14. High-level phylogeny of the Coleoptera inferred with mitochondrial genome sequences.

    PubMed

    Yuan, Ming-Long; Zhang, Qi-Lin; Zhang, Li; Guo, Zhong-Long; Liu, Yong-Jian; Shen, Yu-Ying; Shao, Renfu

    2016-11-01

    The Coleoptera (beetles) exhibits tremendous morphological, ecological, and behavioral diversity. To better understand the phylogenetics and evolution of beetles, we sequenced three complete mitogenomes from two families (Cleridae and Meloidae), which share conserved mitogenomic features with other completely sequenced beetles. We assessed the influence of six datasets and three inference methods on topology and nodal support within the Coleoptera. We found that both Bayesian inference and maximum likelihood with homogeneous-site models were greatly affected by nucleotide compositional heterogeneity, while the heterogeneous-site mixture model in PhyloBayes could provide better phylogenetic signals for the Coleoptera. The amino acid dataset generated more reliable tree topology at the higher taxonomic levels (i.e. suborders and series), where the inclusion of rRNA genes and the third positions of protein-coding genes improved phylogenetic inference at the superfamily level, especially under a heterogeneous-site model. We recovered the suborder relationships as (Archostemata+Adephaga)+(Myxophaga+Polyphaga). The series relationships within Polyphaga were recovered as (Scirtiformia+(Elateriformia+((Bostrichiformia+Scarabaeiformia+Staphyliniformia)+Cucujiformia))). All superfamilies within Cucujiformia were recovered as monophyletic. We obtained a cucujiform phylogeny of (Cleroidea+(Coccinelloidea+((Lymexyloidea+Tenebrionoidea)+(Cucujoidea+(Chrysomeloidea+Curculionoidea))))). This study showed that although tree topologies were sensitive to data types and inference methods, mitogenomic data could provide useful information for resolving the Coleoptera phylogeny at various taxonomic levels by using suitable datasets and heterogeneous-site models.

  15. Maternal folic acid supplementation to dams on marginal protein level alters brain fatty acid levels of their adult offspring.

    PubMed

    Rao, Shobha; Joshi, Sadhana; Kale, Anvita; Hegde, Mahabaleshwar; Mahadik, Sahebarao

    2006-05-01

    Studies on fetal programming of adult diseases have highlighted the importance of maternal nutrition during pregnancy. Folic acid and long-chain essential polyunsaturated fatty acids (LC-PUFAs) have independent effects on fetal growth. However, folic acid effects may also involve alteration of LC-PUFA metabolism. Because marginal deficiency of LC-PUFAs during critical periods of brain growth and development is associated with risks for adult diseases, it is highly relevant to investigate how maternal supplementation of such nutrients can alter brain fatty acid levels. We examined the impact of folic acid supplementation, conventionally used in maternal intervention, on brain essential fatty acid levels and plasma corticosterone concentrations in adult offspring at 11 months of age. Pregnant female rats from 4 groups (6 in each) were fed with casein diets either with 18 g protein/100 g diet (control diet) or treatment diets that were marginal in protein (MP), such as 12 g protein/100 g diet supplemented with 8 mg folic acid (FAS/MP), 12 g protein/100 g diet without folic acid (FAD/MP), or 12 g protein/100 g diet (MP) with 2 mg folic acid. Pups were weaned to a standard laboratory diet with 18 g protein/100 g diet. All male adult offspring in the FAS/MP group showed lower docosahexaenoic acid (P<.05) as compared with control adult offspring (6.04+/-2.28 vs 10.33+/-0.86 g/100 g fatty acids) and higher n-6/n-3 ratio (P<.05). Docosahexaenoic acid levels in FAS/MP adult offspring were also lower (P<.05) when compared with the MP group. Plasma corticosterone concentrations were higher (P<.05) in male adult offspring from the FAS/MP group compared with control as well as the MP adult offspring. Results suggest that maternal folic acid supplementation at MP intake decreased brain docosahexaenoic acid levels probably involving corticosterone increase.

  16. Trichomonas vaginalis acidic phospholipase A2: isolation and partial amino acid sequence.

    PubMed

    Escobedo-Guajardo, Brenda L; González-Salazar, Francisco; Palacios-Corona, Rebeca; Torres de la Cruz, Víctor M; Morales-Vallarta, Mario; Mata-Cárdenas, Benito D; Garza-González, Jesús N; Rivera-Silva, Gerardo; Vargas-Villarreal, Javier

    2013-12-01

    Sexually transmitted diseases are a major cause of acute disease worldwide, and trichomoniasis is the most common and curable disease, generating more than 170 million cases annually worldwide. Trichomonas vaginalis is the causal agent of trichomoniasis and has the ability to destroy in vitro cell monolayers of the vaginal mucosa, where the phospholipases A2 (PLA2) have been reported as potential virulence factors. These enzymes have been partially characterized from the subcellular fraction S30 of pathogenic T. vaginalis strains. The main objective of this study was to purify a phospholipase A2 from T. vaginalis, make a partial characterization, obtain a partial amino acid sequence, and determine its enzymatic participation as hemolytic factor causing lysis of erythrocytes. Trichomonas S30, RF30 and UFF30 sub-fractions from GT-15 strain have the capacity to hydrolyze [2-(14)C-PA]-PC at pH 6.0. Proteins from the UFF30 sub-fraction were separated by affinity chromatography into two eluted fractions with detectable PLA A2 activity. The EDTA-eluted fraction was analyzed by HPLC using on-line HPLC-tandem mass spectrometry and two protein peaks were observed at 8.2 and 13 kDa. Peptide sequences were identified from the proteins present in the eluted EDTA UFF30 fraction; bioinformatic analysis using Protein Link Global Server charged with T. vaginalis protein database suggests that eluted peptides correspond a putative ubiquitin protein in the 8.2 kDa fraction and a phospholipase preserved in the 13 kDa fraction. The EDTA-eluted fraction hydrolyzed [2-(14)C-PA]-PC lyses erythrocytes from Sprague-Dawley in a time and dose-dependent manner. The acidic hemolytic activity decreased by 84% with the addition of 100 μM of Rosenthal's inhibitor.

  17. Uric Acid Levels in Normotensive Children of Hypertensive Parents

    PubMed Central

    Yildirim, Ali; Keles, Fatma; Kosger, Pelin; Ozdemir, Gokmen; Ucar, Birsen; Kilic, Zubeyir

    2015-01-01

    This study evaluated uric acid concentrations in normotensive children of parents with hypertension. Eighty normotensive children from families with and without a history of essential hypertension were included. Concentrations of lipid parameters and uric acid were compared. Demographic and anthropometric characteristics were similar in the groups. Systolic and diastolic blood pressure were higher in the normotensive children of parents with hypertension without statistically significant difference (P > 0.05). Uric acid concentrations were higher in the normotensive children of parents with hypertension (4.61 versus 3.57 mg/dL, P < 0.01). Total cholesterol and triglyceride concentrations were similar in the two groups. Systolic and diastolic blood pressure were significantly higher in control children aged >10 years (P < 0.01). Uric acid levels were significantly higher in all children with more pronounced difference after age 10 of years (P < 0.001). Positive correlations were found between the level of serum uric acid and age, body weight, body mass index, and systolic and diastolic blood pressure in the normotensive children of parents. The higher uric acid levels in the normotensive children of hypertensive parents suggest that uric acid may be a predeterminant of hypertension. Monitoring of uric acid levels in these children may allow for prevention or earlier treatment of future hypertension. PMID:26464873

  18. 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.

  19. 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.

  20. Deoxyribonucleic acid sequence of araBAD promoter mutants of Escherichia coli.

    PubMed

    Horwitz, A H; Morandi, C; Wilcox, G

    1980-05-01

    The controlling site region for the araBAD operon is defined, in part, by two classes of cis-acting constitutive mutations. The aralc mutations allow low-level constitutive expression of ara-BAD in the absence of the positive regulatory protein coded for by the araC gene, whereas the araXc mutations allow expression of araBAD in the absence of the cyclic adenosine monophosphate receptor protein. Six independently isolated aralc mutations and three independently isolated araXc mutations were cloned onto the plasmid pBR322 using in vitro recombinant deoxyribonucleic acid techniques and in vivo recombination between plasmid and chromosomal deoxyribonucleic acid. The location of these mutations was determined by deoxyribonucleic acid sequence analysis. All of the aralc mutations occurred at position -35 within the araBAD promoter (+1 = messenger ribonucleic acid start for araBAD) and resulted from an AT leads to GC transition. All of the araXc mutations occurred at position -10 within the araBAD promoter and resulted from a GC leads to AT transition. Models are presented to explain the mode of action of the aralc and araXc mutations.

  1. 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.

  2. The amino acid sequences of the Fd fragments of two human γ heavy chains

    PubMed Central

    Press, E. M.; Hogg, N. M.

    1970-01-01

    The amino acid sequences of the Fd fragments of two human pathological immunoglobulins of the immunoglobulin G1 class are reported. Comparison of the two sequences shows that the heavy-chain variable regions are similar in length to those of the light chains. The existence of heavy chain variable region subgroups is also deduced, from a comparison of these two sequences with those of another γ 1 chain, Eu, a μ chain, Ou, and the partial sequence of a fourth γ 1 chain, Ste. Carbohydrate has been found to be linked to an aspartic acid residue in the variable region of one of the γ 1 chains, Cor. PMID:5449120

  3. [Hydroxycinnamic acid levels of various batches from mugwort flowering tops].

    PubMed

    Fraisse, D; Carnat, A; Carnat, A-P; Guédon, D; Lamaison, J-L

    2003-07-01

    Dried flowering tops of 24 harvested batches (Artemisia vulgaris: 13; Artemisia verlotiorum: 11) and 12 batches of mugwort from commercial origin were examined. The levels of principal compounds averaged respectively: total hydroxycinnamic acids 6.09; 10.29 and 9.13%, chlorogenic acid 0.79; 2.05 and 1.35%, 1,5-dicaffeoylquinic acid 0.51; 4.01 and 1.25%, 3,5-dicaffeoylquinic acid 2.21; 1.25 and 2.60%. Specifications were discussed for an European Pharmacopoeial monography.

  4. 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

  5. The amino acid sequence of goat beta-lactoglobulin.

    PubMed

    Préaux, G; Braunitzer, G; Schrank, B; Stangl, A

    1979-11-01

    The isolation of beta-lactoglobulin from milk of the goat is described. The purified protein was checked for purity and has been characterized by its gross composition and end groups. The native or the modified protein was then degraded by tryptic and cyanogen bromide cleavage. The cleavage products were isolated and sequenced in the sequenator using a Quadrol and propyne program. These data provide the complete sequence of beta-lactoglobulin of the goat. The results are discussed and compared particularly with bovine beta-lactoglobulin components AB. Some biological aspects are described.

  6. Layered materials with coexisting acidic and basic sites for catalytic one-pot reaction sequences.

    PubMed

    Motokura, Ken; Tada, Mizuki; Iwasawa, Yasuhiro

    2009-06-17

    Acidic montmorillonite-immobilized primary amines (H-mont-NH(2)) were found to be excellent acid-base bifunctional catalysts for one-pot reaction sequences, which are the first materials with coexisting acid and base sites active for acid-base tamdem reactions. For example, tandem deacetalization-Knoevenagel condensation proceeded successfully with the H-mont-NH(2), affording the corresponding condensation product in a quantitative yield. The acidity of the H-mont-NH(2) was strongly influenced by the preparation solvent, and the base-catalyzed reactions were enhanced by interlayer acid sites.

  7. Synthesis of gamma,delta-unsaturated glycolic acids via sequenced brook and Ireland--claisen rearrangements.

    PubMed

    Schmitt, Daniel C; Johnson, Jeffrey S

    2010-03-05

    Organozinc, -magnesium, and -lithium nucleophiles initiate a Brook/Ireland-Claisen rearrangement sequence of allylic silyl glyoxylates resulting in the formation of gamma,delta-unsaturated alpha-silyloxy acids.

  8. 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)

  9. 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.

  10. Genome sequence of the acid-tolerant strain Rhizobium sp. LPU83.

    PubMed

    Wibberg, Daniel; Tejerizo, Gonzalo Torres; Del Papa, María Florencia; Martini, Carla; Pühler, Alfred; Lagares, Antonio; Schlüter, Andreas; Pistorio, Mariano

    2014-04-20

    Rhizobia are important members of the soil microbiome since they enter into nitrogen-fixing symbiosis with different legume host plants. Rhizobium sp. LPU83 is an acid-tolerant Rhizobium strain featuring a broad-host-range. However, it is ineffective in nitrogen fixation. Here, the improved draft genome sequence of this strain is reported. Genome sequence information provides the basis for analysis of its acid tolerance, symbiotic properties and taxonomic classification.

  11. 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).

  12. Single-chain structure of human ceruloplasmin: the complete amino acid sequence of the whole molecule.

    PubMed Central

    Takahashi, N; Ortel, T L; Putnam, F W

    1984-01-01

    We have determined the amino acid sequence of the amino-terminal 67,000-dalton (67-kDa) fragment of human ceruloplasmin and have established overlapping sequences between the 67-kDa and 50-kDa fragments and between the 50-kDa and 19-kDa fragments. The 67-kDa fragment contains 480 amino acid residues and three glucosamine oligosaccharides. These results together with our previous sequence data for the 50-kDa and 19-kDa fragments complete the amino acid sequence of human ceruloplasmin. The polypeptide chain has a total of 1,046 amino acid residues (Mr 120,085) and has attachment sites for four glucosamine oligosaccharides; together these account for the total molecular mass of human ceruloplasmin (132 kDa). The sequence analysis of the peptides overlapping the fragments showed that one additional amino acid, arginine, is present between the 67-kDa and 50-kDa fragments, and another, lysine, is between the 50-kDa and 19-kDa fragments. Only two apparent sites of amino acid interchange have been identified in the polypeptide chain. Both involve a single-point interchange of glycine and lysine that would result in a difference in charge. The results of the complete sequence analysis verified that human ceruloplasmin is composed of a single polypeptide chain and that the subunit-like fragments are produced by proteolytic cleavage during purification (and possibly also in vivo). PMID:6582496

  13. Predicted Molecular Effects of Sequence Variants Link to System Level of Disease

    PubMed Central

    Bromberg, Yana; Rost, Burkhard

    2016-01-01

    Developments in experimental and computational biology are advancing our understanding of how protein sequence variation impacts molecular protein function. However, the leap from the micro level of molecular function to the macro level of the whole organism, e.g. disease, remains barred. Here, we present new results emphasizing earlier work that suggested some links from molecular function to disease. We focused on non-synonymous single nucleotide variants, also referred to as single amino acid variants (SAVs). Building upon OMIA (Online Mendelian Inheritance in Animals), we introduced a curated set of 117 disease-causing SAVs in animals. Methods optimized to capture effects upon molecular function often correctly predict human (OMIM) and animal (OMIA) Mendelian disease-causing variants. We also predicted effects of human disease-causing variants in the mouse model, i.e. we put OMIM SAVs into mouse orthologs. Overall, fewer variants were predicted with effect in the model organism than in the original organism. Our results, along with other recent studies, demonstrate that predictions of molecular effects capture some important aspects of disease. Thus, in silico methods focusing on the micro level of molecular function can help to understand the macro system level of disease. PMID:27536940

  14. Multiple Genome Sequences of Important Beer-Spoiling Lactic Acid Bacteria

    PubMed Central

    Geissler, Andreas J.; Vogel, Rudi F.

    2016-01-01

    Seven strains of important beer-spoiling lactic acid bacteria were sequenced using single-molecule real-time sequencing. Complete genomes were obtained for strains of Lactobacillus paracollinoides, Lactobacillus lindneri, and Pediococcus claussenii. The analysis of these genomes emphasizes the role of plasmids as the genomic foundation of beer-spoiling ability. PMID:27795248

  15. Nucleic acid sequence of an internal image-bearing monoclonal anti-idiotype and its comparison to the sequence of the external antigen.

    PubMed Central

    Bruck, C; Co, M S; Slaoui, M; Gaulton, G N; Smith, T; Fields, B N; Mullins, J I; Greene, M I

    1986-01-01

    The monoclonal anti-idiotypic antibody (mAb2) 87.92.6 directed against the 9B.G5 antibody specific for the virus neutralizing epitope on the mammalian reovirus type 3 hemagglutinin was previously demonstrated to express an internal image of the receptor binding epitope of the reovirus type 3. Furthermore, this mAb2 has autoimmune reactivity to the cell surface receptor of the reovirus. The nucleotide and deduced amino acid sequences of the 87.92.6 mAb2 heavy and light chains are described in this report. The sequence analysis reveals that the same heavy chain variable and joining (VH and JH) gene segments are used by the 87.92.6 anti-idiotypic mAb2 and by the dominant idiotypes of the BALB/c anti-GAT (cGAT) and anti-NP (NPa) responses. [GAT; random polymer that is 60% glutamic acid, 30% alanine, and 10% tyrosine. NP; (4-hydroxy-3-nitrophenyl)-acetyl.] Despite extensive homology at the level of the heavy chain variable regions, the NPa positive BALB/c anti-NP monoclonal antibody 17.2.25 binds neither 9B.G5 nor the cellular receptor for the hemagglutinin. Amino acid sequence comparison between the viral hemagglutinin and the 87.92.6 mAb2 light chain "internal image," reveals an area of significant homology indicating that antigen mimicry by antibodies may be achieved by sharing primary structure. PMID:2428036

  16. Identification of tropomyosins as major allergens in antarctic krill and mantis shrimp and their amino acid sequence characteristics.

    PubMed

    Motoyama, Kanna; Suma, Yota; Ishizaki, Shoichiro; Nagashima, Yuji; Lu, Ying; Ushio, Hideki; Shiomi, Kazuo

    2008-01-01

    Tropomyosin represents a major allergen of decapod crustaceans such as shrimps and crabs, and its highly conserved amino acid sequence (>90% identity) is a molecular basis of the immunoglobulin E (IgE) cross-reactivity among decapods. At present, however, little information is available about allergens in edible crustaceans other than decapods. In this study, the major allergen in two species of edible crustaceans, Antarctic krill Euphausia superba and mantis shrimp Oratosquilla oratoria that are taxonomically distinct from decapods, was demonstrated to be tropomyosin by IgE-immunoblotting using patient sera. The cross-reactivity of the tropomyosins from both species with decapod tropomyosins was also confirmed by inhibition IgE immunoblotting. Sequences of the tropomyosins from both species were determined by complementary deoxyribonucleic acid cloning. The mantis shrimp tropomyosin has high sequence identity (>90% identity) with decapod tropomyosins, especially with fast-type tropomyosins. On the other hand, the Antarctic krill tropomyosin is characterized by diverse alterations in region 13-42, the amino acid sequence of which is highly conserved for decapod tropomyosins, and hence, it shares somewhat lower sequence identity (82.4-89.8% identity) with decapod tropomyosins than the mantis shrimp tropomyosin. Quantification by enzyme-linked immunosorbent assay revealed that Antarctic krill contains tropomyosin at almost the same level as decapods, suggesting that its allergenicity is equivalent to decapods. However, mantis shrimp was assumed to be substantially not allergenic because of the extremely low content of tropomyosin.

  17. PASTA: Ultra-Large Multiple Sequence Alignment for Nucleotide and Amino-Acid Sequences.

    PubMed

    Mirarab, Siavash; Nguyen, Nam; Guo, Sheng; Wang, Li-San; Kim, Junhyong; Warnow, Tandy

    2015-05-01

    We introduce PASTA, a new multiple sequence alignment algorithm. PASTA uses a new technique to produce an alignment given a guide tree that enables it to be both highly scalable and very accurate. We present a study on biological and simulated data with up to 200,000 sequences, showing that PASTA produces highly accurate alignments, improving on the accuracy and scalability of the leading alignment methods (including SATé). We also show that trees estimated on PASTA alignments are highly accurate--slightly better than SATé trees, but with substantial improvements relative to other methods. Finally, PASTA is faster than SATé, highly parallelizable, and requires relatively little memory.

  18. Serum uric acid levels and cardiovascular disease: the Gordian knot

    PubMed Central

    Tugores, Antonio; Rodríguez-González, Fayna

    2016-01-01

    Hyperuricemia is defined as serum uric acid level of more than 7 mg/dL and blood levels of uric acid are causally associated with gout, as implicated by evidence from randomized clinical trials using urate lowering therapies. Uric acid as a cardiovascular risk factor often accompanies metabolic syndrome, hypertension, diabetes, dyslipidemia, chronic renal disease, and obesity. Despite the association of hyperuricemia with cardiovascular risk factors, it has remained controversial as to whether uric acid is an independent predictor of cardiovascular disease. To settle this issue, and in the absence of large randomized controlled trials, Mendelian randomization analysis in which the exposure is defined based on the presence or absence of a specific allele that influences a risk factor of interest have tried to shed light on this. PMID:28066631

  19. 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

  20. SETG: Nucleic Acid Extraction and Sequencing for In Situ Life Detection on Mars

    NASA Astrophysics Data System (ADS)

    Mojarro, A.; Hachey, J.; Tani, J.; Smith, A.; Bhattaru, S. A.; Pontefract, A.; Doebler, R.; Brown, M.; Ruvkun, G.; Zuber, M. T.; Carr, C. E.

    2016-10-01

    We are developing an integrated nucleic acid extraction and sequencing instrument: the Search for Extra-Terrestrial Genomes (SETG) for in situ life detection on Mars. Our goals are to identify related or unrelated nucleic acid-based life on Mars.

  1. Draft Genome Sequence of Cyanobacterium sp. Strain IPPAS B-1200 with a Unique Fatty Acid Composition

    PubMed Central

    Starikov, Alexander Y.; Usserbaeva, Aizhan A.; Sinetova, Maria A.; Sarsekeyeva, Fariza K.; Zayadan, Bolatkhan K.; Ustinova, Vera V.; Kupriyanova, Elena V.; Los, Dmitry A.

    2016-01-01

    Here, we report the draft genome of Cyanobacterium sp. IPPAS strain B-1200, isolated from Lake Balkhash, Kazakhstan, and characterized by the unique fatty acid composition of its membrane lipids, which are enriched with myristic and myristoleic acids. The approximate genome size is 3.4 Mb, and the predicted number of coding sequences is 3,119. PMID:27856596

  2. Sequencing and computational analysis of complete genome sequences of Citrus yellow mosaic badna virus from acid lime and pummelo.

    PubMed

    Borah, Basanta K; Johnson, A M Anthony; Sai Gopal, D V R; Dasgupta, Indranil

    2009-08-01

    Citrus yellow mosaic badna virus (CMBV), a member of the Family Caulimoviridae, Genus Badnavirus, is the causative agent of Citrus mosaic disease in India. Although the virus has been detected in several citrus species, only two full-length genomes, one each from Sweet orange and Rangpur lime, are available in publicly accessible databases. In order to obtain a better understanding of the genetic variability of the virus in other citrus mosaic-affected citrus species, we performed the cloning and sequence analysis of complete genomes of CMBV from two additional citrus species, Acid lime and Pummelo. We show that CMBV genomes from the two hosts share high homology with previously reported CMBV sequences and hence conclude that the new isolates represent variants of the virus present in these species. Based on in silico sequence analysis, we predict the possible function of the protein encoded by one of the five ORFs.

  3. Parvalbumins from coelacanth muscle. III. Amino acid sequence of the major component.

    PubMed

    Jauregui-Adell, J; Pechere, J F

    1978-09-26

    The primary structure of the major parvalbumin (pI = 4.52) from coelacanth muscle (Latimeria chalumnae) has been determined. Sequence analysis of the tryptic peptides, in some cases obtained with beta-trypsin, accounts for the total amino acid content of the protein. Chymotryptic peptides provide appropriate sequence overlaps, to complete the localization of the tryptic peptides. Examination of the amino acid sequence of this protein shows the typical structure of a beta-parvalbumin. Its position in the dendrogram of related calcium-binding proteins corresponds to that usually accepted for crossopterygians.

  4. Independence of sialic acid levels in normal and malignant growth.

    PubMed

    Khadapkar, S V; Sheth, N A; Bhide, S V

    1975-06-01

    Sialic acid content in breast or tumor tissue and serum of mouse strains that are either susceptible or resistant to breast cancer was measured at various age periods. Sialic acid content was also studied in normal lung tissue and in lung adenoma and hepatoma. Sialic acid levels during nonmalignant growth of a tissue were measured in breast tissue during pregnancy and lactation, and in regenerating liver, as well as in newborn and postnatal liver. The sialic acid content, when expressed per mg of protein, increased in mammary tumor, lung adenoma, and hepatoma. It also increased in nonmalignant growth of breast tissue during pregnancy and lactation and of regenerating liver and postnatal liver. Increase in sialic acid per mg DNA was observed only in lung tumors, regenerating liver, and postnatal liver. It appears that the changes in sialic acid level are independent of the normal or malignant growth of a tissue and that these changes might be the function of the parameter used to express the sialic acid values, i.e., either the DNA content or protein content of a given tissue.

  5. New Insights into Poly(Lactic-co-glycolic acid) Microstructure: Using Repeating Sequence Copolymers to Decipher Complex NMR and Thermal Behavior

    PubMed Central

    Stayshich, Ryan M.; Meyer, Tara Y.

    2012-01-01

    Sequence, which Nature uses to spectacular advantage, has not been fully exploited in synthetic copolymers. To investigate the effect of sequence and stereosequence on the physical properties of copolymers a family of complex isotactic, syndiotactic and atactic repeating sequence poly(lactic-co-glycolic acid) copolymers (RSC PLGAs) were prepared and their NMR and thermal behavior was studied. The unique suitability of polymers prepared from the bioassimilable lactic and glycolic acid monomers for biomedical applications makes them ideal candidates for this type of sequence engineering. Polymers with repeating units of LG, GLG and LLG (L = lactic, G = glycolic) with controlled and varied tacticities were synthesized by assembly of sequence specific, stereopure dimeric, trimeric and hexameric segmer units. Specifically labeled deuterated lactic and glycolic acid segmers were likewise prepared and polymerized. Molecular weights for the copolymers ranged from Mn = 12-40 kDa by size exclusion chromatography in THF. Although the effects of sequence-influenced solution conformation were visible in all resonances of the 1H and 13C NMR spectra, the diastereotopic methylene resonances in the 1H NMR (CDCl3) for the glycolic units of the copolymers proved most sensitive. An octad level of resolution, which corresponds to an astounding 31-atom distance between the most separated stereocenters, was observed in some mixed sequence polymers. Importantly, the level of sensitivity of a particular NMR resonance to small differences in sequence was found to depend on the sequence itself. Thermal properties were also correlated with sequence. PMID:20681726

  6. Amino acid sequence of anionic peroxidase from the windmill palm tree Trachycarpus fortunei.

    PubMed

    Baker, Margaret R; Zhao, Hongwei; Sakharov, Ivan Yu; Li, Qing X

    2014-12-10

    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.

  7. Plasma amino acid response to graded levels of escape protein.

    PubMed

    Gibb, D J; Klopfenstein, T J; Britton, R A; Lewis, A J

    1992-09-01

    A trial was conducted to examine the potential of using plasma amino acid responses to graded levels of escape protein to determine limiting amino acids in cattle. Growing calves (n = 120; mean BW = 220 +/- 21 kg) were fed a basal diet of corncob:sorghum silage (61:39) and were individually supplemented with distillers' dried grains (DDG), heat-damaged DDG (H-DDG), feather meal (FTH), or urea. The urea supplement was mixed with DDG and H-DDG to allow 0, 20, 35, 50, 65, or 80% of the supplemental CP to come from distillers' protein and maintain an 11.5% CP diet. Urea supplement was mixed with FTH to allow 0, 22, 39, 56, 73, or 90% of the supplemental CP to come from FTH. Dietary CP ranged from 11.5% at the 0% level to 17.3% at the 90% level. Plasma concentration of most essential plasma amino acids responded (P less than .10) linearly and(or) quadratically to increased escape protein. The broken-line response of plasma methionine at low DDG intake suggested that methionine was limiting at low levels of escape protein. An initial decrease followed by a plateau fit by a broken line indicated that histidine became limiting in FTH diets, and lysine eventually became limiting for DDG, H-DDG, and FTH diets before maximum BW gain was reached. Results indicate that plasma amino acid responses may identify amino acids that become limiting with increasing escape protein.

  8. Amino acid sequence of homologous rat atrial peptides: natriuretic activity of native and synthetic forms.

    PubMed Central

    Seidah, N G; Lazure, C; Chrétien, M; Thibault, G; Garcia, R; Cantin, M; Genest, J; Nutt, R F; Brady, S F; Lyle, T A

    1984-01-01

    A substance called atrial natriuretic factor (ANF), localized in secretory granules of atrial cardiocytes, was isolated as four homologous natriuretic peptides from homogenates of rat atria. The complete sequence of the longest form showed that it is composed of 33 amino acids. The three other shorter forms (2-33, 3-33, and 8-33) represent amino-terminally truncated versions of the 33 amino acid parent molecule as shown by analysis of sequence, amino acid composition, or both. The proposed primary structure agrees entirely with the amino acid composition and reveals no significant sequence homology with any known protein or segment of protein. The short form ANF-(8-33) was synthesized by a multi-fragment condensation approach and the synthetic product was shown to exhibit specific activity comparable to that of the natural ANF-(3-33). PMID:6232612

  9. Nucleotide and deduced amino acid sequences of a new subtilisin from an alkaliphilic Bacillus isolate.

    PubMed

    Saeki, Katsuhisa; Magallones, Marietta V; Takimura, Yasushi; Hatada, Yuji; Kobayashi, Tohru; Kawai, Shuji; Ito, Susumu

    2003-10-01

    The gene for a new subtilisin from the alkaliphilic Bacillus sp. KSM-LD1 was cloned and sequenced. The open reading frame of the gene encoded a 97 amino-acid prepro-peptide plus a 307 amino-acid mature enzyme that contained a possible catalytic triad of residues, Asp32, His66, and Ser224. The deduced amino acid sequence of the mature enzyme (LD1) showed approximately 65% identity to those of subtilisins SprC and SprD from alkaliphilic Bacillus sp. LG12. The amino acid sequence identities of LD1 to those of previously reported true subtilisins and high-alkaline proteases were below 60%. LD1 was characteristically stable during incubation with surfactants and chemical oxidants. Interestingly, an oxidizable Met residue is located next to the catalytic Ser224 of the enzyme as in the cases of the oxidation-susceptible subtilisins reported to date.

  10. Shark myelin basic protein: amino acid sequence, secondary structure, and self-association.

    PubMed

    Milne, T J; Atkins, A R; Warren, J A; Auton, W P; Smith, R

    1990-09-01

    Myelin basic protein (MBP) from the Whaler shark (Carcharhinus obscurus) has been purified from acid extracts of a chloroform/methanol pellet from whole brains. The amino acid sequence of the majority of the protein has been determined and compared with the sequences of other MBPs. The shark protein has only 44% homology with the bovine protein, but, in common with other MBPs, it has basic residues distributed throughout the sequence and no extensive segments that are predicted to have an ordered secondary structure in solution. Shark MBP lacks the triproline sequence previously postulated to form a hairpin bend in the molecule. The region containing the putative consensus sequence for encephalitogenicity in the guinea pig contains several substitutions, thus accounting for the lack of activity of the shark protein. Studies of the secondary structure and self-association have shown that shark MBP possesses solution properties similar to those of the bovine protein, despite the extensive differences in primary structure.

  11. 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.

  12. Association of Renal Manifestations with Serum Uric Acid in Korean Adults with Normal Uric Acid Levels

    PubMed Central

    Jung, Dong-Hyuk; Lee, Yong-Jae; Lee, Hye-Ree; Lee, Jung-Hyun

    2010-01-01

    Several studies have reported that hyperuricemia is associated with the development of hypertension and cardiovascular disease. Increasing evidences also suggest that hyperuricemia may have a pathogenic role in the progression of renal disease. Paradoxically, uric acid is also widely accepted to have antioxidant activity in experimental studies. We aimed to investigate the association between glomerular filtration rate (GFR) and uric acid in healthy individuals with a normal serum level of uric acid. We examined renal function determined by GFR and uric acid in 3,376 subjects (1,896 men; 1,480 women; aged 20-80 yr) who underwent medical examinations at Gangnam Severance Hospital from November 2006 to June 2007. Determinants for renal function and uric acid levels were also investigated. In both men and women, GFR was negatively correlated with systolic and diastolic blood pressures, fasting plasma glucose, total cholesterol, uric acid, log transformed C reactive protein, and log transformed triglycerides. In multivariate regression analysis, total uric acid was found to be an independent factor associated with estimated GFR in both men and women. This result suggests that uric acid appears to contribute to renal impairment in subjects with normal serum level of uric acid. PMID:21165292

  13. 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

  14. An analysis of amino acid sequences surrounding archaeal glycoprotein sequons.

    PubMed

    Abu-Qarn, Mehtap; Eichler, Jerry

    2007-05-01

    Despite having provided the first example of a prokaryal glycoprotein, little is known of the rules governing the N-glycosylation process in Archaea. As in Eukarya and Bacteria, archaeal N-glycosylation takes place at the Asn residues of Asn-X-Ser/Thr sequons. Since not all sequons are utilized, it is clear that other factors, including the context in which a sequon exists, affect glycosylation efficiency. As yet, the contribution to N-glycosylation made by sequon-bordering residues and other related factors in Archaea remains unaddressed. In the following, the surroundings of Asn residues confirmed by experiment as modified were analyzed in an attempt to define sequence rules and requirements for archaeal N-glycosylation.

  15. Increased isoprostane levels in oleic acid-induced lung injury

    SciTech Connect

    Ono, Koichi; Koizumi, Tomonobu; Tsushima, Kenji; Yoshikawa, Sumiko; Yokoyama, Toshiki; Nakagawa, Rikimaru; Obata, Toru

    2009-10-16

    The present study was performed to examine a role of oxidative stress in oleic acid-induced lung injury model. Fifteen anesthetized sheep were ventilated and instrumented with a lung lymph fistula and vascular catheters for blood gas analysis and measurement of isoprostanes (8-epi prostaglandin F2{alpha}). Following stable baseline measurements, oleic acid (0.08 ml/kg) was administered and observed 4 h. Isoprostane was measured by gas chromatography mass spectrometry with the isotope dilution method. Isoprostane levels in plasma and lung lymph were significantly increased 2 h after oleic acid administration and then decreased at 4 h. The percent increases in isoprostane levels in plasma and lung lymph at 2 h were significantly correlated with deteriorated oxygenation at the same time point, respectively. These findings suggest that oxidative stress is involved in the pathogenesis of the pulmonary fat embolism-induced acute lung injury model in sheep and that the increase relates with the deteriorated oxygenation.

  16. Amino acid sequence homology between rat and human C-reactive protein.

    PubMed Central

    Taylor, J A; Bruton, C J; Anderson, J K; Mole, J E; De Beer, F C; Baltz, M L; Pepys, M B

    1984-01-01

    The rat serum protein that undergoes Ca2+-dependent binding to pneumococcal C-polysaccharide and to phosphocholine residues, and that is evidently a member of the pentraxin family of proteins by virtue of its appearance under the electron microscope, has been variously designated as rat C-reactive protein (CRP) [de Beer, Baltz, Munn, Feinstein, Taylor, Bruton, Clamp & Pepys (1982) Immunology 45, 55-70], 'phosphoryl choline-binding protein' [Nagpurkar & Mookerjea (1981) J. Biol. Chem. 256, 7440-7448] and rat serum amyloid P component (SAP) [Pontet, D'Asnieres, Gache, Escaig & Engler (1981) Biochim. Biophys. Acta 671, 202-210]. The partial amino acid sequence (45 residues) towards the C-terminus of this protein was determined, and it showed 71.7% identity with the known sequence of human CRP but only 54.3% identity with human SAP. Since human CRP and SAP are themselves approximately 50% homologous, the level of identity between the rat protein and human SAP is evidence only of membership of the pentraxin family. In contrast, the much greater resemblance to human CRP confirms that the rat C-polysaccharide-binding/phosphocholine-binding protein is in fact rat CRP. PMID:6477504

  17. Classification of mouse VK groups based on the partial amino acid sequence to the first invariant tryptophan: impact of 14 new sequences from IgG myeloma proteins.

    PubMed

    Potter, M; Newell, J B; Rudikoff, S; Haber, E

    1982-12-01

    Fourteen new VK sequences derived from BALB/c IgG myeloma proteins were determined to the first invariant tryptophan (Trp 35). These partial sequences were compared with 65 other published VK sequences using a computer program. The 79 sequences were organized according to the length of the sequence from the amino terminus to the first invariant tryptophan (Trp 35), into seven groups (33, 34, 35, 36, 39, 40 and 41aa). A distance matrix of all 79 sequences was then computed, i.e. the number of amino acid substitutions necessary to convert one sequence to another was determined. From these data a dendrogram was constructed. Most of the VK sequences fell into clusters or closely related groups. The definition of a sequence group is arbitrary but facilitates the classification of VK proteins. We used 12 substitutions as the basis for defining a sequence group based on the known number of substitutions that are found in the VK21 proteins. By this criterion there were 18 groups in the Trp 35 dendrogram. Twelve of the 14 new sequences fell into one of these sequence groups; two formed new sequence groups. Collective amino acid sequencing is still encountering new VK structures indicating more sequences will be required to attain an accurate estimate of the total number of VK groups. Updated dendrograms can be quickly generated to include newly generated sequences.

  18. Relationship between Uric Acid Level and Achievement Motivation. Final Report.

    ERIC Educational Resources Information Center

    Mueller, Ernst F.; French, John R. P., Jr.

    In an investigation of the relationship of uric acid (a metabolic end product) to achievement, this study hypothesized that a person's serum urate level (a factor often associated with gout) is positively related to achievement need as well as indicators of actual achievement. (Speed of promotion and number of yearly publications were chosen as…

  19. 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.

  20. 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.

  1. Amino acid sequence around the active-site serine residue in the acyltransferase domain of goat mammary fatty acid synthetase.

    PubMed Central

    Mikkelsen, J; Højrup, P; Rasmussen, M M; Roepstorff, P; Knudsen, J

    1985-01-01

    Goat mammary fatty acid synthetase was labelled in the acyltransferase domain by formation of O-ester intermediates by incubation with [1-14C]acetyl-CoA and [2-14C]malonyl-CoA. Tryptic-digest and CNBr-cleavage peptides were isolated and purified by high-performance reverse-phase and ion-exchange liquid chromatography. The sequences of the malonyl- and acetyl-labelled peptides were shown to be identical. The results confirm the hypothesis that both acetyl and malonyl groups are transferred to the mammalian fatty acid synthetase complex by the same transferase. The sequence is compared with those of other fatty acid synthetase transferases. PMID:3922356

  2. Ligation with nucleic acid sequence-based amplification.

    PubMed

    Ong, Carmichael; Tai, Warren; Sarma, Aartik; Opal, Steven M; Artenstein, Andrew W; Tripathi, Anubhav

    2012-01-01

    This work presents a novel method for detecting nucleic acid targets using a ligation step along with an isothermal, exponential amplification step. We use an engineered ssDNA with two variable regions on the ends, allowing us to design the probe for optimal reaction kinetics and primer binding. This two-part probe is ligated by T4 DNA Ligase only when both parts bind adjacently to the target. The assay demonstrates that the expected 72-nt RNA product appears only when the synthetic target, T4 ligase, and both probe fragments are present during the ligation step. An extraneous 38-nt RNA product also appears due to linear amplification of unligated probe (P3), but its presence does not cause a false-positive result. In addition, 40 mmol/L KCl in the final amplification mix was found to be optimal. It was also found that increasing P5 in excess of P3 helped with ligation and reduced the extraneous 38-nt RNA product. The assay was also tested with a single nucleotide polymorphism target, changing one base at the ligation site. The assay was able to yield a negative signal despite only a single-base change. Finally, using P3 and P5 with longer binding sites results in increased overall sensitivity of the reaction, showing that increasing ligation efficiency can improve the assay overall. We believe that this method can be used effectively for a number of diagnostic assays.

  3. Thin-film technology for direct visual detection of nucleic acid sequences: applications in clinical research.

    PubMed

    Jenison, Robert D; Bucala, Richard; Maul, Diana; Ward, David C

    2006-01-01

    Certain optical conditions permit the unaided eye to detect thickness changes on surfaces on the order of 20 A, which are of similar dimensions to monomolecular interactions between proteins or hybridization of complementary nucleic acid sequences. Such detection exploits specific interference of reflected white light, wherein thickness changes are perceived as surface color changes. This technology, termed thin-film detection, allows for the visualization of subattomole amounts of nucleic acid targets, even in complex clinical samples. Thin-film technology has been applied to a broad range of clinically relevant indications, including the detection of pathogenic bacterial and viral nucleic acid sequences and the discrimination of sequence variations in human genes causally related to susceptibility or severity of disease.

  4. 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.

  5. RNA internal standard synthesis by nucleic acid sequence-based amplification for competitive quantitative amplification reactions.

    PubMed

    Lo, Wan-Yu; Baeumner, Antje J

    2007-02-15

    Nucleic acid sequence-based amplification (NASBA) reactions have been demonstrated to successfully synthesize new sequences based on deletion and insertion reactions. Two RNA internal standards were synthesized for use in competitive amplification reactions in which quantitative analysis can be achieved by coamplifying the internal standard with the wild type sample. The sequences were created in two consecutive NASBA reactions using the E. coli clpB mRNA sequence as model analyte. The primer sequences of the wild type sequence were maintained, and a 20-nt-long segment inside the amplicon region was exchanged for a new segment of similar GC content and melting temperature. The new RNA sequence was thus amplifiable using the wild type primers and detectable via a new inserted sequence. In the first reaction, the forwarding primer and an additional 20-nt-long sequence was deleted and replaced by a new 20-nt-long sequence. In the second reaction, a forwarding primer containing as 5' overhang sequence the wild type primer sequence was used. The presence of pure internal standard was verified using electrochemiluminescence and RNA lateral-flow biosensor analysis. Additional sequence deletion in order to shorten the internal standard amplicons and thus generate higher detection signals was found not to be required. Finally, a competitive NASBA reaction between one internal standard and the wild type sequence was carried out proving its functionality. This new rapid construction method via NASBA provides advantages over the traditional techniques since it requires no traditional cloning procedures, no thermocyclers, and can be completed in less than 4 h.

  6. Increasing phylogenetic resolution at low taxonomic levels using massively parallel sequencing of chloroplast genomes

    PubMed Central

    2009-01-01

    Background Molecular evolutionary studies share the common goal of elucidating historical relationships, and the common challenge of adequately sampling taxa and characters. Particularly at low taxonomic levels, recent divergence, rapid radiations, and conservative genome evolution yield limited sequence variation, and dense taxon sampling is often desirable. Recent advances in massively parallel sequencing make it possible to rapidly obtain large amounts of sequence data, and multiplexing makes extensive sampling of megabase sequences feasible. Is it possible to efficiently apply massively parallel sequencing to increase phylogenetic resolution at low taxonomic levels? Results We reconstruct the infrageneric phylogeny of Pinus from 37 nearly-complete chloroplast genomes (average 109 kilobases each of an approximately 120 kilobase genome) generated using multiplexed massively parallel sequencing. 30/33 ingroup nodes resolved with ≥ 95% bootstrap support; this is a substantial improvement relative to prior studies, and shows massively parallel sequencing-based strategies can produce sufficient high quality sequence to reach support levels originally proposed for the phylogenetic bootstrap. Resampling simulations show that at least the entire plastome is necessary to fully resolve Pinus, particularly in rapidly radiating clades. Meta-analysis of 99 published infrageneric phylogenies shows that whole plastome analysis should provide similar gains across a range of plant genera. A disproportionate amount of phylogenetic information resides in two loci (ycf1, ycf2), highlighting their unusual evolutionary properties. Conclusion Plastome sequencing is now an efficient option for increasing phylogenetic resolution at lower taxonomic levels in plant phylogenetic and population genetic analyses. With continuing improvements in sequencing capacity, the strategies herein should revolutionize efforts requiring dense taxon and character sampling, such as phylogeographic

  7. Puget Sound acidity levels drop after ASARCO shutdown

    SciTech Connect

    Not Available

    1987-07-01

    The levels of acidity in Puget Sound region rainfall have decreased significantly since the shutdown of the ASARCO copper smelter in Tacoma, Washington, according to a study funded by the US Environmental Protection Agency. Results indicate that sulfate and hydrogen ion concentrations obtained from samples taken before the closure were significantly different than those collected after the shutdown. Rainwater samples collected downwind during smelter operation were also significantly different from those collected upwind. Sulfur dioxide is considered to be one of the principal contributors to acid rain. The smelter was a major source of sulfur dioxide emissions in the Puget Sound region before it shut down in March 1985.

  8. Cloning, sequence, and developmental expression of a type 5, tartrate-resistant, acid phosphatase of rat bone.

    PubMed

    Ek-Rylander, B; Bill, P; Norgård, M; Nilsson, S; Andersson, G

    1991-12-25

    Tartrate-resistant acid phosphatase (TRAP) is a characteristic constituent of osteoclasts and some mononuclear preosteoclasts and, therefore, used as a histochemical and biochemical marker for osteoclasts and bone resorption. We now report the isolation of a 1397-base pair (bp) full-length TRAP/tartrate-resistant acid ATPase (TrATPase) cDNA clone from a neonatal rat calvaria lambda gt11 cDNA library. The cDNA clone consists of a 92-bp untranslated 5'-flank, an open reading frame of 981 bp and a 324-bp untranslated 3'-poly(A)-containing region. The deduced protein sequence of 327 amino acids contains a putative cleavable signal sequence of 21 amino acids. The mature polypeptide of 306 amino acids has a calculated Mr of 34,350 Da and a pI of 9.18, and it contains two potential N-glycosylation sites and the lysosomal targeting sequence DKRFQ. At the protein level, the sequence displays 89-94% homology to TRAP enzymes from human placenta, beef spleen, and uteroferrin and identity to the N terminus of purified rat bone TRAP/TrATPase. An N-terminal amino acid segment is strikingly homologous to the corresponding region in lysosomal and prostatic acid phosphatases. The cDNA recognized a 1.5-kilobase mRNA in long bones and calvaria, and in vitro translation using, as template, mRNA transcribed from the full-length insert yielded an immunoprecipitated product of 34 kDa. In neonatal rats, TRAP/TrATPase mRNA was highly expressed in skeletal tissues, with much lower (less than 10%) levels detected in spleen, thymus, liver, skin, brain, kidney, brain, lung, and heart. In situ hybridization demonstrated specific labeling of osteoclasts at endostal surfaces and bone trabeculae of long bones. Thus, despite the apparent similarity of this osteoclastic TRAP/TrATPase with type 5, tartrate-resistant and purple, acid phosphatases expressed in other mammalian tissues, this gene appears to be preferentially expressed at skeletal sites.

  9. High Level Waste System Impacts from Acid Dissolution of Sludge

    SciTech Connect

    KETUSKY, EDWARD

    2006-04-20

    This research evaluates the ability of OLI{copyright} equilibrium based software to forecast Savannah River Site High Level Waste system impacts from oxalic acid dissolution of Tank 1-15 sludge heels. Without further laboratory and field testing, only the use of oxalic acid can be considered plausible to support sludge heel dissolution on multiple tanks. Using OLI{copyright} and available test results, a dissolution model is constructed and validated. Material and energy balances, coupled with the model, identify potential safety concerns. Overpressurization and overheating are shown to be unlikely. Corrosion induced hydrogen could, however, overwhelm the tank ventilation. While pH adjustment can restore the minimal hydrogen generation, resultant precipitates will notably increase the sludge volume. OLI{copyright} is used to develop a flowsheet such that additional sludge vitrification canisters and other negative system impacts are minimized. Sensitivity analyses are used to assess the processability impacts from variations in the sludge/quantities of acids.

  10. Altered Serum Uric Acid Level in Lichen Planus Patients

    PubMed Central

    Chakraborti, Goutam; Biswas, Rabindranath; Chakraborti, Sandip; Sen, Pradyot Kumar

    2014-01-01

    Background: Lichen planus (LP) is a common disorder whose etiopathogenesis is not clear. Recently, it has been suggested that increased reactive oxygen species (ROS) play important roles in the underlying mechanism of LP. Objectives: The principal aim of this study was to evaluate serum uric acid (UA) levels as a measure of the antioxidant defense status in LP patients. Methods: Serum UA levels were determined in 58 LP patients and 61 controls. Results: Serum UA levels were significantly decreased in patients with respect to controls. Moreover, serum UA level was decreased according to increasing duration of disease. Conclusions: The results of our study suggest that LP is associated with decrease of UA levels in serum. UA may be a potential, useful biomarker of antioxidant status in LP for elaboration of treatment strategy and monitoring. PMID:25484383

  11. Amino acid sequences of two nonspecific lipid-transfer proteins from germinated castor bean.

    PubMed

    Takishima, K; Watanabe, S; Yamada, M; Suga, T; Mamiya, G

    1988-11-01

    The amino acid sequence of two nonspecific lipid-transfer proteins (nsLTP) B and C from germinated castor bean seeds have been determined. Both the proteins consist of 92 residues, as for nsLTP previously reported, and their calculated Mr values are 9847 and 9593 for nsLTP-B and nsLTP-C, respectively. The sequences of nsLTP-B and nsLTP-C, compared to the known sequence of nsLTP-A from the same source, are 68% and 35% similar, respectively. No variation was found at the positions of the cysteine residues, indicating that they might be involved in disulfide bridges.

  12. 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

  13. 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.

  14. A Mutant of Arabidopsis with Increased Levels of Stearic Acid.

    PubMed Central

    Lightner, J.; Wu, J.; Browse, J.

    1994-01-01

    A mutation at the fab2 locus of Arabidopsis caused increased levels of stearate in leaves. The increase in leaf stearate in fab2 varied developmentally, and the largest increase occurred in young leaves, where stearate accounted for almost 20% of total leaf fatty acids. The fatty acid composition of leaf lipids isolated from the fab2 mutant showed increased stearate in all the major glycerolipids of both the chloroplast and extrachloroplast membranes. Although the stearate content was increased, the fab2 mutant still contained abundant amounts of 18:1, 18:2, and 18:3 fatty acids. These results are consistent with the expectations for a mutation partially affecting the action of the stromal stearoyl-acyl carrier protein desaturase. Positional analysis indicated that the extra 18:0 is excluded with high specificity from the sn-2 position of both chloroplast and extrachloroplast glycerolipids. Although stearate content was increased in all the major leaf membrane lipids, the amount of increase varied considerably among the different lipids, from a high of 25% of fatty acids in phosphatidylcholine to a low of 2.9% of fatty acids in monogalactosyldiacylglycerol. PMID:12232421

  15. Complete amino acid sequence of the N-terminal extension of calf skin type III procollagen.

    PubMed Central

    Brandt, A; Glanville, R W; Hörlein, D; Bruckner, P; Timpl, R; Fietzek, P P; Kühn, K

    1984-01-01

    The N-terminal extension peptide of type III procollagen, isolated from foetal-calf skin, contains 130 amino acid residues. To determine its amino acid sequence, the peptide was reduced and carboxymethylated or aminoethylated and fragmented with trypsin, Staphylococcus aureus V8 proteinase and bacterial collagenase. Pyroglutamate aminopeptidase was used to deblock the N-terminal collagenase fragment to enable amino acid sequencing. The type III collagen extension peptide is homologous to that of the alpha 1 chain of type I procollagen with respect to a three-domain structure. The N-terminal 79 amino acids, which contain ten of the 12 cysteine residues, form a compact globular domain. The next 39 amino acids are in a collagenase triplet sequence (Gly- Xaa - Yaa )n with a high hydroxyproline content. Finally, another short non-collagenous domain of 12 amino acids ends at the cleavage site for procollagen aminopeptidase, which cleaves a proline-glutamine bond. In contrast with type I procollagen, the type III procollagen extension peptides contain interchain disulphide bridges located at the C-terminus of the triple-helical domain. PMID:6331392

  16. 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... And/or Amino Acid Sequences § 1.824 Form and format for nucleotide and/or amino acid sequence... readable form may be created by any means, such as word processors, nucleotide/amino acid sequence...

  17. 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... And/or Amino Acid Sequences § 1.824 Form and format for nucleotide and/or amino acid sequence... readable form may be created by any means, such as word processors, nucleotide/amino acid sequence...

  18. 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... And/or Amino Acid Sequences § 1.824 Form and format for nucleotide and/or amino acid sequence... readable form may be created by any means, such as word processors, nucleotide/amino acid sequence...

  19. Complete amino acid sequence of branched-chain amino acid aminotransferase (transaminase B) of Salmonella typhimurium, identification of the coenzyme-binding site and sequence comparison analysis

    SciTech Connect

    Feild, M.J.

    1988-01-01

    The complete amino acid sequence of the subunit of branched-chain amino acid aminotransferase of Salmonella typhimurium was determined by automated Edman degradation of peptide fragments generated by chemical and enzymatic digestion of S-carboxymethylated and S-pyridylethylated transaminase B. Peptide fragments of transaminase B were generated by treatment of the enzyme with trypsin, Staphylococcus aureus V8 protease, endoproteinase Lys-C, and cyanogen bromide. Protocols were developed for separation of the peptide fragments by reverse-phase high performance liquid chromatography (HPLC), ion-exchange HPLC, and SDS-urea gel electrophoresis. The enzyme subunit contains 308 amino acid residues and has a molecular weight of 33,920 daltons. The coenzyme-binding site was determined by treatment of the enzyme, containing bound pyridoxal 5-phosphate, with tritiated sodium borohydride prior to trypsin digestion. Monitoring radioactivity incorporation and peptide map comparisons with an apoenzyme tryptic digest, allowed identification of the pyridoxylated-peptide which was isolated by reverse-phase HPLC and sequenced. The coenzyme-binding site is a lysyl residue at position 159. Some peptides were further characterized by fast atom bombardment mass spectrometry.

  20. Sequence of the canine herpesvirus thymidine kinase gene: taxon-preferred amino acid residues in the alphaherpesviral thymidine kinases.

    PubMed

    Rémond, M; Sheldrick, P; Lebreton, F; Foulon, T

    1995-12-01

    Multiple sequence alignments of evolutionarily related proteins are finding increasing use as indicators of critical amino acid residues necessary for structural stability or involved in functional domains responsible for catalytic activities. In the past, a number of alignments have provided such information for the herpesviral thymidine kinases, for which three-dimensional structures are not yet available. We have sequenced the thymidine kinase gene of a canine herpesvirus, and with a multiple alignment have identified amino acids preferentially conserved in either of two taxons, the genera Varicellovirus and Simplexvirus, of the subfamily Alphaherpesvirinae. Since some regions of the thymidine kinases show otherwise elevated levels of substitutional tolerance, these conserved amino acids are candidates for critical residues which have become fixed through selection during the evolutionary divergence of these enzymes. Several pairs with distinctive patterns of distribution among the various viruses occur in or near highly conserved sequence motifs previously proposed to form the catalytic site, and we speculate that they may represent interacting, co-ordinately variable residues.

  1. The amino acid sequence of cytochromes c-551 from three species of Pseudomonas

    PubMed Central

    Ambler, R. P.; Wynn, Margaret

    1973-01-01

    The amino acid sequences of the cytochromes c-551 from three species of Pseudomonas have been determined. Each resembles the protein from Pseudomonas strain P6009 (now known to be Pseudomonas aeruginosa, not Pseudomonas fluorescens) in containing 82 amino acids in a single peptide chain, with a haem group covalently attached to cysteine residues 12 and 15. In all four sequences 43 residues are identical. Although by bacteriological criteria the organisms are closely related, the differences between pairs of sequences range from 22% to 39%. These values should be compared with the differences in the sequence of mitochondrial cytochrome c between mammals and amphibians (about 18%) or between mammals and insects (about 33%). Detailed evidence for the amino acid sequences of the proteins has been deposited as Supplementary Publication SUP 50015 at 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. (1973), 131, 5. PMID:4352718

  2. Draft Genome Sequence of Sorghum Grain Mold Fungus Epicoccum sorghinum, a Producer of Tenuazonic Acid

    PubMed Central

    Oliveira, Rodrigo C.; Davenport, Karen W.; Hovde, Blake; Silva, Danielle; Chain, Patrick S. G.; Correa, Benedito

    2017-01-01

    ABSTRACT The facultative plant pathogen Epicoccum sorghinum is associated with grain mold of sorghum and produces the mycotoxin tenuazonic acid. This fungus can have serious economic impact on sorghum production. Here, we report the draft genome sequence of E. sorghinum (USPMTOX48). PMID:28126937

  3. 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.

  4. Draft Genome Sequence of Bacillus coagulans NL01, a Wonderful l-Lactic Acid Producer

    PubMed Central

    Zheng, Zhaojuan; Jiang, Ting; Lin, Xi; Zhou, Jie

    2015-01-01

    Here, we report the draft genome sequence of Bacillus coagulans NL01, which could produce high optically pure l-lactic acid using xylose as a sole carbon source. The draft genome is 3,505,081 bp, with 144 contigs. About 3,903 protein-coding genes and 92 rRNAs are predicted from this assembly. PMID:26089419

  5. Does high serum uric acid level cause aspirin resistance?

    PubMed

    Yildiz, Bekir S; Ozkan, Emel; Esin, Fatma; Alihanoglu, Yusuf I; Ozkan, Hayrettin; Bilgin, Murat; Kilic, Ismail D; Ergin, Ahmet; Kaftan, Havane A; Evrengul, Harun

    2016-06-01

    In patients with coronary artery disease (CAD), though aspirin inhibits platelet activation and reduces atherothrombotic complications, it does not always sufficiently inhibit platelet function, thereby causing a clinical situation known as aspirin resistance. As hyperuricemia activates platelet turnover, aspirin resistance may be specifically induced by increased serum uric acid (SUA) levels. In this study, we thus investigated the association between SUA level and aspirin resistance in patients with CAD. We analyzed 245 consecutive patients with stable angina pectoris (SAP) who in coronary angiography showed more than 50% occlusion in a major coronary artery. According to aspirin resistance, two groups were formed: the aspirin resistance group (Group 1) and the aspirin-sensitive group (Group 2). Compared with those of Group 2, patients with aspirin resistance exhibited significantly higher white blood cell counts, neutrophil counts, neutrophil-to-lymphocyte ratios, SUA levels, high-sensitivity C-reactive protein levels, and fasting blood glucose levels. After multivariate analysis, a high level of SUA emerged as an independent predictor of aspirin resistance. The receiver-operating characteristic analysis provided a cutoff value of 6.45 mg/dl for SUA to predict aspirin resistance with 79% sensitivity and 65% specificity. Hyperuricemia may cause aspirin resistance in patients with CAD and high SUA levels may indicate aspirin-resistant patients. Such levels should thus recommend avoiding heart attack and stroke by adjusting aspirin dosage.

  6. Amino acid sequences of heterotrophic and photosynthetic ferredoxins from the tomato plant (Lycopersicon esculentum Mill.).

    PubMed

    Kamide, K; Sakai, H; Aoki, K; Sanada, Y; Wada, K; Green, L S; Yee, B C; Buchanan, B B

    1995-11-01

    Several forms (isoproteins) of ferredoxin in roots, leaves, and green and red pericarps in tomato plants (Lycopersicon esculentum Mill.) were earlier identified on the basis of N-terminal amino acid sequence and chromatographic behavior (Green et al. 1991). In the present study, a large scale preparation made possible determination of the full length amino acid sequence of the two ferredoxins from leaves. The ferredoxins characteristic of fruit and root were sequenced from the amino terminus to the 30th residue or beyond. The leaf ferredoxins were confirmed to be expressed in pericarp of both green and red fruit. The ferredoxins characteristic of fruit and root appeared to be restricted to those tissue. The results extend earlier findings in demonstrating that ferredoxin occurs in the major organs of the tomato plant where it appears to function irrespective of photosynthetic competence.

  7. Amino acid sequence of myoglobin from white-tailed deer (Odocoileus virginianus).

    PubMed

    Joseph, Poulson; Suman, Surendranath P; Li, Shuting; Fontaine, Michele; Steinke, Laurey

    2012-10-01

    Our objective was to determine the primary structure of white-tailed deer myoglobin (Mb). White-tailed deer Mb was isolated from cardiac muscles employing ammonium sulfate precipitation and gel-filtration chromatography. The amino acid sequence was determined by Edman degradation. Sequence analyses of intact Mb as well as tryptic- and cyanogen bromide-peptides yielded the complete primary structure of white-tailed deer Mb, which shared 100% similarity with red deer Mb. White-tailed deer Mb consists of 153 amino acid residues and shares more than 96% sequence similarity with myoglobins from meat-producing ruminants, such as cattle, buffalo, sheep, and goat. Similar to sheep and goat myoglobins, white-tailed deer Mb contains 12 histidine residues. Proximal (position 93) and distal (position 64) histidine residues responsible for maintaining the stability of heme are conserved in white-tailed deer Mb.

  8. Nucleotide sequence and the encoded amino acids of human apolipoprotein A-I mRNA.

    PubMed Central

    Law, S W; Brewer, H B

    1984-01-01

    The cDNA clones encoding the precursor form of human liver apolipoprotein A-I (apoA-I), preproapoA-I, have been isolated from a cDNA library. A 17-base synthetic oligonucleotide based on residues 108-113 of apoA-I and a 26-base primer-extended, dideoxynucleotide-terminated cDNA were used as hybridization probes to select for recombinant plasmids bearing the apoA-I sequence. The complete nucleic acid sequence of human liver preproapoA-I has been determined by analysis of the cloned cDNA. The sequence is composed of 801 nucleotides encoding 267 amino acid residues. PreproapoA-I contains an 18-amino-acid prepeptide and a 6-amino-acid propeptide connected to the amino terminus of the 243-amino acid mature apoA-I. Southern blotting analysis of chromosomal DNA obtained from peripheral blood indicated the apoA-I gene is contained in a 2.1-kilobase-pair Pst I fragment and there is no gross difference in structural organization between the normal apoA-I gene and the Tangier disease apoA-I gene. Images PMID:6198645

  9. Life-history evolution at the molecular level: adaptive amino acid composition of avian vitellogenins

    PubMed Central

    Hughes, Austin L.

    2015-01-01

    Avian genomes typically encode three distinct vitellogenin (VTG) egg yolk proteins (VTG1, VTG2 and VTG3), which arose by gene duplication prior to the most recent common ancestor of birds. Analysis of VTG sequences from 34 avian species in a phylogenetic framework supported the hypothesis that VTG amino acid composition has co-evolved with embryo incubation time. Embryo incubation time was positively correlated with the proportions of dietary essential amino acids (EAAs) in VTG1 and VTG2, and with the proportion of sulfur-containing amino acids in VTG3. These patterns were seen even when only semi-altricial and/or altricial species were considered, suggesting that the duration of embryo incubation is a major selective factor on the amino acid composition of VTGs, rather than developmental mode alone. The results are consistent with the hypothesis that the level of EAAs provided to the egg represents an adaptation to the loss of amino acids through breakdown over the course of incubation and imply that life-history phenotypes and VTG amino acid composition have co-evolved throughout the evolutionary history of birds. PMID:26224713

  10. The Impact of Selection at the Amino Acid Level on the Usage of Synonymous Codons

    PubMed Central

    Błażej, Paweł; Mackiewicz, Dorota; Wnętrzak, Małgorzata; Mackiewicz, Paweł

    2017-01-01

    There are two main forces that affect usage of synonymous codons: directional mutational pressure and selection. The effectiveness of protein translation is usually considered as the main selectional factor. However, biased codon usage can also be a byproduct of a general selection at the amino acid level interacting with nucleotide replacements. To evaluate the validity and strength of such an effect, we superimposed >3.5 billion unrestricted mutational processes on the selection of nonsynonymous substitutions based on the differences in physicochemical properties of the coded amino acids. Using a modified evolutionary optimization algorithm, we determined the conditions in which the effect on the relative codon usage is maximized. We found that the effect is enhanced by mutational processes generating more adenine and thymine than guanine and cytosine, as well as more purines than pyrimidines. Interestingly, this effect is observed only under an unrestricted model of nucleotide substitution, and disappears when the mutational process is time-reversible. Comparison of the simulation results with data for real protein coding sequences indicates that the impact of selection at the amino acid level on synonymous codon usage cannot be neglected. Furthermore, it can considerably interfere, especially in AT-rich genomes, with other selections on codon usage, e.g., translational efficiency. It may also lead to difficulties in the recognition of other effects influencing codon bias, and an overestimation of protein coding sequences whose codon usage is subjected to adaptational selection. PMID:28122952

  11. Mathematical Characterization of Protein Sequences Using Patterns as Chemical Group Combinations of Amino Acids.

    PubMed

    Das, Jayanta Kumar; Das, Provas; Ray, Korak Kumar; Choudhury, Pabitra Pal; Jana, Siddhartha Sankar

    2016-01-01

    Comparison of amino acid sequence similarity is the fundamental concept behind the protein phylogenetic tree formation. By virtue of this method, we can explain the evolutionary relationships, but further explanations are not possible unless sequences are studied through the chemical nature of individual amino acids. Here we develop a new methodology to characterize the protein sequences on the basis of the chemical nature of the amino acids. We design various algorithms for studying the variation of chemical group transitions and various chemical group combinations as patterns in the protein sequences. The amino acid sequence of conventional myosin II head domain of 14 family members are taken to illustrate this new approach. We find two blocks of maximum length 6 aa as 'FPKATD' and 'Y/FTNEKL' without repeating the same chemical nature and one block of maximum length 20 aa with the repetition of chemical nature which are common among all 14 members. We also check commonality with another motor protein sub-family kinesin, KIF1A. Based on our analysis we find a common block of length 8 aa both in myosin II and KIF1A. This motif is located in the neck linker region which could be responsible for the generation of mechanical force, enabling us to find the unique blocks which remain chemically conserved across the family. We also validate our methodology with different protein families such as MYOI, Myosin light chain kinase (MLCK) and Rho-associated protein kinase (ROCK), Na+/K+-ATPase and Ca2+-ATPase. Altogether, our studies provide a new methodology for investigating the conserved amino acids' pattern in different proteins.

  12. 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.

  13. Light-Induced Alterations in Basil Ganglia Kynurenic Acid Levels

    NASA Technical Reports Server (NTRS)

    Sroufe, Angela E.; Whittaker, J. A.; Patrickson, J. W.; Orr, M. C.

    1997-01-01

    The metabolic synthesis, release and breakdown of several known CNS neurotransmitters have been shown to follow a circadian pattern entrained to the environmental light/dark cycle. The levels of excitatory amino acid (EAA) transmitters such as glutamate, have been shown to vary with environmental lighting conditions. Kynurenic Acid (KA), an endogenous tryptophan metabolite and glutamate receptor antagonist, has been reported to have neuroprotective effects against EAA-induced excitotoxic cell damage. Changes in KA's activity within the mammalian basal ganglia has been proposed as being contributory to neurotoxicity in Huntington's Disease. It is not known whether CNS KA levels follow a circadian pattern or exhibit light-induced fluctuations. However, because the symptoms of certain degenerative motor disorders seem to fluctuate with daily 24 hour rhythm, we initiated studies to determine if basal ganglia KA were influenced by the daily light/dark cycle and could influence motor function. Therefore in this study, HPLC-EC was utilized to determine if basal ganglia KA levels in tissue extracts from adult male Long-Evans rats (200-250g) entrained to 24 and 48 hours constant light and dark conditions, respectively. Samples were taken one hour before the onset of the subjective day and one hour prior to the onset of the subjective night in order to detect possible phase differences in KA levels and to allow for accumulation of factors expressed in association with the light or dark phase. Data analysis revealed that KA levels in the basal ganglia vary with environmental lighting conditions; being elevated generally during the dark. Circadian phase differences in KA levels were also evident during the subjective night and subjective day, respectively. Results from these studies are discussed with respect to potential cyclic changes in neuronal susceptibility to excitotoxic damage during the daily 24 hour cycle and its possible relevance to future therapeutic approaches in

  14. Amino acid sequence of band-3 protein from rainbow trout erythrocytes derived from cDNA.

    PubMed Central

    Hübner, S; Michel, F; Rudloff, V; Appelhans, H

    1992-01-01

    In this report we present the first complete band-3 cDNA sequence of a poikilothermic lower vertebrate. The primary structure of the anion-exchange protein band 3 (AE1) from rainbow trout erythrocytes was determined by nucleotide sequencing of cDNA clones. The overlapping clones have a total length of 3827 bp with a 5'-terminal untranslated region of 150 bp, a 2754 bp open reading frame and a 3'-untranslated region of 924 bp. Band-3 protein from trout erythrocytes consists of 918 amino acid residues with a calculated molecular mass of 101 827 Da. Comparison of its amino acid sequence revealed a 60-65% identity within the transmembrane spanning sequence of band-3 proteins published so far. An additional insertion of 24 amino acid residues within the membrane-associated domain of trout band-3 protein was identified, which until now was thought to be a general feature only of mammalian band-3-related proteins. PMID:1637296

  15. 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,

    2016-07-12

    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.

  16. Role of the two-component leader sequence and mature amino acid sequences in extracellular export of endoglucanase EGL from Pseudomonas solanacearum.

    PubMed Central

    Huang, J Z; Schell, M A

    1992-01-01

    The egl gene of Pseudomonas solanacearum encodes a 43-kDa extracellular endoglucanase (mEGL) involved in wilt disease caused by this phytopathogen. Egl is initially translated with a 45-residue, two-part leader sequence. The first 19 residues are apparently removed by signal peptidase II during export of Egl across the inner membrane (IM); the remaining residues of the leader sequence (modified with palmitate) are removed during export across the outer membrane (OM). Localization of Egl-PhoA fusion proteins showed that the first 26 residues of the Egl leader sequence are required and sufficient to direct lipid modification, processing, and export of Egl or PhoA across the IM but not the OM. Fusions of the complete 45-residue leader sequence or of the leader and increasing portions of mEgl sequences to PhoA did not cause its export across the OM. In-frame deletion of portions of mEGL-coding sequences blocked export of the truncated polypeptides across the OM without affecting export across the IM. These results indicate that the first part of the leader sequence functions independently to direct export of Egl across the IM while the second part and sequences and structures in mEGL are involved in export across the OM. Computer analysis of the mEgl amino acid sequence obtained from its nucleotide sequence identified a region of mEGL similar in amino acid sequence to regions in other prokaryotic endoglucanases. Images PMID:1735723

  17. Studies on adenosine triphosphate transphosphorylases. Amino acid sequence of rabbit muscle ATP-AMP transphosphorylase.

    PubMed

    Kuby, S A; Palmieri, R H; Frischat, A; Fischer, A H; Wu, L H; Maland, L; Manship, M

    1984-05-22

    The total amino acid sequence of rabbit muscle adenylate kinase has been determined, and the single polypeptide chain of 194 amino acid residues starts with N-acetylmethionine and ends with leucyllysine at its carboxyl terminus, in agreement with the earlier data on its amino acid composition [Mahowald, T. A., Noltmann, E. A., & Kuby, S. A. (1962) J. Biol. Chem. 237, 1138-1145] and its carboxyl-terminus sequence [Olson, O. E., & Kuby, S. A. (1964) J. Biol. Chem. 239, 460-467]. Elucidation of the primary structure was based on tryptic and chymotryptic cleavages of the performic acid oxidized protein, cyanogen bromide cleavages of the 14C-labeled S-carboxymethylated protein at its five methionine sites (followed by maleylation of peptide fragments), and tryptic cleavages at its 12 arginine sites of the maleylated 14C-labeled S-carboxymethylated protein. Calf muscle myokinase, whose sequence has also been established, differs primarily from the rabbit muscle myokinase's sequence in the following: His-30 is replaced by Gln-30; Lys-56 is replaced by Met-56; Ala-84 and Asp 85 are replaced by Val-84 and Asn-85. A comparison of the four muscle-type adenylate kinases, whose covalent structures have now been determined, viz., rabbit, calf, porcine, and human [for the latter two sequences see Heil, A., Müller, G., Noda, L., Pinder, T., Schirmer, H., Schirmer, I., & Von Zabern, I. (1974) Eur. J. Biochem. 43, 131-144, and Von Zabern, I., Wittmann-Liebold, B., Untucht-Grau, R., Schirmer, R. H., & Pai, E. F. (1976) Eur. J. Biochem. 68, 281-290], demonstrates an extraordinary degree of homology.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Mathematical Characterization of Protein Sequences Using Patterns as Chemical Group Combinations of Amino Acids

    PubMed Central

    Choudhury, Pabitra Pal; Jana, Siddhartha Sankar

    2016-01-01

    Comparison of amino acid sequence similarity is the fundamental concept behind the protein phylogenetic tree formation. By virtue of this method, we can explain the evolutionary relationships, but further explanations are not possible unless sequences are studied through the chemical nature of individual amino acids. Here we develop a new methodology to characterize the protein sequences on the basis of the chemical nature of the amino acids. We design various algorithms for studying the variation of chemical group transitions and various chemical group combinations as patterns in the protein sequences. The amino acid sequence of conventional myosin II head domain of 14 family members are taken to illustrate this new approach. We find two blocks of maximum length 6 aa as ‘FPKATD’ and ‘Y/FTNEKL’ without repeating the same chemical nature and one block of maximum length 20 aa with the repetition of chemical nature which are common among all 14 members. We also check commonality with another motor protein sub-family kinesin, KIF1A. Based on our analysis we find a common block of length 8 aa both in myosin II and KIF1A. This motif is located in the neck linker region which could be responsible for the generation of mechanical force, enabling us to find the unique blocks which remain chemically conserved across the family. We also validate our methodology with different protein families such as MYOI, Myosin light chain kinase (MLCK) and Rho-associated protein kinase (ROCK), Na+/K+-ATPase and Ca2+-ATPase. Altogether, our studies provide a new methodology for investigating the conserved amino acids’ pattern in different proteins. PMID:27930687

  19. The complete amino acid sequence of a trypsin inhibitor from Bauhinia variegata var. candida seeds.

    PubMed

    Di Ciero, L; Oliva, M L; Torquato, R; Köhler, P; Weder, J K; Camillo Novello, J; Sampaio, C A; Oliveira, B; Marangoni, S

    1998-11-01

    Trypsin inhibitors of two varieties of Bauhinia variegata seeds have been isolated and characterized. Bauhinia variegata candida trypsin inhibitor (BvcTI) and B. variegata lilac trypsin inhibitor (BvlTI) are proteins with Mr of about 20,000 without free sulfhydryl groups. Amino acid analysis shows a high content of aspartic acid, glutamic acid, serine, and glycine, and a low content of histidine, tyrosine, methionine, and lysine in both inhibitors. Isoelectric focusing for both varieties detected three isoforms (pI 4.85, 5.00, and 5.15), which were resolved by HPLC procedure. The trypsin inhibitors show Ki values of 6.9 and 1.2 nM for BvcTI and BvlTI, respectively. The N-terminal sequences of the three trypsin inhibitor isoforms from both varieties of Bauhinia variegata and the complete amino acid sequence of B. variegata var. candida L. trypsin inhibitor isoform 3 (BvcTI-3) are presented. The sequences have been determined by automated Edman degradation of the reduced and carboxymethylated proteins of the peptides resulting from Staphylococcus aureus protease and trypsin digestion. BvcTI-3 is composed of 167 residues and has a calculated molecular mass of 18,529. Homology studies with other trypsin inhibitors show that BvcTI-3 belongs to the Kunitz family. The putative active site encompasses Arg (63)-Ile (64).

  20. 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.

  1. SUBGROUPS OF AMINO ACID SEQUENCES IN THE VARIABLE REGIONS OF IMMUNOGLOBULIN HEAVY CHAINS*

    PubMed Central

    Cunningham, Bruce A.; Pflumm, Mollie N.; User, Urs Rutisha; Edelman, Gerald M.

    1969-01-01

    The amino acid sequence of the first 133 residues of the heavy (γ) chain from a human γG immunoglobulin (He) has been determined. This γ-chain is identical in Gm type to that of protein Eu, the complete sequence of which has been reported. Comparison of the two sequences substantiates the previous suggestion that there are subgroups of variable regions of heavy chains. The variable region of Eu has been assigned to subgroup I and that of He to subgroup II; on the other hand, the constant regions of the two proteins appear to be identical. Comparison of the sequence of the heavy chain of He with the heavy chain sequences determined in other laboratories suggests that the variable region of subgroup II is at least 118 residues long. The nature and distribution of amino acid variations in this heavy chain subgroup resemble those observed in light chain subgroups. These studies provide evidence that the translocation hypothesis applies to heavy as well as to light chains, viz., genes for variable regions (V) are somatically translocated to genes for constant regions (C) to form complete VC structural genes. Images PMID:5264153

  2. 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.

  3. Population sequencing of two endocannabinoid metabolic genes identifies rare and common regulatory variants associated with extreme obesity and metabolite level

    PubMed Central

    2010-01-01

    Background Targeted re-sequencing of candidate genes in individuals at the extremes of a quantitative phenotype distribution is a method of choice to gain information on the contribution of rare variants to disease susceptibility. The endocannabinoid system mediates signaling in the brain and peripheral tissues involved in the regulation of energy balance, is highly active in obese patients, and represents a strong candidate pathway to examine for genetic association with body mass index (BMI). Results We sequenced two intervals (covering 188 kb) encoding the endocannabinoid metabolic enzymes fatty-acid amide hydrolase (FAAH) and monoglyceride lipase (MGLL) in 147 normal controls and 142 extremely obese cases. After applying quality filters, we called 1,393 high quality single nucleotide variants, 55% of which are rare, and 143 indels. Using single marker tests and collapsed marker tests, we identified four intervals associated with BMI: the FAAH promoter, the MGLL promoter, MGLL intron 2, and MGLL intron 3. Two of these intervals are composed of rare variants and the majority of the associated variants are located in promoter sequences or in predicted transcriptional enhancers, suggesting a regulatory role. The set of rare variants in the FAAH promoter associated with BMI is also associated with increased level of FAAH substrate anandamide, further implicating a functional role in obesity. Conclusions Our study, which is one of the first reports of a sequence-based association study using next-generation sequencing of candidate genes, provides insights into study design and analysis approaches and demonstrates the importance of examining regulatory elements rather than exclusively focusing on exon sequences. PMID:21118518

  4. 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.

  5. Amino-Acid Sequence of NADP-Specific Glutamate Dehydrogenase of Neurospora crassa

    PubMed Central

    Wootton, John C.; Chambers, Geoffrey K.; Holder, Anthony A.; Baron, Andrew J.; Taylor, John G.; Fincham, John R. S.; Blumenthal, Kenneth M.; Moon, Kenneth; Smith, Emil L.

    1974-01-01

    A tentative primary structure of the NADP-specific glutamate dehydrogenase [L-glutamate: NADP oxidoreductase (deaminating), EC 1.4.1.4] from Neurospora crassa has been determined. The proposed sequence contains 452 amino-acid residues in each of the identical subunits of the hexameric enzyme. Comparison of the sequence with that of the bovine liver enzyme reveals considerable homology in the amino-terminal portion of the chain, including the vicinity of the reactive lysine, with only shorter stretches of homology within the carboxyl-terminal regions. The significance of this distribution of homologous regions is discussed. PMID:4155068

  6. Seq2Logo: a method for construction and visualization of amino acid binding motifs and sequence profiles including sequence weighting, pseudo counts and two-sided representation of amino acid enrichment and depletion

    PubMed Central

    Thomsen, Martin Christen Frølund; Nielsen, Morten

    2012-01-01

    Seq2Logo is a web-based sequence logo generator. Sequence logos are a graphical representation of the information content stored in a multiple sequence alignment (MSA) and provide a compact and highly intuitive representation of the position-specific amino acid composition of binding motifs, active sites, etc. in biological sequences. Accurate generation of sequence logos is often compromised by sequence redundancy and low number of observations. Moreover, most methods available for sequence logo generation focus on displaying the position-specific enrichment of amino acids, discarding the equally valuable information related to amino acid depletion. Seq2logo aims at resolving these issues allowing the user to include sequence weighting to correct for data redundancy, pseudo counts to correct for low number of observations and different logotype representations each capturing different aspects related to amino acid enrichment and depletion. Besides allowing input in the format of peptides and MSA, Seq2Logo accepts input as Blast sequence profiles, providing easy access for non-expert end-users to characterize and identify functionally conserved/variable amino acids in any given protein of interest. The output from the server is a sequence logo and a PSSM. Seq2Logo is available at http://www.cbs.dtu.dk/biotools/Seq2Logo (14 May 2012, date last accessed). PMID:22638583

  7. Seq2Logo: a method for construction and visualization of amino acid binding motifs and sequence profiles including sequence weighting, pseudo counts and two-sided representation of amino acid enrichment and depletion.

    PubMed

    Thomsen, Martin Christen Frølund; Nielsen, Morten

    2012-07-01

    Seq2Logo is a web-based sequence logo generator. Sequence logos are a graphical representation of the information content stored in a multiple sequence alignment (MSA) and provide a compact and highly intuitive representation of the position-specific amino acid composition of binding motifs, active sites, etc. in biological sequences. Accurate generation of sequence logos is often compromised by sequence redundancy and low number of observations. Moreover, most methods available for sequence logo generation focus on displaying the position-specific enrichment of amino acids, discarding the equally valuable information related to amino acid depletion. Seq2logo aims at resolving these issues allowing the user to include sequence weighting to correct for data redundancy, pseudo counts to correct for low number of observations and different logotype representations each capturing different aspects related to amino acid enrichment and depletion. Besides allowing input in the format of peptides and MSA, Seq2Logo accepts input as Blast sequence profiles, providing easy access for non-expert end-users to characterize and identify functionally conserved/variable amino acids in any given protein of interest. The output from the server is a sequence logo and a PSSM. Seq2Logo is available at http://www.cbs.dtu.dk/biotools/Seq2Logo (14 May 2012, date last accessed).

  8. Ocean acidification increases fatty acids levels of larval fish.

    PubMed

    Díaz-Gil, Carlos; Catalán, Ignacio A; Palmer, Miquel; Faulk, Cynthia K; Fuiman, Lee A

    2015-07-01

    Rising levels of anthropogenic carbon dioxide in the atmosphere are acidifying the oceans and producing diverse and important effects on marine ecosystems, including the production of fatty acids (FAs) by primary producers and their transfer through food webs. FAs, particularly essential FAs, are necessary for normal structure and function in animals and influence composition and trophic structure of marine food webs. To test the effect of ocean acidification (OA) on the FA composition of fish, we conducted a replicated experiment in which larvae of the marine fish red drum (Sciaenops ocellatus) were reared under a climate change scenario of elevated CO2 levels (2100 µatm) and under current control levels (400 µatm). We found significantly higher whole-body levels of FAs, including nine of the 11 essential FAs, and altered relative proportions of FAs in the larvae reared under higher levels of CO2. Consequences of this effect of OA could include alterations in performance and survival of fish larvae and transfer of FAs through food webs.

  9. Ocean acidification increases fatty acids levels of larval fish

    PubMed Central

    Díaz-Gil, Carlos; Catalán, Ignacio A.; Palmer, Miquel; Faulk, Cynthia K.; Fuiman, Lee A.

    2015-01-01

    Rising levels of anthropogenic carbon dioxide in the atmosphere are acidifying the oceans and producing diverse and important effects on marine ecosystems, including the production of fatty acids (FAs) by primary producers and their transfer through food webs. FAs, particularly essential FAs, are necessary for normal structure and function in animals and influence composition and trophic structure of marine food webs. To test the effect of ocean acidification (OA) on the FA composition of fish, we conducted a replicated experiment in which larvae of the marine fish red drum (Sciaenops ocellatus) were reared under a climate change scenario of elevated CO2 levels (2100 µatm) and under current control levels (400 µatm). We found significantly higher whole-body levels of FAs, including nine of the 11 essential FAs, and altered relative proportions of FAs in the larvae reared under higher levels of CO2. Consequences of this effect of OA could include alterations in performance and survival of fish larvae and transfer of FAs through food webs. PMID:26179801

  10. Association between serum folic acid level and erectile dysfunction.

    PubMed

    Karabakan, M; Erkmen, A E; Guzel, O; Aktas, B K; Bozkurt, A; Akdemir, S

    2016-06-01

    This study measured the serum folic acid (FA) level in patients with erectile dysfunction (ED) and evaluated the possible association between the serum FA level and erectile function. The study divided 120 patients with ED into 3 groups of 40 patients each: those with severe, moderate and mild ED. Forty healthy men served as controls. Fasting serum samples were obtained, and the total testosterone, cholesterol and FA levels were measured using chemiluminescent immunoassays. There were no significant differences in the mean age, mean body mass index or mean serum total testosterone and cholesterol levels among the three ED groups and controls (P > 0.05). The mean serum FA concentrations were 7.2 ± 3.7, 7.1 ± 3.2, 10.2 ± 4.6 and 10.7 ± 4.6 ng ml(-1) in the severe, moderate and mild ED and control groups respectively. The mean serum FA concentration was significantly higher in the control group than in the severe and moderate ED groups (both P < 0.001), but not the mild ED group (P = 0.95). Considering the significant differences in the serum FA levels between the control and ED groups, serum FA deficiency might reflect the severity of ED.

  11. Cytokine levels affected by gamma-linolenic acid.

    PubMed

    Dirks, J; van Aswegen, C H; du Plessis, D J

    1998-10-01

    This study was undertaken to assess whether gamma-linolenic acid (GLA) in the form of evening primrose oil (EPO) could affect rat serum cytokines, interferon-gamma (IFN-gamma), monocyte chemotactic protein-1 (MCP-1) and tumour necrosis factor-alpha (TNF-alpha). The following diets were administered: control, glucan, Freund's adjuvant and glucan plus Freund's adjuvant with and without GLA. In the presence of GLA, the IFN-gamma and MCP-1 levels were significantly decreased in contrast to the control group of TNF-alpha, which was significantly stimulated. On account of interaction between diets and GLA, the remaining diet groups of TNF-alpha were either not affected or were inhibited in the presence of GLA. The observations indicate that GLA may modulate the level of serum IFN-gamma, MCP-1 and TNF-alpha, which may be a worthwhile line of treatment in certain human diseases.

  12. Level sequence and splitting identification of closely spaced energy levels by angle-resolved analysis of fluorescence light

    NASA Astrophysics Data System (ADS)

    Wu, Z. W.; Volotka, A. V.; Surzhykov, A.; Dong, C. Z.; Fritzsche, S.

    2016-06-01

    The angular distribution and linear polarization of the fluorescence light following the resonant photoexcitation is investigated within the framework of density matrix and second-order perturbation theory. Emphasis has been placed on "signatures" for determining the level sequence and splitting of intermediate (partially) overlapping resonances, if analyzed as a function of photon energy of incident light. Detailed computations within the multiconfiguration Dirac-Fock method have been performed, especially for the 1 s22 s22 p63 s ,Ji=1 /2 +γ1→(1s22 s 2 p63 s ) 13 p3 /2,J =1 /2 ,3 /2 →1 s22 s22 p63 s ,Jf=1 /2 +γ2 photoexcitation and subsequent fluorescence emission of atomic sodium. A remarkably strong dependence of the angular distribution and linear polarization of the γ2 fluorescence emission is found upon the level sequence and splitting of the intermediate (1s22 s 2 p63 s ) 13 p3 /2,J =1 /2 ,3 /2 overlapping resonances owing to their finite lifetime (linewidth). We therefore suggest that accurate measurements of the angular distribution and linear polarization might help identify the sequence and small splittings of closely spaced energy levels, even if they cannot be spectroscopically resolved.

  13. A rotamer energy level study of sulfuric acid.

    PubMed

    Partanen, Lauri; Pesonen, Janne; Sjöholm, Elina; Halonen, Lauri

    2013-10-14

    It is a common approach in quantum chemical calculations for polyatomic molecules to rigidly constrain some of the degrees of freedom in order to make the calculations computationally feasible. However, the presence of the rigid constraints also affects the kinetic energy operator resulting in the frozen mode correction, originally derived by Pesonen [J. Chem. Phys. 139, 144310 (2013)]. In this study, we compare the effects of this correction to several different approximations to the kinetic energy operator used in the literature, in the specific case of the rotamer energy levels of sulfuric acid. The two stable conformers of sulfuric acid are connected by the rotations of the O-S-O-H dihedral angles and possess C2 and Cs symmetry in the order of increasing energy. Our results show that of the models tested, the largest differences with the frozen mode corrected values were obtained by simply omitting the passive degrees of freedom. For the lowest 17 excited states, this inappropriate treatment introduces an increase of 9.6 cm(-1) on average, with an increase of 8.7 cm(-1) in the zero-point energies. With our two-dimensional potential energy surface calculated at the CCSD(T)-F12a/VDZ-F12 level, we observe a radical shift in the density of states compared to the harmonic picture, combined with an increase in zero point energy. Thus, we conclude that the quantum mechanical inclusion of the different conformers of sulfuric acid have a significant effect on its vibrational partition function, suggesting that it will also have an impact on the computational values of the thermodynamic properties of any reactions where sulfuric acid plays a role. Finally, we also considered the effect of the anharmonicities for the other vibrational degrees of freedom with a VSCF-calculation at the DF-MP2-F12/VTZ-F12 level of theory but found that the inclusion of the other conformer had the more important effect on the vibrational partition function.

  14. 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.

  15. 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.

  16. Genetic Architecture of Vitamin B12 and Folate Levels Uncovered Applying Deeply Sequenced Large Datasets

    PubMed Central

    Thorleifsson, Gudmar; Ahluwalia, Tarunveer S.; Steinthorsdottir, Valgerdur; Bjarnason, Helgi; Gudbjartsson, Daniel F.; Magnusson, Olafur T.; Sparsø, Thomas; Albrechtsen, Anders; Kong, Augustine; Masson, Gisli; Tian, Geng; Cao, Hongzhi; Nie, Chao; Kristiansen, Karsten; Husemoen, Lise Lotte; Thuesen, Betina; Li, Yingrui; Nielsen, Rasmus; Linneberg, Allan; Olafsson, Isleifur; Eyjolfsson, Gudmundur I.; Jørgensen, Torben; Wang, Jun; Hansen, Torben; Thorsteinsdottir, Unnur; Stefánsson, Kari; Pedersen, Oluf

    2013-01-01

    Genome-wide association studies have mainly relied on common HapMap sequence variations. Recently, sequencing approaches have allowed analysis of low frequency and rare variants in conjunction with common variants, thereby improving the search for functional variants and thus the understanding of the underlying biology of human traits and diseases. Here, we used a large Icelandic whole genome sequence dataset combined with Danish exome sequence data to gain insight into the genetic architecture of serum levels of vitamin B12 (B12) and folate. Up to 22.9 million sequence variants were analyzed in combined samples of 45,576 and 37,341 individuals with serum B12 and folate measurements, respectively. We found six novel loci associating with serum B12 (CD320, TCN2, ABCD4, MMAA, MMACHC) or folate levels (FOLR3) and confirmed seven loci for these traits (TCN1, FUT6, FUT2, CUBN, CLYBL, MUT, MTHFR). Conditional analyses established that four loci contain additional independent signals. Interestingly, 13 of the 18 identified variants were coding and 11 of the 13 target genes have known functions related to B12 and folate pathways. Contrary to epidemiological studies we did not find consistent association of the variants with cardiovascular diseases, cancers or Alzheimer's disease although some variants demonstrated pleiotropic effects. Although to some degree impeded by low statistical power for some of these conditions, these data suggest that sequence variants that contribute to the population diversity in serum B12 or folate levels do not modify the risk of developing these conditions. Yet, the study demonstrates the value of combining whole genome and exome sequencing approaches to ascertain the genetic and molecular architectures underlying quantitative trait associations. PMID:23754956

  17. Genetic architecture of vitamin B12 and folate levels uncovered applying deeply sequenced large datasets.

    PubMed

    Grarup, Niels; Sulem, Patrick; Sandholt, Camilla H; Thorleifsson, Gudmar; Ahluwalia, Tarunveer S; Steinthorsdottir, Valgerdur; Bjarnason, Helgi; Gudbjartsson, Daniel F; Magnusson, Olafur T; Sparsø, Thomas; Albrechtsen, Anders; Kong, Augustine; Masson, Gisli; Tian, Geng; Cao, Hongzhi; Nie, Chao; Kristiansen, Karsten; Husemoen, Lise Lotte; Thuesen, Betina; Li, Yingrui; Nielsen, Rasmus; Linneberg, Allan; Olafsson, Isleifur; Eyjolfsson, Gudmundur I; Jørgensen, Torben; Wang, Jun; Hansen, Torben; Thorsteinsdottir, Unnur; Stefánsson, Kari; Pedersen, Oluf

    2013-06-01

    Genome-wide association studies have mainly relied on common HapMap sequence variations. Recently, sequencing approaches have allowed analysis of low frequency and rare variants in conjunction with common variants, thereby improving the search for functional variants and thus the understanding of the underlying biology of human traits and diseases. Here, we used a large Icelandic whole genome sequence dataset combined with Danish exome sequence data to gain insight into the genetic architecture of serum levels of vitamin B(12) (B12) and folate. Up to 22.9 million sequence variants were analyzed in combined samples of 45,576 and 37,341 individuals with serum B(12) and folate measurements, respectively. We found six novel loci associating with serum B(12) (CD320, TCN2, ABCD4, MMAA, MMACHC) or folate levels (FOLR3) and confirmed seven loci for these traits (TCN1, FUT6, FUT2, CUBN, CLYBL, MUT, MTHFR). Conditional analyses established that four loci contain additional independent signals. Interestingly, 13 of the 18 identified variants were coding and 11 of the 13 target genes have known functions related to B(12) and folate pathways. Contrary to epidemiological studies we did not find consistent association of the variants with cardiovascular diseases, cancers or Alzheimer's disease although some variants demonstrated pleiotropic effects. Although to some degree impeded by low statistical power for some of these conditions, these data suggest that sequence variants that contribute to the population diversity in serum B(12) or folate levels do not modify the risk of developing these conditions. Yet, the study demonstrates the value of combining whole genome and exome sequencing approaches to ascertain the genetic and molecular architectures underlying quantitative trait associations.

  18. Sequence-specific thermodynamic properties of nucleic acids influence both transcriptional pausing and backtracking in yeast

    PubMed Central

    2017-01-01

    RNA Polymerase II pauses and backtracks during transcription, with many consequences for gene expression and cellular physiology. Here, we show that the energy required to melt double-stranded nucleic acids in the transcription bubble predicts pausing in Saccharomyces cerevisiae far more accurately than nucleosome roadblocks do. In addition, the same energy difference also determines when the RNA polymerase backtracks instead of continuing to move forward. This data-driven model corroborates—in a genome wide and quantitative manner—previous evidence that sequence-dependent thermodynamic features of nucleic acids influence both transcriptional pausing and backtracking. PMID:28301878

  19. Respiratory syncytial virus fusion glycoprotein: nucleotide sequence of mRNA, identification of cleavage activation site and amino acid sequence of N-terminus of F1 subunit.

    PubMed Central

    Elango, N; Satake, M; Coligan, J E; Norrby, E; Camargo, E; Venkatesan, S

    1985-01-01

    The amino acid sequence of respiratory syncytial virus fusion protein (Fo) was deduced from the sequence of a partial cDNA clone of mRNA and from the 5' mRNA sequence obtained by primer extension and dideoxysequencing. The encoded protein of 574 amino acids is extremely hydrophobic and has a molecular weight of 63371 daltons. The site of proteolytic cleavage within this protein was accurately mapped by determining a partial amino acid sequence of the N-terminus of the larger subunit (F1) purified by radioimmunoprecipitation using monoclonal antibodies. Alignment of the N-terminus of the F1 subunit within the deduced amino acid sequence of Fo permitted us to identify a sequence of lys-lys-arg-lys-arg-arg at the C-terminus of the smaller N-terminal F2 subunit that appears to represent the cleavage/activation domain. Five potential sites of glycosylation, four within the F2 subunit, were also identified. Three extremely hydrophobic domains are present in the protein; a) the N-terminal signal sequence, b) the N-terminus of the F1 subunit that is analogous to the N-terminus of the paramyxovirus F1 subunit and the HA2 subunit of influenza virus hemagglutinin, and c) the putative membrane anchorage domain near the C-terminus of F1. Images PMID:2987829

  20. 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

  1. Amino acid sequence of myoglobin from emu (Dromaius novaehollandiae) skeletal muscle.

    PubMed

    Suman, S P; Joseph, P; Li, S; Beach, C M; Fontaine, M; Steinke, L

    2010-11-01

    The objective of the present study was to characterize the primary structure of emu myoglobin (Mb). Emu Mb was isolated from Iliofibularis muscle employing gel-filtration chromatography. Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry was employed to determine the exact molecular mass of emu Mb in comparison with horse Mb, and Edman degradation was utilized to characterize the amino acid sequence. The molecular mass of emu Mb was 17,380 Da and was close to those reported for ratite and poultry myoglobins. Similar to myoglobins from meat-producing livestock and birds, emu Mb has 153 amino acids. Emu Mb contains 9 histidines. Proximal and distal histidines, responsible for coordinating oxygen-binding property of Mb, are conserved in emu. Emu Mb shared more than 90% homology with ratite and chicken myoglobins, whereas it demonstrated only less than 70% sequence similarity with ruminant myoglobins.

  2. Stereochemical Sequence Ion Selectivity: Proline versus Pipecolic-acid-containing Protonated Peptides

    NASA Astrophysics Data System (ADS)

    Abutokaikah, Maha T.; Guan, Shanshan; Bythell, Benjamin J.

    2017-01-01

    Substitution of proline by pipecolic acid, the six-membered ring congener of proline, results in vastly different tandem mass spectra. The well-known proline effect is eliminated and amide bond cleavage C-terminal to pipecolic acid dominates instead. Why do these two ostensibly similar residues produce dramatically differing spectra? Recent evidence indicates that the proton affinities of these residues are similar, so are unlikely to explain the result [Raulfs et al., J. Am. Soc. Mass Spectrom. 25, 1705-1715 (2014)]. An additional hypothesis based on increased flexibility was also advocated. Here, we provide a computational investigation of the "pipecolic acid effect," to test this and other hypotheses to determine if theory can shed additional light on this fascinating result. Our calculations provide evidence for both the increased flexibility of pipecolic-acid-containing peptides, and structural changes in the transition structures necessary to produce the sequence ions. The most striking computational finding is inversion of the stereochemistry of the transition structures leading to "proline effect"-type amide bond fragmentation between the proline/pipecolic acid-congeners: R (proline) to S (pipecolic acid). Additionally, our calculations predict substantial stabilization of the amide bond cleavage barriers for the pipecolic acid congeners by reduction in deleterious steric interactions and provide evidence for the importance of experimental energy regime in rationalizing the spectra.

  3. 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.

  4. Amino acid sequence of atrial natriuretic peptides in human coronary sinus plasma.

    PubMed

    Yandle, T; Crozier, I; Nicholls, G; Espiner, E; Carne, A; Brennan, S

    1987-07-31

    Two atrial natriuretic peptides were purified from pooled human coronary sinus plasma by Sep-Pak extraction, immunoaffinity chromatography and reverse phase HPLC. The amino acid sequences of the two peptides were homologous with 99-126 human atrial natriuretic peptide (hANP) and 106-126 hANP, the latter being most probably linked to 99-105 ANP by the disulphide bond. The molar ratio of the peptides in plasma, as assessed by radioimmunoassay was 10:3.

  5. Amino Acid Sequences Mediating Vascular Cell Adhesion Molecule 1 Binding to Integrin Alpha 4: Homologous DSP Sequence Found for JC Polyoma VP1 Coat Protein

    PubMed Central

    Meyer, Michael Andrew

    2013-01-01

    The JC polyoma viral coat protein VP1 was analyzed for amino acid sequences homologies to the IDSP sequence which mediates binding of VLA-4 (integrin alpha 4) to vascular cell adhesion molecule 1. Although the full sequence was not found, a DSP sequence was located near the critical arginine residue linked to infectivity of the virus and binding to sialic acid containing molecules such as integrins (3). For the JC polyoma virus, a DSP sequence was found at residues 70, 71 and 72 with homology also noted for the mouse polyoma virus and SV40 virus. Three dimensional modeling of the VP1 molecule suggests that the DSP loop has an accessible site for interaction from the external side of the assembled viral capsid pentamer. PMID:24147211

  6. Amino Acid Sequences Mediating Vascular Cell Adhesion Molecule 1 Binding to Integrin Alpha 4: Homologous DSP Sequence Found for JC Polyoma VP1 Coat Protein.

    PubMed

    Meyer, Michael Andrew

    2013-01-01

    The JC polyoma viral coat protein VP1 was analyzed for amino acid sequences homologies to the IDSP sequence which mediates binding of VLA-4 (integrin alpha 4) to vascular cell adhesion molecule 1. Although the full sequence was not found, a DSP sequence was located near the critical arginine residue linked to infectivity of the virus and binding to sialic acid containing molecules such as integrins (3). For the JC polyoma virus, a DSP sequence was found at residues 70, 71 and 72 with homology also noted for the mouse polyoma virus and SV40 virus. Three dimensional modeling of the VP1 molecule suggests that the DSP loop has an accessible site for interaction from the external side of the assembled viral capsid pentamer.

  7. Serum hyaluronic acid levels in patients with ankylosing spondylitis.

    PubMed

    Duruöz, Mehmet Tuncay; Turan, Yasemin; Cerrahoglu, Lale; Isbilen, Banu

    2008-05-01

    Our aim in this study was to investigate serum hyaluronic acid (HA) levels and the relationship between clinical parameters in ankylosing spondylitis (AS). Approximately 30 patients with AS and 30 healthy individuals were recruited in this study consecutively. Cross-sectional study was planned, and demographic, clinical, functional, radiological, and laboratory data of patients were evaluated. Disease activity, functional status, and quality of life were assessed, respectively, with Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI), and Short-Form 36 (SF-36). Mander Enthesis Index (MEI) was used for evaluation of enthesis involvement. We examined serum concentrations of HA (ng/ml) in patients with AS and controls. The mean ages of patients and control group were 38.3 (SD=10.8) and 42.7 (SD=10.6) years, respectively. The mean of serum HA levels in AS patients was 40.4 (SD=34.8) ng/ml and in controls was 24.9 (SD=20.2). There was significant difference of HA levels between two groups (p=0.04). Furthermore, there was a significant correlation between HA level and distance of hand-floor (r=0.444, p=0.014), modified lumbar Schober's (r= -0.413, p=0.023), distance of chin to chest (r=0.436, p=0.016), right sacroiliit grade (r=0.601, p<0.001), left sacroiliit grade (r=0.610, p<0.001), C reactive protein level (r=0.404, p=0.027), albumin (r= -0.464, p=0.010), C3 (p=0.449, p=0.013), and IgA levels (r=0.369, p=0.045). However, there was no significant correlation between HA levels with MEI, BASFI, BASDAI, and SF-36 (p >or= 0.05). Serum HA level was significantly higher in AS patients than controls. However, there was no significant correlation between serum HA level and disease-specific measures as BASFI and BASDAI; it had significant relation with spinal mobility limitation, sacroiliitis, and laboratory parameters related with acute inflammation. The serum HA level may be a potential biomarker of axial

  8. Amino acid sequence similarity between rabies virus glycoprotein and snake venom curaremimetic neurotoxins.

    PubMed

    Lentz, T L; Wilson, P T; Hawrot, E; Speicher, D W

    1984-11-16

    Evidence was presented earlier that a host-cell receptor for the highly neurotropic rabies virus might be the acetylcholine receptor. The amino acid sequence of the glycoprotein of rabies virus was compared by computer analysis with that of snake venom curaremimetic neurotoxins, potent ligands of the acetylcholine receptor. A statistically significant sequence relation was found between a segment of the rabies glycoprotein and the entire sequence of long neurotoxins. The greatest identity occurs with residues considered most important in neurotoxicity, including those interacting with the acetylcholine binding site of the acetylcholine receptor. Because of the similarity between the glycoprotein and the receptor-binding region of the neurotoxins, this region of the viral glycoprotein may function as a recognition site for the acetylcholine receptor. Direct binding of the rabies virus glycoprotein to the acetylcholine receptor could contribute to the neurotropism of this virus.

  9. Partial amino acid sequence of human pancreatic stone protein, a novel pancreatic secretory protein.

    PubMed Central

    Montalto, G; Bonicel, J; Multigner, L; Rovery, M; Sarles, H; De Caro, A

    1986-01-01

    Pancreatic stone protein (PSP) is the major organic component of human pancreatic stones. With the use of monoclonal antibody immunoadsorbents, five immunoreactive forms (PSP-S) with close Mr values (14,000-19,000) were isolated from normal pancreatic juice. By CM-Trisacryl M chromatography the lowest-Mr form (PSP-S1) was separated from the others and some of its molecular characteristics were investigated. The Mr of the PSP-S1 polypeptide chain calculated from the amino acid composition was about 16,100. The N-terminal sequences (40 residues) of PSP and PSP-S1 are identical, which suggests that the peptide backbone is the same for both of these polypeptides. The PSP-S1 sequence was determined up to residue 65 and was found to be different from all other known protein sequences. Images Fig. 1. PMID:3541906

  10. 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.

  11. [MOLECULAR EVOLUTION OF ION CHANNELS: AMINO ACID SEQUENCES AND 3D STRUCTURES].

    PubMed

    Korkosh, V S; Zhorov, B S; Tikhonov, D B

    2016-01-01

    An integral part of modern evolutionary biology is comparative analysis of structure and function of macromolecules such as proteins. The first and critical step to understand evolution of homologous proteins is their amino acid sequence alignment. However, standard algorithms fop not provide unambiguous sequence alignments for proteins of poor homology. More reliable results can be obtained by comparing experimental 3D structures obtained at atomic resolution, for instance, with the aid of X-ray structural analysis. If such structures are lacking, homology modeling is used, which may take into account indirect experimental data on functional roles of individual amino-acid residues. An important problem is that the sequence alignment, which reflects genetic modifications, does not necessarily correspond to the functional homology. The latter depends on three-dimensional structures which are critical for natural selection. Since alignment techniques relying only on the analysis of primary structures carry no information on the functional properties of proteins, including 3D structures into consideration is very important. Here we consider several examples involving ion channels and demonstrate that alignment of their three-dimensional structures can significantly improve sequence alignments obtained by traditional methods.

  12. The amino acid sequence of the aspartate aminotransferase from baker's yeast (Saccharomyces cerevisiae).

    PubMed Central

    Cronin, V B; Maras, B; Barra, D; Doonan, S

    1991-01-01

    1. The single (cytosolic) aspartate aminotransferase was purified in high yield from baker's yeast (Saccharomyces cerevisiae). 2. Amino-acid-sequence analysis was carried out by digestion of the protein with trypsin and with CNBr; some of the peptides produced were further subdigested with Staphylococcus aureus V8 proteinase or with pepsin. Peptides were sequenced by the dansyl-Edman method and/or by automated gas-phase methods. The amino acid sequence obtained was complete except for a probable gap of two residues as indicated by comparison with the structures of counterpart proteins in other species. 3. The N-terminus of the enzyme is blocked. Fast-atom-bombardment m.s. was used to identify the blocking group as an acetyl one. 4. Alignment of the sequence of the enzyme with those of vertebrate cytosolic and mitochondrial aspartate aminotransferases and with the enzyme from Escherichia coli showed that about 25% of residues are conserved between these distantly related forms. 5. Experimental details and confirmatory data for the results presented here are given in a Supplementary Publication (SUP 50164, 25 pages) that has been deposited at the British Library Document Supply Centre, Boston Spa. Wetherby, West Yorkshire LS23 7 BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1991) 273, 5. PMID:1859361

  13. 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.

  14. Late Cretaceous base level lowering in Campanian and Maastrichtian depositional sequences, Kure Beach, North Carolina

    USGS Publications Warehouse

    Harris, W.B.; ,

    2006-01-01

    Campanian through Maastrichtian mixed carbonate and siliciclastic sediments in a 422 m continuous core drilled at Kure Beach, NC provide a record of sea-level change. Based on lithology and stratigraphy, depositional sequences are defined, and calcareous nannofossil zones and 87Sr/86Sr ratios and corresponding ages using the LOWESS Table determined. Campanian and Maastrichtian sediments comprise six depositional sequences. The oldest is Tar Heel 1 and contains calcareous nannofossils that indicate assignment to the upper part of Zones CC18a, CC18c and the lower part of CC19. 87Sr/86 Sr ratios indicate ages from 83.2 to 80.0 Ma or lower Campanian. Tar Heel II contains calcareous nannofossils that indicate assignment to the upper part of Zone CC 19 and CC20. 87Sr/86Sr ratios indicate ages from 78.0 to 76.3 Ma or middle Campanian. Donoho Creek I and II are thin and contain calcareous nannofossils referable to upper Zone CC21 and Zone CC22, and to CC23, respectively. The top of Donoho Creek II marks the Campanian-Maastrichtian boundary. Donoho Creek I 87Sr/86Sr ratios cluster into two groups, and provide ages from 78.0 to 76.2 Ma and 73.7 to 72.3 Ma, respectively. 87Sr/86Sr ratios in Donoho Creek II indicate ages from 71.4 to 69.6 Ma. Two Maastrichtian sequences are present; the lowermost Peedee I contains calcareous nannofossils that place it in Zones CC25a and CC25b. 87Sr/86Sr r ratios indicate an age from 69.3 to 66.9 Ma or late Maastrichtian. Peedee II is assigned to calcareous nannofossil Zone CC26a. 87Sr/86Sr ratios indicate ages from 66.4 to 65.2 Ma or late Maastrichtian. The four Campanian sequences correlate to three depositional sequences in New Jersey; the sequence boundary between upper Campanian Donoho Creek I and Donoho Creek II is not recognized in New Jersey. This boundary is interpreted to result from Gulf Stream impingement and subsequent erosion on the outer shelf. The two Maastrichtian sequences recognized in the Kure Beach core correlate to the two

  15. Endogenous Levels of Abscisic Acid and Decanoic Acid in Dutch Iris Bulbs and the Influence of Abscisic Acid and Decanoic Acid on Iris Meristems Cultured In Vitro1

    PubMed Central

    Doss, Robert P.; Kimura, Yosh; Christian, James K.

    1983-01-01

    Abscisic acid (ABA) and decanoic acid inhibited shoot elongation and floral development of Dutch iris (Iris hollandica Hoog. cv Ideal) meristems cultured in vitro. No synergism with respect to inhibition of leaf growth between ABA and decanoic acid was observed. With monthly harvest dates, from July 10, 1981 to October 10, 1981, there was a progressive decrease in endogenous level of free ABA in `Ideal' iris bulbs. Bulbs subjected to a full set of the usual preplanting storage conditions flowered, on average, 46 days after planting versus 194 days after planting for bulbs planted directly after harvest. ABA levels at harvest were 4- to 5-fold those after the preplanting storage treatment. In general, ABA levels did not correlate well with the length of time from planting until flowering of iris bulbs. Endogenous decanoic acid levels did not follow any pattern with respect to harvest date or postharvest treatment. After the postharvest high temperature treatment, there was about a 3-fold increase in nonscale decanoic acid concentration. Decanoic acid levels, in nonscale tissue, remained high after each of the other postharvest treatments. It is concluded that there is no good evidence to support the contention that either ABA or decanoic acid is directly involved in iris bulb dormancy. PMID:16663072

  16. Momentum sequence and environmental climate influence levels of perceived psychological momentum within a sport competition.

    PubMed

    Briki, Walid; Markman, Keith D; Coudevylle, Guillaume; Sinnapah, Stéphane; Hue, Olivier

    2016-01-01

    The present study examined the influence of momentum sequence (positive vs. negative) and environmental climate (hot-wet vs. neutral) on supporters' (i.e. virtual observers') reported levels of perceived psychological momentum (PM) during a simulated cycling competition. Participants supported one of two competing cyclists involved in a race that was displayed on a screen in a lecture hall. The race scenario was manipulated so that the supported cyclist appeared to undergo either a positive or negative momentum sequence. In addition, participants were either exposed to a hot-wet environmental climate or to a neutral environmental climate while observing the race scenario. According to the results, reported levels of PM were higher in the positive momentum sequence condition than in the negative momentum sequence condition, consistent with the notion that supporters' PM is influenced by a positivity bias, and reported levels of PM were also found to be higher in the hot-wet climate condition than in the neutral climate condition, consistent with the notion that environmental climate is a contextual factor that influences PM through the operation of a causal augmenting mechanism.

  17. Processing and amino acid sequence analysis of the mouse mammary tumor virus env gene product.

    PubMed Central

    Arthur, L O; Copeland, T D; Oroszlan, S; Schochetman, G

    1982-01-01

    The envelope proteins of mouse mammary tumor virus (MMTV) are synthesized from a subgenomic 24S mRNA as a 75,000-dalton glycosylated precursor polyprotein which is eventually processed to the mature glycoproteins gp52 and gp36. In vivo synthesis of this env precursor in the presence of the core glycosylation inhibitor tunicamycin yielded a precursor of approximately 61,000 daltons (P61env). However, a 67,000-dalton protein (P67env) was obtained from cell-free translation with the MMTV 24S mRNA as the template. To determine whether the portion of the protein cleaved from P67env to give P61env was removed from the NH2-terminal end of P67env and as such would represent a leader sequence, the NH2-terminal amino acid sequence of the terminal peptide gp52 was determined. Glutamic acid, and not methionine, was found to be the amino-terminal residue of gp52, indicating that the cleaved portion was derived from the NH2-terminal end of P67env. The NH2-terminal amino acid sequences of gp52's from endogenous and exogenous C3H MMTVs were determined though 46 residues and found to be identical. However, amino acid composition and type-specific gp52 radioimmunoassays from MMTVs grown in heterologous cells indicated primary structure differences between gp52's of the two viruses. The nucleic acid sequence of cloned MMTV DNA fragments (J. Majors and H. E. Varmus, personal communication) in conjunction with the NH2-terminal sequence of gp52 allowed localization of the env gene in the MMTV genome. Nucleotides coding for the NH2 terminus of gp52 begin approximately 0.8 kilobase to the 3' side of the single EcoRI cleavage site. Localization of the env gene at that point agrees with the proposed gene order -gag-pol-env- and also allows sufficient coding potential for the glycoprotein precursor without extending into the long terminal repeat. Images PMID:6281457

  18. 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

  19. BeadCons: detection of nucleic acid sequences by flow cytometry.

    PubMed

    Horejsh, Douglas; Martini, Federico; Capobianchi, Maria Rosaria

    2005-11-01

    Molecular beacons are single-stranded nucleic acid structures with a terminal fluorophore and a distal, terminal quencher. These molecules are typically used in real-time PCR assays, but have also been conjugated with solid matrices. This unit describes protocols related to molecular beacon-conjugated beads (BeadCons), whose specific hybridization with complementary target sequences can be resolved by cytometry. Assay sensitivity is achieved through the concentration of fluorescence signal on discrete particles. By using molecular beacons with different fluorophores and microspheres of different sizes, it is possible to construct a fluid array system with each bead corresponding to a specific target nucleic acid. Methods are presented for the design, construction, and use of BeadCons for the specific, multiplexed detection of unlabeled nucleic acids in solution. The use of bead-based detection methods will likely lead to the design of new multiplex molecular diagnostic tools.

  20. Measuring nanometer distances in nucleic acids using a sequence-independent nitroxide probe

    PubMed Central

    Qin, Peter Z; Haworth, Ian S; Cai, Qi; Kusnetzow, Ana K; Grant, Gian Paola G; Price, Eric A; Sowa, Glenna Z; Popova, Anna; Herreros, Bruno; He, Honghang

    2008-01-01

    This protocol describes the procedures for measuring nanometer distances in nucleic acids using a nitroxide probe that can be attached to any nucleotide within a given sequence. Two nitroxides are attached to phosphorothioates that are chemically substituted at specific sites of DNA or RNA. Inter-nitroxide distances are measured using a four-pulse double electron–electron resonance technique, and the measured distances are correlated to the parent structures using a Web-accessible computer program. Four to five days are needed for sample labeling, purification and distance measurement. The procedures described herein provide a method for probing global structures and studying conformational changes of nucleic acids and protein/nucleic acid complexes. PMID:17947978

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

    PubMed

    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

    2011-12-31

    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.

  2. 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.

  3. Low irradiances affect abscisic acid, indole-3-acidic acid, and cytokinin levels of wheat (Triticum aestivum L.) tissues

    NASA Technical Reports Server (NTRS)

    Nan, R.; Carman, J. G.; Salisbury, F. B.

    1999-01-01

    Wheat (Triticum aestivum L.) plants were grown under four irradiance levels: 1,400, 400, 200, and 100 micromol m-2 s-1. Leaves and roots were sampled before, during, and after the boot stage, and levels of abscisic acid (ABA), indole-3-acetic acid (IAA), zeatin, zeatin riboside, dihydrozeatin, dihydrozeatin riboside, isopentenyl adenine, and isopentenyl adenosine were quantified using noncompetitive indirect ELISA systems. Levels of IAA in leaves and roots of plants exposed to 100 micromol m-2 s-1 of irradiance were 0.7 and 2.9 micromol kg-1 dry mass (DM), respectively. These levels were 0.2 and 1.0 micromol kg-1 DM, respectively, when plants were exposed to 1,400 micromol m-2 s-1. Levels of ABA in leaves and roots of plants exposed to 100 micromol m-2 s-1 were 0.65 and 0.55 micromol kg-1 DM, respectively. They were 0.24 micromol kg-1 DM (both leaves and roots) when plants were exposed to 1,400 micromol m-2 s-1. Levels of isopentenyl adenosine in leaves (24.3 nmol kg-1 DM) and roots (29.9 nmol kg-1 DM) were not affected by differences in the irradiance regime. Similar values were obtained in a second experiment. Other cytokinins could not be detected (<10 nmol kg 1 DM) in either experiment with the sample sizes used (150-600 mg DM for roots and shoots, respectively).

  4. SigniSite: Identification of residue-level genotype-phenotype correlations in protein multiple sequence alignments

    PubMed Central

    Jessen, Leon Eyrich; Hoof, Ilka; Lund, Ole; Nielsen, Morten

    2013-01-01

    Identifying which mutation(s) within a given genotype is responsible for an observable phenotype is important in many aspects of molecular biology. Here, we present SigniSite, an online application for subgroup-free residue-level genotype–phenotype correlation. In contrast to similar methods, SigniSite does not require any pre-definition of subgroups or binary classification. Input is a set of protein sequences where each sequence has an associated real number, quantifying a given phenotype. SigniSite will then identify which amino acid residues are significantly associated with the data set phenotype. As output, SigniSite displays a sequence logo, depicting the strength of the phenotype association of each residue and a heat-map identifying ‘hot’ or ‘cold’ regions. SigniSite was benchmarked against SPEER, a state-of-the-art method for the prediction of specificity determining positions (SDP) using a set of human immunodeficiency virus protease-inhibitor genotype–phenotype data and corresponding resistance mutation scores from the Stanford University HIV Drug Resistance Database, and a data set of protein families with experimentally annotated SDPs. For both data sets, SigniSite was found to outperform SPEER. SigniSite is available at: http://www.cbs.dtu.dk/services/SigniSite/. PMID:23761454

  5. Alterations of amino Acid level in depressed rat brain.

    PubMed

    Yang, Pei; Li, Xuechun; Ni, Jian; Tian, Jingchen; Jing, Fu; Qu, Changhai; Lin, Longfei; Zhang, Hui

    2014-10-01

    Amino-acid neurotransmitter system dysfunction plays a major role in the pathophysiology of depression. Several studies have demonstrated the potential of amino acids as a source of neuro-specific biomarkers could be used in future diagnosis of depression. Only partial amino acids such as glycine and asparagine were determined from certain parts of rats' brain included hippocampi and cerebral cortex in previous studies. However, according to systematic biology, amino acids in different area of brain are interacted and interrelated. Hence, the determination of 34 amino acids through entire rats' brain was conducted in this study in order to demonstrate more possibilities for biomarkers of depression by discovering other potential amino acids in more areas of rats' brain. As a result, 4 amino acids (L-aspartic acid, L-glutamine, taurine and γ-amino-n-butyric acid) among 34 were typically identified as potentially primary biomarkers of depression by data statistics. Meanwhile, an antidepressant called Fluoxetine was employed to verify other potential amino acids which were not identified by data statistics. Eventually, we found L-α-amino-adipic acid could also become a new potentially secondary biomarker of depression after drug validation. In conclusion, we suggested that L-aspartic acid, L-glutamine, taurine, γ-amino-n-butyric acid and L-α-amino-adipic acid might become potential biomarkers for future diagnosis of depression and development of antidepressant.

  6. A 25-Amino Acid Sequence of the Arabidopsis TGD2 Protein Is Sufficient for Specific Binding of Phosphatidic Acid*

    PubMed Central

    Lu, Binbin; Benning, Christoph

    2009-01-01

    Genetic analysis suggests that the TGD2 protein of Arabidopsis is required for the biosynthesis of endoplasmic reticulum derived thylakoid lipids. TGD2 is proposed to be the substrate-binding protein of a presumed lipid transporter consisting of the TGD1 (permease) and TGD3 (ATPase) proteins. The TGD1, -2, and -3 proteins are localized in the inner chloroplast envelope membrane. TGD2 appears to be anchored with an N-terminal membrane-spanning domain into the inner envelope membrane, whereas the C-terminal domain faces the intermembrane space. It was previously shown that the C-terminal domain of TGD2 binds phosphatidic acid (PtdOH). To investigate the PtdOH binding site of TGD2 in detail, the C-terminal domain of the TGD2 sequence lacking the transit peptide and transmembrane sequences was fused to the C terminus of the Discosoma sp. red fluorescent protein (DR). This greatly improved the solubility of the resulting DR-TGD2C fusion protein following production in Escherichia coli. The DR-TGD2C protein bound PtdOH with high specificity, as demonstrated by membrane lipid-protein overlay and liposome association assays. Internal deletion and truncation mutagenesis identified a previously undescribed minimal 25-amino acid fragment in the C-terminal domain of TGD2 that is sufficient for PtdOH binding. Binding characteristics of this 25-mer were distinctly different from those of TGD2C, suggesting that additional sequences of TGD2 providing the proper context for this 25-mer are needed for wild type-like PtdOH binding. PMID:19416982

  7. Nucleotide sequence of the luxC gene encoding fatty acid reductase of the lux operon from Photobacterium leiognathi.

    PubMed

    Lin, J W; Chao, Y F; Weng, S F

    1993-02-26

    The nucleotide sequence of the luxC gene (EMBL Accession No. 65156) encoding fatty acid reductase (FAR) of the lux operon from Photobacterium leiognathi PL741 was determined and the encoded amino acid sequence deduced. The fatty acid reductase is a component of the fatty acid reductase complex. The complex is responsible for converting fatty acid to aldehyde which serves as the substrate in the luciferase-catalyzed bioluminescent reaction. The protein comprises 478 amino acid residues and has a calculated M(r) of 53,858. Alignment and comparison of the fatty acid reductase of P. leiognathi with that of Vibrio harveyi B392 and Vibrio fischeri ATCC 7744 shows that there is 70% and 59% amino acid residues identity, respectively.

  8. Elevation of serum levels of beta-aminoisobutyric acid in uremic patients and the toxicity of the amino acid.

    PubMed

    Gejyo, F; Kinoshita, Y; Ikenaka, T

    1977-12-01

    A reliable method for the determination of beta-aminoisobutyric acid in serum was developed utilizing an automated amino acid analyzer. The serum concentrations of beta-aminoisobutyric acid were determined in 20 normal subjects and in 71 uremic patients. The mean serum level of beta-aminoisobutyric acid was markedly increased in the uremic patients to 0.856 +/- 0.910 (mean +/- SD) mg/100 ml as compared with a normal value of 0.026 +/- 0.027 mg/100 ml. The distribution of serum beta-aminoisobutyric acid level in uremic patients was wide-spread, and there was no correlation between the serum levels of the amino acid and those of urea nitrogen, creatinine and uric acid. The toxicity of beta-aminoisobutyric acid on mice with acute renal failure induced by uranyl acetate was investigated and compared with that of alpha-amino-n-butyric acid and gamma-amino-n-butyric acid. All mice given more than 4 g/kg body wt of beta-aminoisobutyric acid showed twitching and cramps, and some of them died. However, the control mice given an equivalent dose of alpha-amino-n-butyric acid or gamma-amino-n-butyric acid showed no change. These results suggest that beta-aminoisobutyric acid might be a factor influencing the development and progression of uremic toxemia.

  9. Intermediate divergence levels maximize the strength of structure-sequence correlations in enzymes and viral proteins.

    PubMed

    Jackson, Eleisha L; Shahmoradi, Amir; Spielman, Stephanie J; Jack, Benjamin R; Wilke, Claus O

    2016-07-01

    Structural properties such as solvent accessibility and contact number predict site-specific sequence variability in many proteins. However, the strength and significance of these structure-sequence relationships vary widely among different proteins, with absolute correlation strengths ranging from 0 to 0.8. In particular, two recent works have made contradictory observations. Yeh et al. (Mol. Biol. Evol. 31:135-139, 2014) found that both relative solvent accessibility (RSA) and weighted contact number (WCN) are good predictors of sitewise evolutionary rate in enzymes, with WCN clearly out-performing RSA. Shahmoradi et al. (J. Mol. Evol. 79:130-142, 2014) considered these same predictors (as well as others) in viral proteins and found much weaker correlations and no clear advantage of WCN over RSA. Because these two studies had substantial methodological differences, however, a direct comparison of their results is not possible. Here, we reanalyze the datasets of the two studies with one uniform analysis pipeline, and we find that many apparent discrepancies between the two analyses can be attributed to the extent of sequence divergence in individual alignments. Specifically, the alignments of the enzyme dataset are much more diverged than those of the virus dataset, and proteins with higher divergence exhibit, on average, stronger structure-sequence correlations. However, the highest structure-sequence correlations are observed at intermediate divergence levels, where both highly conserved and highly variable sites are present in the same alignment.

  10. Phylum-Level Conservation of Regulatory Information in Nematodes despite Extensive Non-coding Sequence Divergence.

    PubMed

    Gordon, Kacy L; Arthur, Robert K; Ruvinsky, Ilya

    2015-05-01

    Gene regulatory information guides development and shapes the course of evolution. To test conservation of gene regulation within the phylum Nematoda, we compared the functions of putative cis-regulatory sequences of four sets of orthologs (unc-47, unc-25, mec-3 and elt-2) from distantly-related nematode species. These species, Caenorhabditis elegans, its congeneric C. briggsae, and three parasitic species Meloidogyne hapla, Brugia malayi, and Trichinella spiralis, represent four of the five major clades in the phylum Nematoda. Despite the great phylogenetic distances sampled and the extensive sequence divergence of nematode genomes, all but one of the regulatory elements we tested are able to drive at least a subset of the expected gene expression patterns. We show that functionally conserved cis-regulatory elements have no more extended sequence similarity to their C. elegans orthologs than would be expected by chance, but they do harbor motifs that are important for proper expression of the C. elegans genes. These motifs are too short to be distinguished from the background level of sequence similarity, and while identical in sequence they are not conserved in orientation or position. Functional tests reveal that some of these motifs contribute to proper expression. Our results suggest that conserved regulatory circuitry can persist despite considerable turnover within cis elements.

  11. Phylum-Level Conservation of Regulatory Information in Nematodes despite Extensive Non-coding Sequence Divergence

    PubMed Central

    Gordon, Kacy L.; Arthur, Robert K.; Ruvinsky, Ilya

    2015-01-01

    Gene regulatory information guides development and shapes the course of evolution. To test conservation of gene regulation within the phylum Nematoda, we compared the functions of putative cis-regulatory sequences of four sets of orthologs (unc-47, unc-25, mec-3 and elt-2) from distantly-related nematode species. These species, Caenorhabditis elegans, its congeneric C. briggsae, and three parasitic species Meloidogyne hapla, Brugia malayi, and Trichinella spiralis, represent four of the five major clades in the phylum Nematoda. Despite the great phylogenetic distances sampled and the extensive sequence divergence of nematode genomes, all but one of the regulatory elements we tested are able to drive at least a subset of the expected gene expression patterns. We show that functionally conserved cis-regulatory elements have no more extended sequence similarity to their C. elegans orthologs than would be expected by chance, but they do harbor motifs that are important for proper expression of the C. elegans genes. These motifs are too short to be distinguished from the background level of sequence similarity, and while identical in sequence they are not conserved in orientation or position. Functional tests reveal that some of these motifs contribute to proper expression. Our results suggest that conserved regulatory circuitry can persist despite considerable turnover within cis elements. PMID:26020930

  12. Stacking sequence and shape optimization of laminated composite plates via a level-set method

    NASA Astrophysics Data System (ADS)

    Allaire, G.; Delgado, G.

    2016-12-01

    We consider the optimal design of composite laminates by allowing a variable stacking sequence and in-plane shape of each ply. In order to optimize both variables we rely on a decomposition technique which aggregates the constraints into one unique constraint margin function. Thanks to this approach, an exactly equivalent bi-level optimization problem is established. This problem is made up of an inner level represented by the combinatorial optimization of the stacking sequence and an outer level represented by the topology and geometry optimization of each ply. We propose for the stacking sequence optimization an outer approximation method which iteratively solves a set of mixed integer linear problems associated to the evaluation of the constraint margin function. For the topology optimization of each ply, we lean on the level set method for the description of the interfaces and the Hadamard method for boundary variations by means of the computation of the shape gradient. Numerical experiments are performed on an aeronautic test case where the weight is minimized subject to different mechanical constraints, namely compliance, reserve factor and buckling load.

  13. Serum uric acid levels during leprosy reaction episodes

    PubMed Central

    Alves-Junior, Eduardo R.; Arruda, Talita A.; Lopes, Jose C.; Fontes, Cor J.F.

    2016-01-01

    Background. Leprosy reactions are acute inflammatory episodes that occur mainly in the multibacillary forms of the disease. The reactions are classified as type 1 (reverse reaction) or type 2 (erythema nodosum leprosum). Leprosy-associated oxidative stress has been widely demonstrated. Several recent studies have shown uric acid (UA) to have antioxidative effects under pathologic conditions. The objective of this study was to assess serum levels of UA in patients with leprosy reactions, with the aim of monitoring their levels before and after treatment, compared with levels in leprosy patients without reactions. Methods. The study included patients aged 18–69 years assisted at a leprosy treatment reference center in the Central Region of Brazil. Patients who were pregnant; were using immunosuppressant drugs or immunobiologicals; or had an autoimmune disease, human immunodeficiency virus infection, acquired immune deficiency syndrome, or tuberculosis were excluded. Upon recruitment, all individuals were clinically assessed for skin lesions and neural or systemic impairment. Some patients had already completed treatment for leprosy, while others were still undergoing treatment or had initiated treatment after being admitted. The treatment of the reactional episode was started only after the initial evaluation. Laboratory assessments were performed upon admission (baseline) and at approximately 30 and 60 days (time points 1 and 2, respectively). Results. A total of 123 leprosy patients were recruited between June 2012 and June 2015; among them, 56, 42, and 25 presented with type 1, type 2, and no reactions, respectively. Serum UA levels were significantly reduced in patients with type 2 leprosy reactions compared with patients in the control group and remained lower in the two subsequent assessments, after initiation of anti-reaction treatments, with similar values to those recorded before the treatment. Discussion. The decreased serum UA levels in patients with

  14. Nucleotide sequence of the Klebsiella pneumoniae nifD gene and predicted amino acid sequence of the alpha-subunit of nitrogenase MoFe protein.

    PubMed Central

    Ioannidis, I; Buck, M

    1987-01-01

    The nucleotide sequence of the Klebsiella pneumoniae nifD gene is presented and together with the accompanying paper [Holland, Zilberstein, Zamir & Sussman (1987) Biochem. J. 247, 277-285] completes the sequence of the nifHDK genes encoding the nitrogenase polypeptides. The K. pneumoniae nifD gene encodes the 483-amino acid-residue nitrogenase alpha-subunit polypeptide of Mr 54156. The alpha-subunit has five strongly conserved cysteine residues at positions 63, 89, 155, 184 and 275, some occurring in a region showing both primary sequence and potential structural homology to the K. pneumoniae nitrogenase beta-subunit. A comparison with six other alpha-subunit amino acid sequences has been made, which indicates a number of potentially important domains within alpha-subunits. PMID:3322262

  15. Complete amino acid sequence of the A chain of human complement-classical-pathway enzyme C1r.

    PubMed Central

    Arlaud, G J; Willis, A C; Gagnon, J

    1987-01-01

    The amino acid sequence of human C1r A chain was determined, from sequence analysis performed on fragments obtained from C1r autolytic cleavage, cleavage of methionyl bonds, tryptic cleavages at arginine and lysine residues, and cleavages by staphylococcal proteinase. The polypeptide chain has an N-terminal serine residue and contains 446 amino acid residues (Mr 51,200). The sequence data allow chemical characterization of fragments alpha (positions 1-211), beta (positions 212-279) and gamma (positions 280-446) yielded from C1r autolytic cleavage, and identification of the two major cleavage sites generating these fragments. Position 150 of C1r A chain is occupied by a modified amino acid residue that, upon acid hydrolysis, yields erythro-beta-hydroxyaspartic acid, and that is located in a sequence homologous to the beta-hydroxyaspartic acid-containing regions of Factor IX, Factor X, protein C and protein Z. Sequence comparison reveals internal homology between two segments (positions 10-78 and 186-257). Two carbohydrate moieties are attached to the polypeptide chain, both via asparagine residues at positions 108 and 204. Combined with the previously determined sequence of C1r B chain [Arlaud & Gagnon (1983) Biochemistry 22, 1758-1764], these data give the complete sequence of human C1r. PMID:3036070

  16. Mutation-selection models of coding sequence evolution with site-heterogeneous amino acid fitness profiles

    PubMed Central

    Rodrigue, Nicolas; Philippe, Hervé; Lartillot, Nicolas

    2010-01-01

    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

  17. Upper Cretaceous sequences and sea-level history, New Jersey Coastal Plain

    USGS Publications Warehouse

    Miller, K.G.; Sugarman, P.J.; Browning, J.V.; Kominz, M.A.; Olsson, R.K.; Feigenson, M.D.; Hernandez, J.C.

    2004-01-01

    We developed a Late Cretaceous sealevel estimate from Upper Cretaceous sequences at Bass River and Ancora, New Jersey (ODP [Ocean Drilling Program] Leg 174AX). We dated 11-14 sequences by integrating Sr isotope and biostratigraphy (age resolution ??0.5 m.y.) and then estimated paleoenvironmental changes within the sequences from lithofacies and biofacies analyses. Sequences generally shallow upsection from middle-neritic to inner-neritic paleodepths, as shown by the transition from thin basal glauconite shelf sands (transgressive systems tracts [TST]), to medial-prodelta silty clays (highstand systems tracts [HST]), and finally to upper-delta-front quartz sands (HST). Sea-level estimates obtained by backstripping (accounting for paleodepth variations, sediment loading, compaction, and basin subsidence) indicate that large (>25 m) and rapid (???1 m.y.) sea-level variations occurred during the Late Cretaceous greenhouse world. The fact that the timing of Upper Cretaceous sequence boundaries in New Jersey is similar to the sea-level lowering records of Exxon Production Research Company (EPR), northwest European sections, and Russian platform outcrops points to a global cause. Because backstripping, seismicity, seismic stratigraphic data, and sediment-distribution patterns all indicate minimal tectonic effects on the New Jersey Coastal Plain, we interpret that we have isolated a eustatic signature. The only known mechanism that can explain such global changes-glacio-eustasy-is consistent with foraminiferal ??18O data. Either continental ice sheets paced sea-level changes during the Late Cretaceous, or our understanding of causal mechanisms for global sea-level change is fundamentally flawed. Comparison of our eustatic history with published ice-sheet models and Milankovitch predictions suggests that small (5-10 ?? 106 km3), ephemeral, and areally restricted Antarctic ice sheets paced the Late Cretaceous global sea-level change. New Jersey and Russian eustatic estimates

  18. Acetic acid vapor levels associated with facial prosthetics

    SciTech Connect

    McElroy, T.H.; Guerra, O.N.; Lee, S.A.

    1985-01-01

    The use of Silastic Medical Adhesive Type A in the fabrication of facial prostheses may cause health hazards to the patient and the operator because of acetic acid emissions. Caution must be exercised to remove acetic acid vapors from the air and unliberated acetic acid from material applied directly to the skin.

  19. Nucleotide sequences of the Pseudomonas savastanoi indoleacetic acid genes show homology with Agrobacterium tumefaciens T-DNA

    PubMed Central

    Yamada, Tetsuji; Palm, Curtis J.; Brooks, Bob; Kosuge, Tsune

    1985-01-01

    We report the nucleotide sequences of iaaM and iaaH, the genetic determinants for, respectively, tryptophan 2-monooxygenase and indoleacetamide hydrolase, the enzymes that catalyze the conversion of L-tryptophan to indoleacetic acid in the tumor-forming bacterium Pseudomonas syringae pv. savastanoi. The sequence analysis indicates that the iaaM locus contains an open reading frame encoding 557 amino acids that would comprise a protein with a molecular weight of 61,783; the iaaH locus contains an open reading frame of 455 amino acids that would comprise a protein with a molecular weight of 48,515. Significant amino acid sequence homology was found between the predicted sequence of the tryptophan monooxygenase of P. savastanoi and the deduced product of the T-DNA tms-1 gene of the octopine-type plasmid pTiA6NC from Agrobacterium tumefaciens. Strong homology was found in the 25 amino acid sequence in the putative FAD-binding region of tryptophan monooxygenase. Homology was also found in the amino acid sequences representing the central regions of the putative products of iaaH and tms-2 T-DNA. The results suggest a strong similarity in the pathways for indoleacetic acid synthesis encoded by genes in P. savastanoi and in A. tumefaciens T-DNA. Images PMID:16593610

  20. Lower serum uric acid level predicts mortality in dialysis patients

    PubMed Central

    Bae, Eunjin; Cho, Hyun-Jeong; Shin, Nara; Kim, Sun Moon; Yang, Seung Hee; Kim, Dong Ki; Kim, Yong-Lim; Kang, Shin-Wook; Yang, Chul Woo; Kim, Nam Ho; Kim, Yon Su; Lee, Hajeong

    2016-01-01

    Abstract We evaluated the impact of serum uric acid (SUA) on mortality in patients with chronic dialysis. A total of 4132 adult patients on dialysis were enrolled prospectively between August 2008 and September 2014. Among them, we included 1738 patients who maintained dialysis for at least 3 months and had available SUA in the database. We categorized the time averaged-SUA (TA-SUA) into 5 groups: <5.5, 5.5–6.4, 6.5–7.4, 7.5–8.4, and ≥8.5 mg/dL. Cox regression analysis was used to calculate the hazard ratio (HR) of all-cause mortality according to SUA group. The mean TA-SUA level was slightly higher in men than in women. Patients with lower TA-SUA level tended to have lower body mass index (BMI), phosphorus, serum albumin level, higher proportion of diabetes mellitus (DM), and higher proportion of malnourishment on the subjective global assessment (SGA). During a median follow-up of 43.9 months, 206 patients died. Patients with the highest SUA had a similar risk to the middle 3 TA-SUA groups, but the lowest TA-SUA group had a significantly elevated HR for mortality. The lowest TA-SUA group was significantly associated with increased all-cause mortality (adjusted HR, 1.720; 95% confidence interval, 1.007–2.937; P = 0.047) even after adjusting for demographic, comorbid, nutritional covariables, and medication use that could affect SUA levels. This association was prominent in patients with well nourishment on the SGA, a preserved serum albumin level, a higher BMI, and concomitant DM although these parameters had no significant interaction in the TA-SUA-mortality relationship except DM. In conclusion, a lower TA-SUA level <5.5 mg/dL predicted all-cause mortality in patients with chronic dialysis. PMID:27310949

  1. Prediction of flexible/rigid regions from protein sequences using k-spaced amino acid pairs

    PubMed Central

    Chen, Ke; Kurgan, Lukasz A; Ruan, Jishou

    2007-01-01

    Background Traditionally, it is believed that the native structure of a protein corresponds to a global minimum of its free energy. However, with the growing number of known tertiary (3D) protein structures, researchers have discovered that some proteins can alter their structures in response to a change in their surroundings or with the help of other proteins or ligands. Such structural shifts play a crucial role with respect to the protein function. To this end, we propose a machine learning method for the prediction of the flexible/rigid regions of proteins (referred to as FlexRP); the method is based on a novel sequence representation and feature selection. Knowledge of the flexible/rigid regions may provide insights into the protein folding process and the 3D structure prediction. Results The flexible/rigid regions were defined based on a dataset, which includes protein sequences that have multiple experimental structures, and which was previously used to study the structural conservation of proteins. Sequences drawn from this dataset were represented based on feature sets that were proposed in prior research, such as PSI-BLAST profiles, composition vector and binary sequence encoding, and a newly proposed representation based on frequencies of k-spaced amino acid pairs. These representations were processed by feature selection to reduce the dimensionality. Several machine learning methods for the prediction of flexible/rigid regions and two recently proposed methods for the prediction of conformational changes and unstructured regions were compared with the proposed method. The FlexRP method, which applies Logistic Regression and collocation-based representation with 95 features, obtained 79.5% accuracy. The two runner-up methods, which apply the same sequence representation and Support Vector Machines (SVM) and Naïve Bayes classifiers, obtained 79.2% and 78.4% accuracy, respectively. The remaining considered methods are characterized by accuracies below 70

  2. 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.

  3. Shewanella olleyana sp. nov., a marine species isolated from a temperate estuary which produces high levels of polyunsaturated fatty acids.

    PubMed

    Skerratt, Jennifer H; Bowman, John P; Nichols, Peter D

    2002-11-01

    Two polyunsaturated fatty acid (PUFA) producing strains (ACEM 6 and ACEM 9(T)) isolated from a temperate, humic-rich river estuary in Tasmania, Australia, were found to be members of the genus Shewanella. These strains were able to utilize humic compounds (tannic acid) and derivatives (2,6-anthraquinone disulfonate) as sole carbon sources and as electron acceptors for anaerobic respiration. The major fatty acids were typical of the genus Shewanella; however, PUFAs mostly made up of eicosapentaenoic acid were produced at high levels (10.2-23.6% of total fatty acids) and at relatively high incubation temperatures (10.2% at 24 degrees C). Sequence analysis indicated that ACEM 6 and ACEM 9(T) had identical 16S rDNA sequences and were most closely related to Shewanella japonica (sequence similarity 97.1%). DNA hybridization and phenotypic characteristics confirmed that the isolates constituted a novel species of the genus Shewanella, which is designated Shewanella olleyana sp. nov. (type strain ACEM 9(T) = ACAM 644(T) = LMG 21437(T)).

  4. 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

  5. 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.

  6. 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.

  7. 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.

  8. 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.

  9. 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.

  10. Complete amino acid sequence of ananain and a comparison with stem bromelain and other plant cysteine proteases.

    PubMed Central

    Lee, K L; Albee, K L; Bernasconi, R J; Edmunds, T

    1997-01-01

    The amino acid sequences of ananain (EC3.4.22.31) and stem bromelain (3.4.22.32), two cysteine proteases from pineapple stem, are similar yet ananain and stem bromelain possess distinct specificities towards synthetic peptide substrates and different reactivities towards the cysteine protease inhibitors E-64 and chicken egg white cystatin. We present here the complete amino acid sequence of ananain and compare it with the reported sequences of pineapple stem bromelain, papain and chymopapain from papaya and actinidin from kiwifruit. Ananain is comprised of 216 residues with a theoretical mass of 23464 Da. This primary structure includes a sequence insert between residues 170 and 174 not present in stem bromelain or papain and a hydrophobic series of amino acids adjacent to His-157. It is possible that these sequence differences contribute to the different substrate and inhibitor specificities exhibited by ananain and stem bromelain. PMID:9355753

  11. Microbial community dynamics in bioaugmented sequencing batch reactors for bromoamine acid removal.

    PubMed

    Qu, Yuanyuan; Zhou, Jiti; Wang, Jing; Fu, Xiang; Xing, Linlin

    2005-05-01

    Sphingomonas xenophaga QYY with the ability to degrade bromoamine acid (BAA) was previously isolated from sludge samples. The enhancement of BAA removal by strain QYY in sequencing batch reactors (SBRs) was investigated in this study. The results showed that augmented SBRs exhibited stronger abilities to degrade BAA than the non-augmented control one. In order to estimate the relationship between community dynamics and function of augmented SBRs, a combined method based on fingerprints (ribosomal intergenic spacer analysis, RISA) and 16S rRNA gene sequencing was used. The results indicated that the microbial community dynamics were substantially changed, and the introduced strain QYY was persistent in the augmented systems. This study suggests that it is feasible and potentially useful to enhance BAA removal using BAA-degrading bacteria, such as S. xenophaga QYY.

  12. Higher transcription levels in ascorbic acid biosynthetic and recycling genes were associated with higher ascorbic acid accumulation in blueberry.

    PubMed

    Liu, Fenghong; Wang, Lei; Gu, Liang; Zhao, Wei; Su, Hongyan; Cheng, Xianhao

    2015-12-01

    In our preliminary study, the ripe fruits of two highbush blueberry (Vaccinium corymbosum L.) cultivars, cv 'Berkeley' and cv 'Bluecrop', were found to contain different levels of ascorbic acid. However, factors responsible for these differences are still unknown. In the present study, ascorbic acid content in fruits was compared with expression profiles of ascorbic acid biosynthetic and recycling genes between 'Bluecrop' and 'Berkeley' cultivars. The results indicated that the l-galactose pathway was the predominant route of ascorbic acid biosynthesis in blueberry fruits. Moreover, higher expression levels of the ascorbic acid biosynthetic genes GME, GGP, and GLDH, as well as the recycling genes MDHAR and DHAR, were associated with higher ascorbic acid content in 'Bluecrop' compared with 'Berkeley', which indicated that a higher efficiency ascorbic acid biosynthesis and regeneration was likely to be responsible for the higher ascorbic acid accumulation in 'Bluecrop'.

  13. [Measurement of the amino acid sequence for the fusion protein FP3 with LC-MS/MS].

    PubMed

    Li, Xiang; Gao, Xiang-Dong; Tao, Lei; Pei, De-Ning; Guo, Ying; Rao, Chun-Ming; Wang, Jun-Zhi

    2012-02-01

    The amino acid sequence of the fusion protein FP3 was measured by two types of LC-MS/MS and its primary structure was confirmed. After reduction and alkylation, the protein was digested with trypsin and glycosyl groups in glycopeptide were removed by PNGase F. The mixed peptides were separated by LC, then Q-TOF and Ion trap tandem mass spectrometry were used to measure b, y fragment ions of each peptide to analyze the amino acid sequence of fusion protein FP3. Seventy-six percent of full amino acid sequence of the fusion protein FP3 was measured by LC-ESI-Q-TOF with the remaining 24% completed by LC-ESI-Trap. As LC-MS and tandem mass spectrometry are rapid, sensitive, accurate to measure the protein amino acid sequence, they are important approach to structure analysis and identification of recombinant protein.

  14. NullSeq: A Tool for Generating Random Coding Sequences with Desired Amino Acid and GC Contents

    PubMed Central

    Liu, Sophia S.; Hockenberry, Adam J.; Lancichinetti, Andrea; Jewett, Michael C.

    2016-01-01

    The existence of over- and under-represented sequence motifs in genomes provides evidence of selective evolutionary pressures on biological mechanisms such as transcription, translation, ligand-substrate binding, and host immunity. In order to accurately identify motifs and other genome-scale patterns of interest, it is essential to be able to generate accurate null models that are appropriate for the sequences under study. While many tools have been developed to create random nucleotide sequences, protein coding sequences are subject to a unique set of constraints that complicates the process of generating appropriate null models. There are currently no tools available that allow users to create random coding sequences with specified amino acid composition and GC content for the purpose of hypothesis testing. Using the principle of maximum entropy, we developed a method that generates unbiased random sequences with pre-specified amino acid and GC content, which we have developed into a python package. Our method is the simplest way to obtain maximally unbiased random sequences that are subject to GC usage and primary amino acid sequence constraints. Furthermore, this approach can easily be expanded to create unbiased random sequences that incorporate more complicated constraints such as individual nucleotide usage or even di-nucleotide frequencies. The ability to generate correctly specified null models will allow researchers to accurately identify sequence motifs which will lead to a better understanding of biological processes as well as more effective engineering of biological systems. PMID:27835644

  15. 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.

  16. Dependence of intestinal amino acid uptake on dietary protein or amino acid levels

    SciTech Connect

    Karasov, W.H.; Solberg, D.H.; Diamond, J.M.

    1987-05-01

    To understand how intestinal amino acid (AA) transport is regulated by dietary substrate levels, the authors measured uptake of seven radioactively-labelled AAs and glucose across the jejunal brush-border membrane of mice kept on one of three isocaloric rations differing in nitrogen content. In the high-protein ration, uptake increased by 77-81% for the nonessential, less toxic AAs, proline, and aspartate but only by 32-61% for the more toxic essential AAs tested. In the nitrogen-deficient ration, uptake decreased for the nonessential aspartate and proline but stayed constant or increased for essential AAs and for the nonessential alanine. These patterns imply independent regulation of the intestine's various AA transporters. With decreasing dietary AA (or protein), the imino acid and acidic AA private transporters are repressed, while activities of the basic AA transporter and the neutral AA public transporter decrease to an asymptote or else go through a minimum. These regulatory patterns can be understood as a compromise among conflicting constraints imposed by protein's multiple roles as a source of calories, nitrogen, and essential AAs and by the toxicity of essential AAs at high concentrations.

  17. Sequence selective recognition of double-stranded RNA using triple helix-forming peptide nucleic acids.

    PubMed

    Zengeya, Thomas; Gupta, Pankaj; Rozners, Eriks

    2014-01-01

    Noncoding RNAs are attractive targets for molecular recognition because of the central role they play in gene expression. Since most noncoding RNAs are in a double-helical conformation, recognition of such structures is a formidable problem. Herein, we describe a method for sequence-selective recognition of biologically relevant double-helical RNA (illustrated on ribosomal A-site RNA) using peptide nucleic acids (PNA) that form a triple helix in the major grove of RNA under physiologically relevant conditions. Protocols for PNA preparation and binding studies using isothermal titration calorimetry are described in detail.

  18. 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.

  19. ADS genes for reducing saturated fatty acid levels in seed oils

    DOEpatents

    Heilmann, Ingo H.; Shanklin, John

    2010-02-02

    The present invention relates to enzymes involved in lipid metabolism. In particular, the present invention provides coding sequences for Arabidopsis Desaturases (ADS), the encoded ADS polypeptides, and methods for using the sequences and encoded polypeptides, where such methods include decreasing and increasing saturated fatty acid content in plant seed oils.

  20. ADS genes for reducing saturated fatty acid levels in seed oils

    DOEpatents

    Heilmann, Ingo H; Shanklin, John

    2014-03-18

    The present invention relates to enzymes involved in lipid metabolism. In particular, the present invention provides coding sequences for Arabidopsis Desaturases (ADS), the encoded ADS polypeptides, and methods for using the sequences and encoded polypeptides, where such methods include decreasing and increasing saturated fatty acid content in plant seed oils.

  1. Fluorescence energy transfer as a probe for nucleic acid structures and sequences.

    PubMed Central

    Mergny, J L; Boutorine, A S; Garestier, T; Belloc, F; Rougée, M; Bulychev, N V; Koshkin, A A; Bourson, J; Lebedev, A V; Valeur, B

    1994-01-01

    The primary or secondary structure of single-stranded nucleic acids has been investigated with fluorescent oligonucleotides, i.e., oligonucleotides covalently linked to a fluorescent dye. Five different chromophores were used: 2-methoxy-6-chloro-9-amino-acridine, coumarin 500, fluorescein, rhodamine and ethidium. The chemical synthesis of derivatized oligonucleotides is described. Hybridization of two fluorescent oligonucleotides to adjacent nucleic acid sequences led to fluorescence excitation energy transfer between the donor and the acceptor dyes. This phenomenon was used to probe primary and secondary structures of DNA fragments and the orientation of oligodeoxynucleotides synthesized with the alpha-anomers of nucleoside units. Fluorescence energy transfer can be used to reveal the formation of hairpin structures and the translocation of genes between two chromosomes. PMID:8152922

  2. Amino acid sequence of two neurotoxins from the venom of the Egyptian black snake (Walterinnesia aegyptia).

    PubMed

    Samejima, Y; Aoki-Tomomatsu, Y; Yanagisawa, M; Mebs, D

    1997-02-01

    The venom of the Egyptian black snake Walterinnesia aegyptia contains at least three toxins, which act postsynaptically to block the neuromuscular transmission of isolated rat phrenic nerve-diaphragm and chicken biventer cervicis muscle. The complete amino acid sequence of the two toxins, W-III and W-IV, consisting of 62 amino acid residues, was elucidated by Edman degradation of fragments obtained after Staphylococcus aureus protease and prolylpeptidase digestion. Although the toxins exhibit close structural homology to other short-chain postsynaptic neurotoxins from Elapidae venoms, toxin IV is unique by having a free SH-group (cysteine) at position 16. In position 35 of W-III, which is located at the tip of the central loop, threonine is replaced by lysine, which may alter the interaction of the toxin with the acetylcholine receptor, since the toxin is seven times less lethal than toxin W-IV.

  3. Exceptionally high heterologous protein levels in transgenic dicotyledonous seeds using Phaseolus vulgaris regulatory sequences.

    PubMed

    De Jaeger, Geert; Angenon, Geert; Depicker, Ann

    2003-01-01

    Seeds are concentrated sources of protein and thus may be ideal 'bioreactors' for the production of heterologous proteins. For this application, strong seed-specific expression signals are required. A set of expression cassettes were designed using 5' and 3' regulatory sequences of the seed storage protein gene arcelin 5-I (arc5-I) from Phaseolus vulgaris, and evaluated for the production of heterologous proteins in dicotyledonous plant species. A murine single-chain variable fragment (scFv) was chosen as model protein because of the current industrial interest to produce antibodies and derived fragments in crops. Because the highest scFv accumulation in seed had previously been achieved in the endoplasmic reticulum (ER), the scFv-encoding sequence was provided with signal sequences for accumulation in the ER. Transgenic Arabidopsis seed stocks, expressing the scFv under control of the 35S promoter, contained scFv accumulation levels in the range of 1% of total soluble protein (TSP). However, the seed storage promoter constructs boosted the scFv to exceptionally high levels. Maximum scFv levels were obtained in homozygous seed stocks, being 12.5% of TSP under control of the arc5-I regulatory sequences and even up to 36.5% of TSP upon replacing the arc5-I promoter by the beta-phaseolin promoter of Phaseolus vulgaris. Even at such very high levels, the scFv proteins retain their full antigen-binding activity. Moreover, the presence of very high scFv levels has only minory effects on seed germination and no effect on seed production. These results demonstrate that the expression levels of arcelin 5-I and beta-phaseolin seed storage protein genes can be transferred to heterologous proteins, giving exceptionally high levels of heterologous proteins, which can be of great value for the molecular farming industry by raising production yield and lowering bio-mass production and purification costs. Finally, the feasibility of heterologous protein production using the

  4. Complete genome sequence of Bacillus amyloliquefaciens LL3, which exhibits glutamic acid-independent production of poly-γ-glutamic acid.

    PubMed

    Geng, Weitao; Cao, Mingfeng; Song, Cunjiang; Xie, Hui; Liu, Li; Yang, Chao; Feng, Jun; Zhang, Wei; Jin, Yinghong; Du, Yang; Wang, Shufang

    2011-07-01

    Bacillus amyloliquefaciens is one of most prevalent Gram-positive aerobic spore-forming bacteria with the ability to synthesize polysaccharides and polypeptides. Here, we report the complete genome sequence of B. amyloliquefaciens LL3, which was isolated from fermented food and presents the glutamic acid-independent production of poly-γ-glutamic acid.

  5. 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

  6. Interaction of aminooxyacetic acid and ethacrynic acid with intense sound at the level of the cochlea.

    PubMed

    Kisiel, D L; Bobbin, R P

    1982-02-01

    Results of previous investigation of the interaction of intense sound and drugs have, in general, failed to show a protective effect mediated by pre-administration with a drug having transient ototoxic effects. The present investigation was designed to further evaluate a protective effect found previously at the anatomical level and explained with an electrochemical theory of noise damage. The alternating current (a.c) potential and compound eighth nerve action potential (CAP) amplitude were monitored in aminooxyacetic acid (AOAA)- or ethacrynic acid (EA)-treated guinea pigs exposed to either moderate or high levels of intense sound and compared to changes observed in the same potentials in animals exposed to the intense sounds alone. Results showed protective effects only in the moderate--intense sound-exposure groups, with changes in sensitivity and voltage on the linear part of the input--output curve of the a.c cochlear potential found to be the only conditions where differences occurred. These results were difficult to interpret in terms of a protective effect and point to the need for obtaining additional data before an electrochemical mechanism is shown to play a role in the effect of intense sound on the cochlea.

  7. 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

  8. In the TTF-1 homeodomain the contribution of several amino acids to DNA recognition depends on the bound sequence.

    PubMed Central

    Fabbro, D; Tell, G; Leonardi, A; Pellizzari, L; Pucillo, C; Lonigro, R; Formisano, S; Damante, G

    1996-01-01

    The thyroid transcription factor-1 homeodomain (TTF-1HD) shows a peculiar DNA binding specificity, preferentially recognizing sequences containing the 5'-CAAG-3' core motif. Most other homeodomains instead recognize sites containing the 5'-TAAT-3' core motif. Here, we show that TTF-1HD efficiently recognizes another sequence, called D1, devoid of the 5'-CAAG-3' core motif. Different experimental approaches indicate that TTF-1HD contacts the D1 sequence in a manner which is different to that used to interact with sequences containing the 5'-CAAG-3' core motif. The binding activities that mutants of TTF-1HD display with the D1 sequence or with the sequence containing the 5'-CAAG-3' core motif indicate that the role of several DNA-contacting amino acids is different. In particular, during recognition of the D1 sequence, backbone-interacting amino acids not relevant in binding to sequences containing the 5'-CAAG-3' core motif play an important role. In the TTF-1HD, therefore, the contribution of several amino acids to DNA recognition depends on the bound sequence. These data indicate that although a common bonding network exists in all of the HD/DNA complexes, peculiarities important for DNA recognition may occur in single cases. PMID:8811078

  9. Sequence-Level Analysis of the Major European Huntington Disease Haplotype

    PubMed Central

    Lee, Jong-Min; Kim, Kyung-Hee; Shin, Aram; Chao, Michael J.; Abu Elneel, Kawther; Gillis, Tammy; Mysore, Jayalakshmi Srinidhi; Kaye, Julia A.; Zahed, Hengameh; Kratter, Ian H.; Daub, Aaron C.; Finkbeiner, Steven; Li, Hong; Roach, Jared C.; Goodman, Nathan; Hood, Leroy; Myers, Richard H.; MacDonald, Marcy E.; Gusella, James F.

    2015-01-01

    Huntington disease (HD) reflects the dominant consequences of a CAG-repeat expansion in HTT. Analysis of common SNP-based haplotypes has revealed that most European HD subjects have distinguishable HTT haplotypes on their normal and disease chromosomes and that ∼50% of the latter share the same major HD haplotype. We reasoned that sequence-level investigation of this founder haplotype could provide significant insights into the history of HD and valuable information for gene-targeting approaches. Consequently, we performed whole-genome sequencing of HD and control subjects from four independent families in whom the major European HD haplotype segregates with the disease. Analysis of the full-sequence-based HTT haplotype indicated that these four families share a common ancestor sufficiently distant to have permitted the accumulation of family-specific variants. Confirmation of new CAG-expansion mutations on this haplotype suggests that unlike most founders of human disease, the common ancestor of HD-affected families with the major haplotype most likely did not have HD. Further, availability of the full sequence data validated the use of SNP imputation to predict the optimal variants for capturing heterozygosity in personalized allele-specific gene-silencing approaches. As few as ten SNPs are capable of revealing heterozygosity in more than 97% of European HD subjects. Extension of allele-specific silencing strategies to the few remaining homozygous individuals is likely to be achievable through additional known SNPs and discovery of private variants by complete sequencing of HTT. These data suggest that the current development of gene-based targeting for HD could be extended to personalized allele-specific approaches in essentially all HD individuals of European ancestry. PMID:26320893

  10. Chicoric Acid Levels in Basil (Ocimum basilicum L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recently, we reported the presence of chicoric acid in basil leaves (confirmed by co-chromatography with purchased standard). Chicoric acid being the chief phenolic of the Echinacea purpurea plant which is popularly consumed as a dietary supplement. For this study, basil products commonly purchased ...

  11. Circulating Levels of Uric Acid and Risk for Metabolic Syndrome.

    PubMed

    Rubio-Guerra, Alberto F; Morales-López, Herlinda; Garro-Almendaro, Ana K; Vargas-Ayala, German; Durán-Salgado, Montserrat B; Huerta-Ramírez, Saul; Lozano-Nuevo, Jose J

    2017-01-01

    Hyperuricemia leads to insulin resistance, whereas insulin resistance decreases renal excretion of uric acid, both mechanisms link elevated serum uric acid with metabolic syndrome. The aim of this study is to evaluate the probability for the development of metabolic syndrome in low-income young adults with hyperuricaemia.

  12. Molecular cloning, encoding sequence, and expression of vaccinia virus nucleic acid-dependent nucleoside triphosphatase gene.

    PubMed Central

    Rodriguez, J F; Kahn, J S; Esteban, M

    1986-01-01

    A rabbit poxvirus genomic library contained within the expression vector lambda gt11 was screened with polyclonal antiserum prepared against vaccinia virus nucleic acid-dependent nucleoside triphosphatase (NTPase)-I enzyme. Five positive phage clones containing from 0.72- to 2.5-kilobase-pair (kbp) inserts expressed a beta-galactosidase fusion protein that was reactive by immunoblotting with the NTPase-I antibody. Hybridization analysis allowed the location of this gene within the vaccinia HindIIID restriction fragment. From the known nucleotide sequence of the 16-kbp vaccinia HindIIID fragment, we identified a region that contains a 1896-base open reading frame coding for a 631-amino acid protein. Analysis of the complete sequence revealed a highly basic protein, with hydrophilic COOH and NH2 termini, various hydrophobic domains, and no significant homology to other known proteins. Translational studies demonstrate that NTPase-I belongs to a late class of viral genes. This protein is highly conserved among Orthopoxviruses. Images PMID:3025846

  13. 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.

  14. Complete amino acid sequence of a Lolium perenne (perennial rye grass) pollen allergen, Lol p II.

    PubMed

    Ansari, A A; Shenbagamurthi, P; Marsh, D G

    1989-07-05

    The complete amino acid sequence of a Lolium perenne (rye grass) pollen allergen, Lol p II was determined by automated Edman degradation of the protein and selected fragments. Cleavage of the protein by enzymatic and chemical techniques established an unambiguous sequence for the protein. Lol p II contains 97 amino acid residues, with a calculated molecular weight of 10,882. The protein lacks cysteine and glutamine and shows no evidence of glycosylation. Theoretical predictions by Fraga's (Fraga, S. (1982) Can. J. Chem. 60, 2606-2610) and Hopp and Woods' (Hopp, T. P., and Woods, K. R. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 3824-3828) methods indicate the presence of four hydrophilic regions, which may contribute to sequential or parts of conformational B-cell epitopes. Analysis of amphipathic regions by Berzofsky's method indicates the presence of a highly amphipathic region, which may contain, or contribute to, an Ia/T-cell epitope. This latter segment of Lol p II was found to be highly homologous with an antibody-binding segment of the major rye allergen Lol p I and may explain why immune responsiveness to both the allergens is associated with HLA-DR3.

  15. The Sequence-Specific Cellular Uptake of Spherical Nucleic Acid Nanoparticle Conjugates

    PubMed Central

    Narayan, Suguna P.; Choi, Chung Hang J.; Hao, Liangliang; Calabrese, Colin M.; Auyeung, Evelyn; Zhang, Chuan; Goor, Olga J.G.M.

    2015-01-01

    We investigated the sequence-dependent cellular uptake of spherical nucleic acid nanoparticle conjugates (SNAs). This process occurs by interaction with class A scavenger receptors (SR-A) and caveolae-mediated endocytosis. It is known that linear poly(guanine) (poly G) is a natural ligand for SR-A, and it has been proposed that interaction of poly G with SR-A is dependent on the formation of G-quadruplexes. Since G-rich oligonucleotides are known to interact strongly with SR-A, we hypothesized that SNAs with higher G contents would be able to enter cells in larger amounts than SNAs composed of other nucleotides, and as such we measured cellular internalization of SNAs as a function of constituent oligonucleotide sequence. Indeed, SNAs with enriched G content show the highest cellular uptake. Using this hypothesis, we chemically conjugated a small molecule (camptothecin) with SNAs to create drug-SNA conjugates and observed that poly G SNAs deliver the most camptothecin to cells and have the highest cytotoxicity in cancer cells. Our data elucidate important design considerations for enhancing the intracellular delivery of spherical nucleic acids. PMID:26097111

  16. Partial amino acid sequences around sulfhydryl groups of soybean beta-amylase.

    PubMed

    Nomura, K; Mikami, B; Morita, Y

    1987-08-01

    Sulfhydryl (SH) groups of soybean beta-amylase were modified with 5-(iodoaceto-amidoethyl)aminonaphthalene-1-sulfonate (IAEDANS) and the SH-containing peptides exhibiting fluorescence were purified after chymotryptic digestion of the modified enzyme. The sequence analysis of the peptides derived from the modification of all SH groups in the denatured enzyme revealed the existence of six SH groups, in contrast to five reported previously. One of them was found to have extremely low reactivity toward SH-reagents without reduction. In the native state, IAEDANS reacted with 2 mol of SH groups per mol of the enzyme (SH1 and SH2) accompanied with inactivation of the enzyme owing to the modification of SH2 located near the active site of this enzyme. The selective modification of SH2 with IAEDANS was attained after the blocking of SH1 with 5,5'-dithiobis-(2-nitrobenzoic acid). The amino acid sequences of the peptides containing SH1 and SH2 were determined to be Cys-Ala-Asn-Pro-Gln and His-Gln-Cys-Gly-Gly-Asn-Val-Gly-Asp-Ile-Val-Asn-Ile-Pro-Ile-Pro-Gln-Trp, respectively.

  17. Genome Sequence of Lactobacillus rhamnosus Strain CASL, an Efficient l-Lactic Acid Producer from Cheap Substrate Cassava

    PubMed Central

    Yu, Bo; Su, Fei; Wang, Limin; Zhao, Bo; Qin, Jiayang; Ma, Cuiqing; Xu, Ping; Ma, Yanhe

    2011-01-01

    Lactobacillus rhamnosus is a type of probiotic bacteria with industrial potential for l-lactic acid production. We announce the draft genome sequence of L. rhamnosus CASL (2,855,156 bp with a G+C content of 46.6%), which is an efficient producer of l-lactic acid from cheap, nonfood substrate cassava with a high production titer. PMID:22123765

  18. Convergent Evolution of Hemoglobin Function in High-Altitude Andean Waterfowl Involves Limited Parallelism at the Molecular Sequence Level

    PubMed Central

    Natarajan, Chandrasekhar; Projecto-Garcia, Joana; Moriyama, Hideaki; Weber, Roy E.; Muñoz-Fuentes, Violeta; Green, Andy J.; Kopuchian, Cecilia; Tubaro, Pablo L.; Alza, Luis; Bulgarella, Mariana; Smith, Matthew M.; Wilson, Robert E.; Fago, Angela; McCracken, Kevin G.; Storz, Jay F.

    2015-01-01

    A fundamental question in evolutionary genetics concerns the extent to which adaptive phenotypic convergence is attributable to convergent or parallel changes at the molecular sequence level. Here we report a comparative analysis of hemoglobin (Hb) function in eight phylogenetically replicated pairs of high- and low-altitude waterfowl taxa to test for convergence in the oxygenation properties of Hb, and to assess the extent to which convergence in biochemical phenotype is attributable to repeated amino acid replacements. Functional experiments on native Hb variants and protein engineering experiments based on site-directed mutagenesis revealed the phenotypic effects of specific amino acid replacements that were responsible for convergent increases in Hb-O2 affinity in multiple high-altitude taxa. In six of the eight taxon pairs, high-altitude taxa evolved derived increases in Hb-O2 affinity that were caused by a combination of unique replacements, parallel replacements (involving identical-by-state variants with independent mutational origins in different lineages), and collateral replacements (involving shared, identical-by-descent variants derived via introgressive hybridization). In genome scans of nucleotide differentiation involving high- and low-altitude populations of three separate species, function-altering amino acid polymorphisms in the globin genes emerged as highly significant outliers, providing independent evidence for adaptive divergence in Hb function. The experimental results demonstrate that convergent changes in protein function can occur through multiple historical paths, and can involve multiple possible mutations. Most cases of convergence in Hb function did not involve parallel substitutions and most parallel substitutions did not affect Hb-O2 affinity, indicating that the repeatability of phenotypic evolution does not require parallelism at the molecular level. PMID:26637114

  19. Convergent Evolution of Hemoglobin Function in High-Altitude Andean Waterfowl Involves Limited Parallelism at the Molecular Sequence Level.

    PubMed

    Natarajan, Chandrasekhar; Projecto-Garcia, Joana; Moriyama, Hideaki; Weber, Roy E; Muñoz-Fuentes, Violeta; Green, Andy J; Kopuchian, Cecilia; Tubaro, Pablo L; Alza, Luis; Bulgarella, Mariana; Smith, Matthew M; Wilson, Robert E; Fago, Angela; McCracken, Kevin G; Storz, Jay F

    2015-12-01

    A fundamental question in evolutionary genetics concerns the extent to which adaptive phenotypic convergence is attributable to convergent or parallel changes at the molecular sequence level. Here we report a comparative analysis of hemoglobin (Hb) function in eight phylogenetically replicated pairs of high- and low-altitude waterfowl taxa to test for convergence in the oxygenation properties of Hb, and to assess the extent to which convergence in biochemical phenotype is attributable to repeated amino acid replacements. Functional experiments on native Hb variants and protein engineering experiments based on site-directed mutagenesis revealed the phenotypic effects of specific amino acid replacements that were responsible for convergent increases in Hb-O2 affinity in multiple high-altitude taxa. In six of the eight taxon pairs, high-altitude taxa evolved derived increases in Hb-O2 affinity that were caused by a combination of unique replacements, parallel replacements (involving identical-by-state variants with independent mutational origins in different lineages), and collateral replacements (involving shared, identical-by-descent variants derived via introgressive hybridization). In genome scans of nucleotide differentiation involving high- and low-altitude populations of three separate species, function-altering amino acid polymorphisms in the globin genes emerged as highly significant outliers, providing independent evidence for adaptive divergence in Hb function. The experimental results demonstrate that convergent changes in protein function can occur through multiple historical paths, and can involve multiple possible mutations. Most cases of convergence in Hb function did not involve parallel substitutions and most parallel substitutions did not affect Hb-O2 affinity, indicating that the repeatability of phenotypic evolution does not require parallelism at the molecular level.

  20. Amino acid sequence of versutoxin, a lethal neurotoxin from the venom of the funnel-web spider Atrax versutus.

    PubMed

    Brown, M R; Sheumack, D D; Tyler, M I; Howden, M E

    1988-03-01

    The complete amino acid sequence of versutoxin, a lethal neurotoxic polypeptide isolated from the venom of male and female funnel-web spiders of the species Atrax versutus, was determined. Sequencing was performed in a gas-phase protein sequencer by automated Edman degradation of the S-carboxymethylated toxin and fragments of it produced by reaction with CNBr. Versutoxin consisted of a single chain of 42 amino acid residues. It was found to have a high proportion of basic residues and of cystine. The primary structure showed marked homology with that of robustoxin, a novel neurotoxin recently isolated from the venom of another funnel-web-spider species, Atrax robustus.

  1. Amino acid sequence of versutoxin, a lethal neurotoxin from the venom of the funnel-web spider Atrax versutus.

    PubMed Central

    Brown, M R; Sheumack, D D; Tyler, M I; Howden, M E

    1988-01-01

    The complete amino acid sequence of versutoxin, a lethal neurotoxic polypeptide isolated from the venom of male and female funnel-web spiders of the species Atrax versutus, was determined. Sequencing was performed in a gas-phase protein sequencer by automated Edman degradation of the S-carboxymethylated toxin and fragments of it produced by reaction with CNBr. Versutoxin consisted of a single chain of 42 amino acid residues. It was found to have a high proportion of basic residues and of cystine. The primary structure showed marked homology with that of robustoxin, a novel neurotoxin recently isolated from the venom of another funnel-web-spider species, Atrax robustus. PMID:3355530

  2. Amino acid sequence of neurotoxin III of the scorpion Androctonus austrialis Hector.

    PubMed

    Kopeyan, C; Martinez, G; Rochat, H

    1979-03-01

    The amino acid sequence of neurotoxin III, purified from the venom of the North African scorpion Androctonus australis Hector, has been determined by Edman degradation using a liquid-phase sequencer. Carboxypeptidase A hydrolyses confirmed not only the sequence of the five last residues but also the presence of a free alpha-carboxylic group at the C-terminus. Edman degradation was conducted on one hand with the Quadrol [N,N,N',N'-tetrakis(2-hydroxypropyl)ethylene diamine] program and S-alkylated protein before or after coupling with sulfophenylisothiocynate (the first 34 residues were thus identified), on the other hand on tryptic and chymotryptic peptides with a dimethylbenzylamine program (residues 1--23 and 31--34 were confirmed, the positions of residues 35-64 were established). Neurotoxin III was found to belong to the same group of scorpion toxins active on mammals as neurotoxin I purified from the same venom (50 homologous positions exist in the two proteins).

  3. Purification, amino acid sequence and characterisation of kangaroo IGF-I.

    PubMed

    Yandell, C A; Francis, G L; Wheldrake, J F; Upton, Z

    1998-01-01

    Insulin-like growth factor-I (IGF-I) and IGF-II have been purified to homogeneity from kangaroo (Macropus fuliginosus) serum, thus this represents the first report of the purification, sequencing and characterisation of marsupial IGFs. N-Terminal protein sequencing reveals that there are six amino acid differences between kangaroo and human IGF-I. Kangaroo IGF-II has been partially sequenced and no differences were found between human and kangaroo IGF-II in the 53 residues identified. Thus the IGFs appear to be remarkably structurally conserved during mammalian radiation. In addition, in vitro characterisation of kangaroo IGF-I demonstrated that the functional properties of human, kangaroo and chicken IGF-I are very similar. In an assay measuring the ability of the proteins to stimulate protein synthesis in rat L6 myoblasts, all IGF-I proteins were found to be equally potent. The ability of all three proteins to compete for binding with radiolabelled human IGF-I to type-1 IGF receptors in L6 myoblasts and in Sminthopsis crassicaudata transformed lung fibroblasts, a marsupial cell line, was comparable. Furthermore, kangaroo and human IGF-I react equally in a human IGF-I RIA using a human reference standard, radiolabelled human IGF-I and a polyclonal antibody raised against recombinant human IGF-I. This study indicates that not only is the primary structure of eutherian and metatherian IGF-I conserved, but also the proteins appear to be functionally similar.

  4. Complete Genome Sequence of the Prototype Lactic Acid Bacterium Lactococcus lactis subsp. cremoris MG1363▿

    PubMed Central

    Wegmann, Udo; O'Connell-Motherway, Mary; Zomer, Aldert; Buist, Girbe; Shearman, Claire; Canchaya, Carlos; Ventura, Marco; Goesmann, Alexander; Gasson, Michael J.; Kuipers, Oscar P.; van Sinderen, Douwe; Kok, Jan

    2007-01-01

    Lactococcus lactis is of great importance for the nutrition of hundreds of millions of people worldwide. This paper describes the genome sequence of Lactococcus lactis subsp. cremoris MG1363, the lactococcal strain most intensively studied throughout the world. The 2,529,478-bp genome contains 81 pseudogenes and encodes 2,436 proteins. Of the 530 unique proteins, 47 belong to the COG (clusters of orthologous groups) functional category “carbohydrate metabolism and transport,” by far the largest category of novel proteins in comparison with L. lactis subsp. lactis IL1403. Nearly one-fifth of the 71 insertion elements are concentrated in a specific 56-kb region. This integration hot-spot region carries genes that are typically associated with lactococcal plasmids and a repeat sequence specifically found on plasmids and in the “lateral gene transfer hot spot” in the genome of Streptococcus thermophilus. Although the parent of L. lactis MG1363 was used to demonstrate lysogeny in Lactococcus, L. lactis MG1363 carries four remnant/satellite phages and two apparently complete prophages. The availability of the L. lactis MG1363 genome sequence will reinforce its status as the prototype among lactic acid bacteria through facilitation of further applied and fundamental research. PMID:17307855

  5. N-terminal amino acid sequences of D-serine deaminases of wild-type and operator-constitutive strains of Escherichia coli K-12.

    PubMed Central

    Heincz, M C; McFall, E

    1975-01-01

    The N-terminal amino acid sequences of the D-serine deaminases from strains of Escherichia coli K-12 that harbor wild-type and high-level constitutive catabolite-insensitive operator-initiator regions are identical: Met-Ser-GluNH2-Ser-Gly-Arg-His-Cys. This result indicates that the operator-initiator region is probably distinct from the D-serine deaminase structural gene. Images PMID:1099073

  6. The ABRF Edman Sequencing Research Group 2008 Study: Investigation into Homopolymeric Amino Acid N-Terminal Sequence Tags and Their Effects on Automated Edman Degradation

    PubMed Central

    Thoma, R. S.; Smith, J. S.; Sandoval, W.; Leone, J. W.; Hunziker, P.; Hampton, B.; Linse, K. D.; Denslow, N. D.

    2009-01-01

    The Edman Sequence Research Group (ESRG) of the Association of Biomolecular Resource designs and executes interlaboratory studies investigating the use of automated Edman degradation for protein and peptide analysis. In 2008, the ESRG enlisted the help of core sequencing facilities to investigate the effects of a repeating amino acid tag at the N-terminus of a protein. Commonly, to facilitate protein purification, an affinity tag containing a polyhistidine sequence is conjugated to the N-terminus of the protein. After expression, polyhistidine-tagged protein is readily purified via chelation with an immobilized metal affinity resin. The addition of the polyhistidine tag presents unique challenges for the determination of protein identity using Edman degradation chemistry. Participating laboratories were asked to sequence one protein engineered in three configurations: with an N-terminal polyhistidine tag; with an N-terminal polyalanine tag; or with no tag. Study participants were asked to return a data file containing the uncorrected amino acid picomole yields for the first 17 cycles. Initial and repetitive yield (R.Y.) information and the amount of lag were evaluated. Information about instrumentation and sample treatment was also collected as part of the study. For this study, the majority of participating laboratories successfully called the amino acid sequence for 17 cycles for all three test proteins. In general, laboratories found it more difficult to call the sequence containing the polyhistidine tag. Lag was observed earlier and more consistently with the polyhistidine-tagged protein than the polyalanine-tagged protein. Histidine yields were significantly less than the alanine yields in the tag portion of each analysis. The polyhistidine and polyalanine protein-R.Y. calculations were found to be equivalent. These calculations showed that the nontagged portion from each protein was equivalent. The terminal histidines from the tagged portion of the protein

  7. The amino acid sequence around the active-site cysteine and histidine residues, and the buried cysteine residue in ficin.

    PubMed

    Husain, S S; Lowe, G

    1970-04-01

    Ficin that had been prepared from the latex of Ficus glabrata by salt fractionation and chromatography on carboxymethylcellulose was completely and irreversibly inhibited with 1,3-dibromo[2-(14)C]acetone and then treated with N-(4-dimethylamino-3,5-dinitrophenyl)maleimide in 6m-guanidinium chloride. After reduction and carboxymethylation of the labelled protein, it was digested with trypsin and alpha-chymotrypsin. Two radioactive peptides and two coloured peptides were isolated chromatographically and their sequences determined. The radioactive peptides revealed the amino acid sequences around the active-site cysteine and histidine residues and showed a high degree of homology with the omino acid sequence around the active-site cysteine and histidine residues in papain. The coloured peptides allowed the amino acid sequence around the buried cysteine residue in ficin to be determined.

  8. The `heavy' subunit of the photosynthetic reaction centre from Rhodopseudomonas viridis: isolation of the gene, nucleotide and amino acid sequence

    PubMed Central

    Michel, H.; Weyer, K. A.; Gruenberg, H.; Lottspeich, F.

    1985-01-01

    The gene coding for the `heavy' subunit of the photosynthetic reaction centre from Rhodopseudomonas viridis was isolated in an expression vector. Expression of the heavy subunit in Escherichia coli was detected with antibodies raised against crystalline reaction centres. The entire subunit, and not a fusion protein, was expressed in E. coli. The protein coding region of the gene was sequenced and the amino acid sequence derived. Part of the amino acid sequence was confirmed by chemical sequence analysis of the protein. The heavy subunit consists of 258 amino acids and its mol. wt. is 28 345. It possesses one membrane-spanning α-helical segment, as was revealed by the concomitant X-ray structure analysis. ImagesFig. 1.Fig. 2. PMID:16453623

  9. Purification, amino acid sequence and immunological characterization of Ole e 6, a cysteine-enriched allergen from olive tree pollen.

    PubMed

    Batanero, E; Ledesma, A; Villalba, M; Rodríguez, R

    1997-06-30

    The Ole e 6 allergen from olive tree pollen has been isolated by combining gel permeation and reverse-phase chromatographies. It is a single and highly acidic (pI 4.2) polypeptide chain protein. Its NH2-terminal amino acid sequence has been determined by Edman degradation. Total RNA from the olive tree pollen was isolated, and a specific cDNA was amplified by the polymerase chain reaction using a degenerate oligonucleotide primer designed according to the NH2-terminal sequence of the protein. The nucleotide sequencing of the cDNA rendered an open reading frame encoding a 50 amino acid polypeptide chain, in which two sets of the sequential motif Cys-X3-Cys-X3-Cys are present. No sequence similarity has been found between this protein and other previously described polypeptides.

  10. 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

  11. The evolution of proteins from random amino acid sequences: II. Evidence from the statistical distributions of the lengths of modern protein sequences.

    PubMed

    White, S H

    1994-04-01

    This paper continues an examination of the hypothesis that modern proteins evolved from random heteropeptide sequences. In support of the hypothesis, White and Jacobs (1993, J Mol Evol 36:79-95) have shown that any sequence chosen randomly from a large collection of nonhomologous proteins has a 90% or better chance of having a lengthwise distribution of amino acids that is indistinguishable from the random expectation regardless of amino acid type. The goal of the present study was to investigate the possibility that the random-origin hypothesis could explain the lengths of modern protein sequences without invoking specific mechanisms such as gene duplication or exon splicing. The sets of sequences examined were taken from the 1989 PIR database and consisted of 1,792 "super-family" proteins selected to have little sequence identity, 623 E. coli sequences, and 398 human sequences. The length distributions of the proteins could be described with high significance by either of two closely related probability density functions: The gamma distribution with parameter 2 or the distribution for the sum of two exponential random independent variables. A simple theory for the distributions was developed which assumes that (1) protoprotein sequences had exponentially distributed random independent lengths, (2) the length dependence of protein stability determined which of these protoproteins could fold into compact primitive proteins and thereby attain the potential for biochemical activity, (3) the useful protein sequences were preserved by the primitive genome, and (4) the resulting distribution of sequence lengths is reflected by modern proteins. The theory successfully predicts the two observed distributions which can be distinguished by the functional form of the dependence of protein stability on length. The theory leads to three interesting conclusions. First, it predicts that a tetra-nucleotide was the signal for primitive translation termination. This prediction is

  12. Arthropod Phylogenetics in Light of Three Novel Millipede (Myriapoda: Diplopoda) Mitochondrial Genomes with Comments on the Appropriateness of Mitochondrial Genome Sequence Data for Inferring Deep Level Relationships

    PubMed Central

    Brewer, Michael S.; Swafford, Lynn; Spruill, Chad L.; Bond, Jason E.

    2013-01-01

    Background Arthropods are the most diverse group of eukaryotic organisms, but their phylogenetic relationships are poorly understood. Herein, we describe three mitochondrial genomes representing orders of millipedes for which complete genomes had not been characterized. Newly sequenced genomes are combined with existing data to characterize the protein coding regions of myriapods and to attempt to reconstruct the evolutionary relationships within the Myriapoda and Arthropoda. Results The newly sequenced genomes are similar to previously characterized millipede sequences in terms of synteny and length. Unique translocations occurred within the newly sequenced taxa, including one half of the Appalachioria falcifera genome, which is inverted with respect to other millipede genomes. Across myriapods, amino acid conservation levels are highly dependent on the gene region. Additionally, individual loci varied in the level of amino acid conservation. Overall, most gene regions showed low levels of conservation at many sites. Attempts to reconstruct the evolutionary relationships suffered from questionable relationships and low support values. Analyses of phylogenetic informativeness show the lack of signal deep in the trees (i.e., genes evolve too quickly). As a result, the myriapod tree resembles previously published results but lacks convincing support, and, within the arthropod tree, well established groups were recovered as polyphyletic. Conclusions The novel genome sequences described herein provide useful genomic information concerning millipede groups that had not been investigated. Taken together with existing sequences, the variety of compositions and evolution of myriapod mitochondrial genomes are shown to be more complex than previously thought. Unfortunately, the use of mitochondrial protein-coding regions in deep arthropod phylogenetics appears problematic, a result consistent with previously published studies. Lack of phylogenetic signal renders the

  13. Fragmentation Characteristics of Deprotonated N-linked Glycopeptides: Influences of Amino Acid Composition and Sequence

    NASA Astrophysics Data System (ADS)

    Nishikaze, Takashi; Kawabata, Shin-ichirou; Tanaka, Koichi

    2014-06-01

    Glycopeptide structural analysis using tandem mass spectrometry is becoming a common approach for elucidating site-specific N-glycosylation. The analysis is generally performed in positive-ion mode. Therefore, fragmentation of protonated glycopeptides has been extensively investigated; however, few studies are available on deprotonated glycopeptides, despite the usefulness of negative-ion mode analysis in detecting glycopeptide signals. Here, large sets of glycopeptides derived from well-characterized glycoproteins were investigated to understand the fragmentation behavior of deprotonated N-linked glycopeptides under low-energy collision-induced dissociation (CID) conditions. The fragment ion species were found to be significantly variable depending on their amino acid sequence and could be classified into three types: (i) glycan fragment ions, (ii) glycan-lost fragment ions and their secondary cleavage products, and (iii) fragment ions with intact glycan moiety. The CID spectra of glycopeptides having a short peptide sequence were dominated by type (i) glycan fragments (e.g., 2,4AR, 2,4AR-1, D, and E ions). These fragments define detailed structural features of the glycan moiety such as branching. For glycopeptides with medium or long peptide sequences, the major fragments were type (ii) ions (e.g., [peptide + 0,2X0-H]- and [peptide-NH3-H]-). The appearance of type (iii) ions strongly depended on the peptide sequence, and especially on the presence of Asp, Asn, and Glu. When a glycosylated Asn is located on the C-terminus, an interesting fragment having an Asn residue with intact glycan moiety, [glycan + Asn-36]-, was abundantly formed. Observed fragments are reasonably explained by a combination of existing fragmentation rules suggested for N-glycans and peptides.

  14. An amino acid sequence motif sufficient for subnuclear localization of an arginine/serine-rich splicing factor.

    PubMed

    Hedley, M L; Amrein, H; Maniatis, T

    1995-12-05

    We have identified an amino acid sequence in the Drosophila Transformer (Tra) protein that is capable of directing a heterologous protein to nuclear speckles, regions of the nucleus previously shown to contain high concentrations of spliceosomal small nuclear RNAs and splicing factors. This sequence contains a nucleoplasmin-like bipartite nuclear localization signal (NLS) and a repeating arginine/serine (RS) dipeptide sequence adjacent to a short stretch of basic amino acids. Sequence comparisons from a number of other splicing factors that colocalize to nuclear speckles reveal the presence of one or more copies of this motif. We propose a two-step subnuclear localization mechanism for splicing factors. The first step is transport across the nuclear envelope via the nucleoplasmin-like NLS, while the second step is association with components in the speckled domain via the RS dipeptide sequence.

  15. 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.

  16. Variation in seed fatty acid composition and sequence divergence in the FAD2 gene coding region between wild and cultivated sesame.

    PubMed

    Chen, Zhenbang; Tonnis, Brandon; Morris, Brad; Wang, Richard B; Zhang, Amy L; Pinnow, David; Wang, Ming Li

    2014-12-03

    Sesame germplasm harbors genetic diversity which can be useful for sesame improvement in breeding programs. Seven accessions with different levels of oleic acid were selected from the entire USDA sesame germplasm collection (1232 accessions) and planted for morphological observation and re-examination of fatty acid composition. The coding region of the FAD2 gene for fatty acid desaturase (FAD) in these accessions was also sequenced. Cultivated sesame accessions flowered and matured earlier than the wild species. The cultivated sesame seeds contained a significantly higher percentage of oleic acid (40.4%) than the seeds of the wild species (26.1%). Nucleotide polymorphisms were identified in the FAD2 gene coding region between wild and cultivated species. Some nucleotide polymorphisms led to amino acid changes, one of which was located in the enzyme active site and may contribute to the altered fatty acid composition. Based on the morphology observation, chemical analysis, and sequence analysis, it was determined that two accessions were misnamed and need to be reclassified. The results obtained from this study are useful for sesame improvement in molecular breeding programs.

  17. Salivary Sialic Acid Levels in Smokeless Tobacco Users

    PubMed Central

    Farhad Mollashahi, Leila; Honarmand, Marieh; Nakhaee, Alireza; Mollashahi, Ghasem

    2016-01-01

    Background Smokeless tobacco chewing is one of the known risk factors for oral cancer. It is consumed widely by residents of southeastern Iran. Objectives In this study, salivary free and total sialic acid, and total protein were compared in paan consumers and non-consumers. Patients and Methods In this cross-sectional study, unstimulated saliva of 94 subjects (44 paan consumers and 50 non-consumers) who were referred to the oral medicine department of the dentistry school of Zahedan were collected. Salivary free and total sialic acid, and total protein concentration were measured by standard biochemical methods, and the obtained data were analyzed using SPSS 20 through the non-parametric Mann-Whitney test. Results The concentration of salivary free sialic acid (23.21 ± 18.98 mg/L) was significantly increased in paan consumers. The concentration of salivary Total sialic acid (TSA) (39.57 ± 26.58 mg/L) and total protein (0.77 ± 0.81 mg/mL) showed increases in paan consumers, however, the results were not statistically significant. Conclusions Salivary free and total sialic acid, and total protein were higher in the paan consumers compared to non-consumers. Due to the carcinogenic effect of smokeless tobacco, measurement of these parameters in saliva may be useful in early detection of oral cancer. PMID:27622172

  18. Exozodiacal Dust Levels for Nearby Main-sequence Stars: A Survey with the Keck Interferometer Nuller

    NASA Astrophysics Data System (ADS)

    Millan-Gabet, R.; Serabyn, E.; Mennesson, B.; Traub, W. A.; Barry, R. K.; Danchi, W. C.; Kuchner, M.; Stark, C. C.; Ragland, S.; Hrynevych, M.; Woillez, J.; Stapelfeldt, K.; Bryden, G.; Colavita, M. M.; Booth, A. J.

    2011-06-01

    The Keck Interferometer Nuller (KIN) was used to survey 25 nearby main-sequence stars in the mid-infrared, in order to assess the prevalence of warm circumstellar (exozodiacal) dust around nearby solar-type stars. The KIN measures circumstellar emission by spatially blocking the star but transmitting the circumstellar flux in a region typically 0.1-4 AU from the star. We find one significant detection (η Crv), two marginal detections (γ Oph and α Aql), and 22 clear non-detections. Using a model of our own solar system's zodiacal cloud, scaled to the luminosity of each target star, we estimate the equivalent number of target zodis needed to match our observations. Our three zodi detections are η Crv (1250 ± 260), γ Oph (200 ± 80), and α Aql (600 ± 200), where the uncertainties are 1σ. The 22 non-detected targets have an ensemble weighted average consistent with zero, with an average individual uncertainty of 160 zodis (1σ). These measurements represent the best limits to date on exozodi levels for a sample of nearby main-sequence stars. A statistical analysis of the population of 23 stars not previously known to contain circumstellar dust (excluding η Crv and γ Oph) suggests that, if the measurement errors are uncorrelated (for which we provide evidence) and if these 23 stars are representative of a single class with respect to the level of exozodi brightness, the mean exozodi level for the class is <150 zodis (3σ upper limit, corresponding to 99% confidence under the additional assumption that the measurement errors are Gaussian). We also demonstrate that this conclusion is largely independent of the shape and mean level of the (unknown) true underlying exozodi distribution.

  19. Alignment of 700 globin sequences: extent of amino acid substitution and its correlation with variation in volume.

    PubMed Central

    Kapp, O. H.; Moens, L.; Vanfleteren, J.; Trotman, C. N.; Suzuki, T.; Vinogradov, S. N.

    1995-01-01

    Seven-hundred globin sequences, including 146 nonvertebrate sequences, were aligned on the basis of conservation of secondary structure and the avoidance of gap penalties. Of the 182 positions needed to accommodate all the globin sequences, only 84 are common to all, including the absolutely conserved PheCD1 and HisF8. The mean number of amino acid substitutions per position ranges from 8 to 13 for all globins and 5 to 9 for internal positions. Although the total sequence volumes have a variation approximately 2-3%, the variation in volume per position ranges from approximately 13% for the internal to approximately 21% for the surface positions. Plausible correlations exist between amino acid substitution and the variation in volume per position for the 84 common and the internal but not the surface positions. The amino acid substitution matrix derived from the 84 common positions was used to evaluate sequence similarity within the globins and between the globins and phycocyanins C and colicins A, via calculation of pairwise similarity scores. The scores for globin-globin comparisons over the 84 common positions overlap the globin-phycocyanin and globin-colicin scores, with the former being intermediate. For the subset of internal positions, overlap is minimal between the three groups of scores. These results imply a continuum of amino acid sequences able to assume the common three-on-three alpha-helical structure and suggest that the determinants of the latter include sites other than those inaccessible to solvent. PMID:8535255

  20. Amino acid substitutions in genetic variants of human serum albumin and in sequences inferred from molecular cloning

    SciTech Connect

    Takahashi, N.; Takahashi, Y.; Blumberg, B.S.; Putnam, F.W.

    1987-07-01

    The structural changes in four genetic variants of human serum albumin were analyzed by tandem high-pressure liquid chromatography (HPLC) of the tryptic peptides, HPLC mapping and isoelectric focusing of the CNBr fragments, and amino acid sequence analysis of the purified peptides. Lysine-372 of normal (common) albumin A was changed to glutamic acid both in albumin Naskapi, a widespread polymorphic variant of North American Indians, and in albumin Mersin found in Eti Turks. The two variants also exhibited anomalous migration in NaDodSO/sub 4//PAGE, which is attributed to a conformational change. The identity of albumins Naskapi and Mersin may have originated through descent from a common mid-Asiatic founder of the two migrating ethnic groups, or it may represent identical but independent mutations of the albumin gene. In albumin Adana, from Eti Turks, the substitution site was not identified but was localized to the region from positions 447 through 548. The substitution of aspartic acid-550 by glycine was found in albumin Mexico-2 from four individuals of the Pima tribe. Although only single-point substitutions have been found in these and in certain other genetic variants of human albumin, five differences exist in the amino acid sequences inferred from cDNA sequences by workers in three other laboratories. However, our results on albumin A and on 14 different genetic variants accord with the amino acid sequence of albumin deduced from the genomic sequence. The apparent amino acid substitutions inferred from comparison of individual cDNA sequences probably reflect artifacts in cloning or in cDNA sequence analysis rather than polymorphism of the coding sections of the albumin gene.

  1. Assessment of Folic Acid Supplementation in Pregnant Women by Estimation of Serum Levels of Tetrahydrofolic Acid, Dihydrofolate Reductase, and Homocysteine.

    PubMed

    Naithani, Manisha; Saxena, Vartika; Mirza, Anissa Atif; Kumari, Ranjeeta; Sharma, Kapil; Bharadwaj, Jyoti

    2016-01-01

    Background. Status of folic acid use in pregnant women of the hilly regions in North India was little known. This study was carried out to assess the folic acid use and estimate folate metabolites in pregnant women of this region. Materials and Methods. This cross-sectional study is comprised of 76 pregnant women, whose folic acid supplementation was assessed by a questionnaire and serum levels of homocysteine, tetrahydrofolic acid (THFA), and dihydrofolate reductase (DHFR) were estimated using Enzyme Linked Immunoassays. Results. The study data revealed awareness of folic acid use during pregnancy was present in 46.1% and 23.7% were taking folic acid supplements. The study depicted that there was no statistically significant difference between serum levels of THFA and DHFR in pregnant women with and without folic acid supplements (p = 0.790). Hyperhomocysteinemia was present in 15.78% of the participants. Conclusion. Less awareness about folic acid supplementation and low use of folic acid by pregnant women were observed in this region. Sufficient dietary ingestion may suffice for the escalated requirements in pregnancy, but since this cannot be ensured, hence folic acid supplementation should be made as an integral part of education and reproductive health programs for its better metabolic use, growth, and development of fetus.

  2. Assessment of Folic Acid Supplementation in Pregnant Women by Estimation of Serum Levels of Tetrahydrofolic Acid, Dihydrofolate Reductase, and Homocysteine

    PubMed Central

    Saxena, Vartika; Mirza, Anissa Atif; Kumari, Ranjeeta; Sharma, Kapil; Bharadwaj, Jyoti

    2016-01-01

    Background. Status of folic acid use in pregnant women of the hilly regions in North India was little known. This study was carried out to assess the folic acid use and estimate folate metabolites in pregnant women of this region. Materials and Methods. This cross-sectional study is comprised of 76 pregnant women, whose folic acid supplementation was assessed by a questionnaire and serum levels of homocysteine, tetrahydrofolic acid (THFA), and dihydrofolate reductase (DHFR) were estimated using Enzyme Linked Immunoassays. Results. The study data revealed awareness of folic acid use during pregnancy was present in 46.1% and 23.7% were taking folic acid supplements. The study depicted that there was no statistically significant difference between serum levels of THFA and DHFR in pregnant women with and without folic acid supplements (p = 0.790). Hyperhomocysteinemia was present in 15.78% of the participants. Conclusion. Less awareness about folic acid supplementation and low use of folic acid by pregnant women were observed in this region. Sufficient dietary ingestion may suffice for the escalated requirements in pregnancy, but since this cannot be ensured, hence folic acid supplementation should be made as an integral part of education and reproductive health programs for its better metabolic use, growth, and development of fetus. PMID:27064332

  3. Real-Time Nucleic Acid Sequence-Based Amplification Assay for Detection of Hepatitis A Virus

    PubMed Central

    Abd El Galil, Khaled H.; El Sokkary, M. A.; Kheira, S. M.; Salazar, Andre M.; Yates, Marylynn V.; Chen, Wilfred; Mulchandani, Ashok

    2005-01-01

    A nucleic acid sequence-based amplification (NASBA) assay in combination with a molecular beacon was developed for the real-time detection and quantification of hepatitis A virus (HAV). A 202-bp, highly conserved 5′ noncoding region of HAV was targeted. The sensitivity of the real-time NASBA assay was tested with 10-fold dilutions of viral RNA, and a detection limit of 1 PFU was obtained. The specificity of the assay was demonstrated by testing with other environmental pathogens and indicator microorganisms, with only HAV positively identified. When combined with immunomagnetic separation, the NASBA assay successfully detected as few as 10 PFU from seeded lake water samples. Due to its isothermal nature, its speed, and its similar sensitivity compared to the real-time RT-PCR assay, this newly reported real-time NASBA method will have broad applications for the rapid detection of HAV in contaminated food or water. PMID:16269748

  4. 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.

  5. Sequence-defined shuttles for targeted nucleic acid and protein delivery.

    PubMed

    Röder, Ruth; Wagner, Ernst

    2014-01-01

    Molecular medicine opens into a space of novel specific therapeutic agents: intracellularly active drugs such as peptides, proteins or nucleic acids, which are not able to cross cell membranes and enter the intracellular space on their own. Through the development of cell-targeted shuttles for specific delivery, this restriction in delivery has the potential to be converted into an advantage. On the one hand, due to the multiple extra- and intracellular barriers, such carrier systems need to be multifunctional. On the other hand, they must be precise and reproducibly manufactured due to pharmaceutical reasons. Here we review the design of precise sequence-defined delivery carriers, including solid-phase synthesized peptides and nonpeptidic oligomers, or nucleotide-based carriers such as aptamers and origami nanoboxes.

  6. Identification of amino acid sequences in the polyomavirus capsid proteins that serve as nuclear localization signals

    NASA Technical Reports Server (NTRS)

    Chang, D.; Haynes, J. I. Jr; Brady, J. N.; Consigli, R. A.; Spooner, B. S. (Principal Investigator)

    1993-01-01

    The molecular mechanism participating in the transport of newly synthesized proteins from the cytoplasm to the nucleus in mammalian cells is poorly understood. Recently, the nuclear localization signal sequences (NLS) of many nuclear proteins have been identified, and most have been found to be composed of a highly basic amino acid stretch. A genetic "subtractive" and a biochemical "additive" approach were used in our studies to identify the NLS's of the polyomavirus structural capsid proteins. An NLS was identified at the N-terminus (Ala1-Pro-Lys-Arg-Lys-Ser-Gly-Val-Ser-Lys-Cys11) of the major capsid protein VP1 and at the C-terminus (Glu307 -Glu-Asp-Gly-Pro-Glu-Lys-Lys-Lys-Arg-Arg-Leu318) of the VP2/VP3 minor capsid proteins.

  7. The amino acid sequence of a carbohydrate-containing fragment of hen ovotransferrin.

    PubMed Central

    Kingston, I B; Williams, J

    1975-01-01

    1. Hen ovotransferrin was treated with CNBr and fractionated by gel filtration. 2. After further treatment by reduction and carboxymethylation a carbohydrate-containing fragment of molecular weight 11990 was obtained (fragment BCd). 3. The amino acid sequence of this fragment was determined. It consists of a single chain of 94 residues. 4. The structure of a tryptic glycopeptide derived from whole ovotransferrin permitted a further eight residues to be assigned at the N-terminus of fragment BCd. 5. Heterogeneity was found at two positions. 6. Further evidence has been deposited as Supplementary Publication SUP 50045 (19 pages) at the British Library (Lending Division), Boston Spa, Wetherby, W. Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1975), 145, 5. PMID:1172663

  8. Phylogenetic analysis of beta-papillomaviruses as inferred from nucleotide and amino acid sequence data.

    PubMed

    Gottschling, Marc; Köhler, Anja; Stockfleth, Eggert; Nindl, Ingo

    2007-01-01

    Human papillomaviruses (HPV) of the beta-group seem to be involved in the pathogenesis of non-melanoma skin cancer. Papillomaviruses are host specific and are considered closely co-evolving with their hosts. Evolutionary incongruence between early genes and late genes has been reported among oncogenic genital alpha-papillomaviruses and considerably challenge phylogenetic reconstructions. We investigated the relationships of 29 beta-HPV (25 types plus four putative new types, subtypes, or variants) as inferred from codon aligned and amino acid sequence data of the genes E1, E2, E6, E7, L1, and L2 using likelihood, distance, and parsimony approaches. An analysis of a L1 fragment included additional nucleotide and amino acid sequences from seven non-human beta-papillomaviruses. Early genes and late genes evolution did not conflict significantly in beta-papillomaviruses based on partition homogeneity tests (p > or = 0.001). As inferred from the complete genome analyses, beta-papillomaviruses were monophyletic and segregated into four highly supported monophyletic assemblages corresponding to the species 1, 2, 3, and fused 4/5. They basically split into the species 1 and the remainder of beta-papillomaviruses, whose species 3, 4, and 5 constituted the sistergroup of species 2. beta-Papillomaviruses have been isolated from humans, apes, and monkeys, and phylogenetic analyses of the L1 fragment showed non-human papillomaviruses highly polyphyletic nesting within the HPV species. Thus, host and virus phylogenies were not congruent in beta-papillomaviruses, and multiple invasions across species borders may contribute (additionally to host-linked evolution) to their diversification.

  9. Complete amino acid sequence and in vitro expression of rat NF-M, the middle molecular weight neurofilament protein.

    PubMed

    Napolitano, E W; Chin, S S; Colman, D R; Liem, R K

    1987-08-01

    A lambda gtII expression library was prepared from rat brain and screened with a polyclonal antiserum, which recognizes both NF-H and NF-M. An NF-M cDNA clone (pNF-M3C = 1.6 kb) was isolated and characterized. The fusion protein of NF-M3C, when used as an affinity matrix for the anti-neurofilament serum, isolated a subpopulation of antibodies specific for NF-M. Northern analysis demonstrates a single band of approximately 3000 nt and a constant message level for NF-M during postnatal development from postnatal day 0 (PO) to adulthood. Using pNF-M3C as a probe, a second cDNA clone was isolated from a lambda gtII rat brain expression library (pNF-M2D = 2.7 kb). The 2 clones were sequenced and pNF-M2D was found to encode the entire rat NF-M protein. The calculated molecular weight is 95,600, which is only 65% of the molecular weight determined by SDS-PAGE. The amino acid sequence of rat NF-M shows the conserved rod segment present in all intermediate filament proteins. The molecule also contains an unusual C-terminal extension with stretches of glutamic acid, which could contribute to the anomalous migration of this protein on SDS-PAGE and the fact that NF-M does not readily assemble into filaments. The pNF-M2D clone was transcribed and translated in vitro utilizing a rabbit reticulocyte lysate system. The resulting radiolabeled translation products were unexpectedly shown to comigrate with purified rat NF-M on 1- and 2-dimensional gels, even though the translated protein is not phosphorylated.

  10. Plio-Quaternary coastal sequences, sea-level changes and coastal geodynamics :

    NASA Astrophysics Data System (ADS)

    Pedoja, Kevin; Husson, Laurent; Nexer, Maëlle; Regard, Vincent; Delcaillau, Bernard

    2013-04-01

    Sequences of Plio-Quaternary shorelines generated by sea-level cyclicity and tectonics massively shape the coasts worldwide. Here, we compile a worldwide synthesis of sea-level changes for the following highstands : MIS 1, MIS 3, MIS5e, and MIS 11 and explore the relationships to regional geodynamics. We identified 896 sequences including the MIS 5e benchmark, out of which 177 also feature the highest Holocene shorelines, 6 where the elevation of the MIS 3 shorelines are known, and 40 including that of MIS 11 shorelines. We considered 8 main geodynamical contexts for a statistical analysis: passive margin (337 sites, 14 domains, U=0.06 ± 0.01 mm/a), hot spot chains (45 sites, 14 domains, U=0.02 ± 0.02 mm/a), rifts (45 sites, 2 domains, U=0.12 ± 0.02 mm/a), ridges (3 sites, 1 domain, U=0.14 ± 0.02 mm/a), transform plate boundaries (119 sites, 7 domains, U=0.25 ± 0.03 mm/a), intra-oceanic subductions (131 sites, 4 domains, U=0.43 ± 0.04 mm/a), continental subductions underneath oceanic plates (12 sites, 1 domain, U=0.54+/- 0.04 mm/a), oceanic subductions underneath continental plates (166 sites, 7 domains, U=0.06 ± 0.01 mm/a), intra-continental convergence (10 sites, 2 domains, U=1.47 ± 0.08 mm/a). Counter-intuitive is the fact that passive margin are ubiquitously uplifting, while tectonic segmentation is more important on active margins. Additionally, 511 sites document the elevation of the highest shoreline of the sequence, from which we extrapolate a minimum age of onset for the sequence (based on MIS 5e uplift rates). 99% (511 sites) were formed at least during Pliocene and/or Quaternary, showing that potentially more ancient records are either eroded or non-existent.

  11. Isolation, amino acid sequence and biological characterization of an "aspartic-49" phospholipase A₂ from Bothrops (Rhinocerophis) ammodytoides venom.

    PubMed

    Clement, Herlinda; Costa de Oliveira, Vanessa; Zamudio, Fernando Z; Lago, Néstor R; Valdez-Cruz, Norma A; Bérnard Valle, Melisa; Hajos, Silvia E; Alagón, Alejandro; Possani, Lourival D; de Roodt, Adolfo R

    2012-12-01

    A phospholipase enzyme was separated by chromatography from the venom of the snake Bothrops (Rhinocerophis) ammodytoides and characterized. The experimentally determined molecular weight was 13,853.65 Da, and the full primary structure was determined by Edman degradation and mass spectrometry analysis. The enzyme contains 122 amino acids residues closely stabilized by 7 disulfide bridges with an isoelectric point of 6.13. Sequence comparison with other known secretory PLA2 shows that the enzyme isolated belongs to the group II, presenting an aspartic acid residue at position 48 (numbered by convention as Asp49) of the active site, and accordingly displaying enzymatic activity. The enzyme corresponds to 3% of the total mass of the venom. The enzyme is mildly toxic to mice. The intravenous LD₅₀ of this phospholipase in CD-1 mice was around 6 μg/g of mouse body weight (more exactly 117 μg/mouse of 20 g) and the minimal mortal dose (MMD) was estimated to be close to 10 μg/g. In contrast, the LD₅₀ of the venom was circa 2 μg/g mouse body weight. Toxicological analyses of the purified enzyme were performed in vitro and in vivo using experimental animals (mice and rats). The enzyme at high doses caused pulmonary congestion, intraperitoneal bleeding, inhibition of clot retraction and muscle tissue alterations with increasing of creatine kinase levels.

  12. Amino acid sequences of alpha-helical segments from S-carbosymethylkerateine-A. Complete sequence of a type-I segment.

    PubMed Central

    Gough, K H; Inglis, A S; Crewther, W G

    1978-01-01

    The amino acid sequence of a type-I helical segment from the low-sulphur protein (S-carboxymethylkerateine-A) of wool was determined by combining automatic and manual-sequencing data. Whereas in the type-II helical segment most of the cationic groups occur in pairs, 11 of the 22 anionic residues in the sequence of the type-I segment were situated next to a second anionic residue. This suggests possible interactions between type-I and type-II helical segments in alpha-keratin. As observed with the sequence of a type-II helical segment a model constructed on 3.6 residues per turn of helix shows a line of hydrophobic residues along the helix, thereby supporting the physicochemical evidence that the molecule is predominantly helical and forms part of a coiled-coil structure. Examination of the sequence data by predictive methods indicates the possibilty of extensive sections of alpha-helix interspersed with discontinuities. The molecule contains a number of regions with peptide sequences identical with those found by other workers after enzymic digestion of fractions from oxidized wool. Images Fig. 1. PMID:697725

  13. Draft Genome Sequences of Biosafety Level 2 Opportunistic Pathogens Isolated from the Environmental Surfaces of the International Space Station.

    PubMed

    Checinska Sielaff, Aleksandra; Singh, Nitin K; Allen, Jonathan E; Thissen, James; Jaing, Crystal; Venkateswaran, Kasthuri

    2016-12-29

    The draft genome sequences of 20 biosafety level 2 (BSL-2) opportunistic pathogens isolated from the environmental surfaces of the International Space Station (ISS) were presented. These genomic sequences will help in understanding the influence of microgravity on the pathogenicity and virulence of these strains when compared with Earth strains.

  14. Draft Genome Sequences of Biosafety Level 2 Opportunistic Pathogens Isolated from the Environmental Surfaces of the International Space Station

    PubMed Central

    Checinska Sielaff, Aleksandra; Singh, Nitin K.; Allen, Jonathan E.; Thissen, James; Jaing, Crystal

    2016-01-01

    The draft genome sequences of 20 biosafety level 2 (BSL-2) opportunistic pathogens isolated from the environmental surfaces of the International Space Station (ISS) were presented. These genomic sequences will help in understanding the influence of microgravity on the pathogenicity and virulence of these strains when compared with Earth strains. PMID:28034853

  15. Spermatogenesis of the lizard Lacerta vivipara: histological studies and amino acid sequence of a protamine lacertine 1.

    PubMed

    Martinage, A; Depeiges, A; Wouters, D; Morel, L; Sautière, P

    1996-06-01

    The lizard Lacerta vivipara is a seasonal breeder with a well characterized reproductive cycle. An histological study of the lizard testis has been performed at different stages of spermatogenesis and the nuclear basic proteins content was assessed by electrophoretical analysis. Two protamines, lacertines 1 and 2, are present in spermatozoa in April and May. We have isolated lacertine1 and characterized a protamine with a mass of 4,963.7 Da. Amino acid sequence of this protamine (41 residues) was established from data provided by automated Edman degradation. It is characterized by a basic amino acid stretch in the N- and C-terminal regions and by a central part which only consists of 3 different intermingled amino acids. This protamine presents 62% homology with scylliorhinine Z3 from dog-fish Scylliorhinus caniculus and 58% homology with quail protamine. The reported lizard protamine sequence is the first reptilian protamine sequence available so far.

  16. The amino acid sequence of the cytochrome c-554(547) from the chemolithotrophic bacterium Thiobacillus neapolitanus.

    PubMed Central

    Ambler, R P; Meyer, T E; Trudinger, P A; Kamen, M D

    1985-01-01

    An amino acid sequence is proposed for the cytochrome c-554(547) from the bacterium Thiobacillus neapolitanus N.C.I.B. 8539). It consists of a polypeptide chain of 91 residues, with a pair of haem-attachment cysteine residues at positions 15 and 18. There is similarity in sequence with each of the halves of the sequence of the dihaem cytochromes c4 and with a cytochrome c-554(548) from a halophilic strain of Paracoccus. Detailed evidence for the amino acid sequence of the protein has been deposited as Supplementary Publication SUP 50127 (11 pages) at the British Library (Lending Division), Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1985) 225, 5. PMID:2988504

  17. 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.

  18. Draft Genome Sequence of Escherichia coli O157:H7 ATCC 35150 and a Nalidixic Acid-Resistant Mutant Derivative

    PubMed Central

    Markell, James A.; Koziol, Adam G.

    2015-01-01

    Shiga toxin-producing Escherichia coli strains, occasionally isolated from food, are of public health importance. Here, we report on the 5.30-Mbp draft genome sequence of E. coli O157:H7 EDL931 (strain ATCC 35150) and the 5.32-Mbp draft genome sequence of a nalidixic acid-resistant mutant derivative used as a distinguishable control strain in food-testing laboratories. PMID:26205873

  19. Microwave-assisted acid and base hydrolysis of intact proteins containing disulfide bonds for protein sequence analysis by mass spectrometry.

    PubMed

    Reiz, Bela; Li, Liang

    2010-09-01

    Controlled hydrolysis of proteins to generate peptide ladders combined with mass spectrometric analysis of the resultant peptides can be used for protein sequencing. In this paper, two methods of improving the microwave-assisted protein hydrolysis process are described to enable rapid sequencing of proteins containing disulfide bonds and increase sequence coverage, respectively. It was demonstrated that proteins containing disulfide bonds could be sequenced by MS analysis by first performing hydrolysis for less than 2 min, followed by 1 h of reduction to release the peptides originally linked by disulfide bonds. It was shown that a strong base could be used as a catalyst for microwave-assisted protein hydrolysis, producing complementary sequence information to that generated by microwave-assisted acid hydrolysis. However, using either acid or base hydrolysis, amide bond breakages in small regions of the polypeptide chains of the model proteins (e.g., cytochrome c and lysozyme) were not detected. Dynamic light scattering measurement of the proteins solubilized in an acid or base indicated that protein-protein interaction or aggregation was not the cause of the failure to hydrolyze certain amide bonds. It was speculated that there were some unknown local structures that might play a role in preventing an acid or base from reacting with the peptide bonds therein.

  20. The effects of evening primrose oil, safflower oil and paraffin on plasma fatty acid levels in humans: choice of an appropriate placebo for clinical studies on primrose oil.

    PubMed

    Horrobin, D F; Ells, K M; Morse-Fisher, N; Manku, M S

    1991-04-01

    In a number of diseases, plasma levels of linoleic acid are normal or elevated while those of gamma-linolenic acid (18:3n-6, GLA) and further metabolites are below normal. Evening primrose oil (EPO), similar to safflower oil (SFO) except that it contains 8-9% of 18:3n-6, has been proposed as a therapeutic agent in these diseases, such as atopic eczema. There is argument as to whether an appropriate placebo for clinical studies on EPO should be an inert material such as paraffin, or a linoleic acid--containing oil such as SFO. We have therefore compared in normal humans the effects on plasma fatty acids of administering EPO, SFO and paraffin for 10 days. Paraffin had no effect on any fatty acid in any fraction. EPO raised the level of 20:3n-6 (dihomo-gamma-linolenic acid, DGLA) the immediate metabolite of GLA but had no significant effect on arachidonic acid. In surprising contrast, SFO raised the levels of linoleic and of arachidonic acids, without raising those of DGLA. This suggests that linoleic acid may be rapidly converted to arachidonic acid by a tightly linked enzyme sequence: GLA, in contrast, may be rapidly converted to DGLA but then only slowly on to arachidonic acid. These results are consistent with recent in vitro observations by others on rat hepatocytes and human fibroblasts.

  1. S-matrix Calculations of Energy Levels of the Lithium Isoelectronic Sequence

    SciTech Connect

    sapirstein, J; Cheng, K T

    2010-11-02

    A QED approach to the calculation of the spectra of the lithium isoelectronic sequence is implemented. A modified Furry representation based on the Kohn-Sham potential is used to evaluate all one- and two-photon diagrams with the exception of the two-loop Lamb shift. Three-photon diagrams are estimated with Hamiltonian methods. After incorporating recent calculations of the two-loop Lamb shift and recoil corrections a comprehensive tabulation of the 2s, 2p{sub 1/2} and 2p{sub 3/2} energy levels as well as the 2s - 2p{sub 1/2} and 2s - 2p{sub 3/2} transition energies for Z = 10 - 100 is presented.

  2. 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.

  3. Rare Variants Association Analysis in Large-Scale Sequencing Studies at the Single Locus Level

    PubMed Central

    Lu, Wenbin; Tzeng, Jung-Ying

    2016-01-01

    Genetic association analyses of rare variants in next-generation sequencing (NGS) studies are fundamentally challenging due to the presence of a very large number of candidate variants at extremely low minor allele frequencies. Recent developments often focus on pooling multiple variants to provide association analysis at the gene instead of the locus level. Nonetheless, pinpointing individual variants is a critical goal for genomic researches as such information can facilitate the precise delineation of molecular mechanisms and functions of genetic factors on diseases. Due to the extreme rarity of mutations and high-dimensionality, significances of causal variants cannot easily stand out from those of noncausal ones. Consequently, standard false-positive control procedures, such as the Bonferroni and false discovery rate (FDR), are often impractical to apply, as a majority of the causal variants can only be identified along with a few but unknown number of noncausal variants. To provide informative analysis of individual variants in large-scale sequencing studies, we propose the Adaptive False-Negative Control (AFNC) procedure that can include a large proportion of causal variants with high confidence by introducing a novel statistical inquiry to determine those variants that can be confidently dispatched as noncausal. The AFNC provides a general framework that can accommodate for a variety of models and significance tests. The procedure is computationally efficient and can adapt to the underlying proportion of causal variants and quality of significance rankings. Extensive simulation studies across a plethora of scenarios demonstrate that the AFNC is advantageous for identifying individual rare variants, whereas the Bonferroni and FDR are exceedingly over-conservative for rare variants association studies. In the analyses of the CoLaus dataset, AFNC has identified individual variants most responsible for gene-level significances. Moreover, single-variant results

  4. d-Amino Acid Levels in Perfused Mouse Brain Tissue and Blood: A Comparative Study.

    PubMed

    Weatherly, Choyce A; Du, Siqi; Parpia, Curran; Santos, Polan T; Hartman, Adam L; Armstrong, Daniel W

    2017-02-16

    The l-enantiomer is the predominant type of amino acid in all living systems. However, d-amino acids, once thought to be "unnatural", have been found to be indigenous even in mammalian systems and increasingly appear to be functioning in essential biological and neurological roles. Both d- and l-amino acid levels in the hippocampus, cortex, and blood samples from NIH Swiss mice are reported. Perfused brain tissues were analyzed for the first time, thereby eliminating artifacts due to endogenous blood, and decreased the mouse-to-mouse variability in amino acid levels. Total amino acid levels (l- plus d-enantiomers) in brain tissue are up to 10 times higher than in blood. However, all measured d-amino acid levels in brain tissue are typically ∼10 to 2000 times higher than blood levels. There was a 13% reduction in almost all measured d-amino acid levels in the cortex compared to those in the hippocampus. There is an approximate inverse relationship between the prevalence of an amino acid and the percentage of its d-enantiomeric form. Interestingly, glutamic acid, unlike all other amino acids, had no quantifiable level of its d-antipode. The bioneurological reason for the unique and conspicuous absence/removal of this d-amino acid is yet unknown. However, results suggest that d-glutamate metabolism is likely a unidirectional process and not a cycle, as per the l-glutamate/glutamine cycle. The results suggest that there might be unreported d-amino acid racemases in mammalian brains. The regulation and function of specific other d-amino acids are discussed.

  5. High-level production of poly (β-L: -malic acid) with a new isolated Aureobasidium pullulans strain.

    PubMed

    Zhang, Huili; Cai, Jin; Dong, Jiaqi; Zhang, Danping; Huang, Lei; Xu, Zhinan; Cen, Peilin

    2011-10-01

    Poly (β-L: -malic acid) (PMLA) is a water-soluble polyester with many attractive properties in chemical industry and medicine development. However, the low titer of PMLA in the available producer strains limits further industrialization efforts and restricts its many potential applications. In order to solve this problem, a new strain with the distinguished high productivity of PMLA was isolated from fresh plants samples. It was characterized as the candidate of Aureobasidium pullulans based on the morphology and phylogenetic analyses of the internal transcribed spacer sequences. After the optimization of culture conditions, the highest PMLA concentration (62.27 g l(-1)) could be achieved in the shake flask scale. In addition, the contribution of the carbon flux to exopolysaccharide (EPS) and PMLA could be regulated by the addition of CaCO₃ in the medium. This high-level fermentation process was further scaled up in the 10 l benchtop fermentor with a high PMLA concentration (57.2 g l(-1)) and productivity (0.35 g l(-1) h(-1)), which are the highest level in all the literature. Finally, the suitable acid hydrolysis conditions of PMLA were also investigated with regard to the production of L: -malic acid, and the kinetics of PMLA acid hydrolysis was modeled to simulate the whole degradation process. The present work paved the road to produce this multifunctional biomaterial (PMLA) at industrial scale and promised one alternative method to produce L: -malic acid in the future.

  6. Purification, characterization, gene cloning and nucleotide sequencing of D: -stereospecific amino acid amidase from soil bacterium: Delftia acidovorans.

    PubMed

    Hongpattarakere, Tipparat; Komeda, Hidenobu; Asano, Yasuhisa

    2005-12-01

    The D-amino acid amidase-producing bacterium was isolated from soil samples using an enrichment culture technique in medium broth containing D-phenylalanine amide as a sole source of nitrogen. The strain exhibiting the strongest activity was identified as Delftia acidovorans strain 16. This strain produced intracellular D-amino acid amidase constitutively. The enzyme was purified about 380-fold to homogeneity and its molecular mass was estimated to be about 50 kDa, on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was active preferentially toward D-amino acid amides rather than their L-counterparts. It exhibited strong amino acid amidase activity toward aromatic amino acid amides including D-phenylalanine amide, D-tryptophan amide and D-tyrosine amide, yet it was not specifically active toward low-molecular-weight D-amino acid amides such as D-alanine amide, L-alanine amide and L-serine amide. Moreover, it was not specifically active toward oligopeptides. The enzyme showed maximum activity at 40 degrees C and pH 8.5 and appeared to be very stable, with 92.5% remaining activity after the reaction was performed at 45 degrees C for 30 min. However, it was mostly inactivated in the presence of phenylmethanesulfonyl fluoride or Cd2+, Ag+, Zn2+, Hg2+ and As3+ . The NH2 terminal and internal amino acid sequences of the enzyme were determined; and the gene was cloned and sequenced. The enzyme gene damA encodes a 466-amino-acid protein (molecular mass 49,860.46 Da); and the deduced amino acid sequence exhibits homology to the D-amino acid amidase from Variovorax paradoxus (67.9% identity), the amidotransferase A subunit from Burkholderia fungorum (50% identity) and other enantioselective amidases.

  7. New alleles of FATB-1A to reduce palmitic acid levels in soybean

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In wild-type soybeans, palmitic acid typically constitutes 10% of the total seed oil. Palmitic acid is a saturated fat linked to increased cholesterol levels, and reducing levels of saturated fats in soybean oil has been a breeding target. To identify novel and useful variation that could help in re...

  8. Plasma Amino Acid Levels in Children with Autism and Their Families.

    ERIC Educational Resources Information Center

    Aldred, Sarah; Moore, Kieran M.; Fitzgerald, Michael; Waring, Rosemary H.

    2003-01-01

    Plasma amino acid levels were measured in autistic (n=12), Asperger syndrome (n=11) patients, their parents and siblings. Patients with autism or Asperger syndrome and their siblings and parents all had raised glutamic acid, phenyalanine, asparagine, tyrosine, alanine, and lysine levels than age-matched controls. Results suggest dysregulated amino…

  9. Effects of Acidic Peptide Size and Sequence on Trivalent Praseodymium Adduction and Electron Transfer Dissociation Mass Spectrometry.

    PubMed

    Commodore, Juliette J; Cassady, Carolyn J

    2017-02-07

    Using the lanthanide ion praseodymium, Pr(III), metallated ion formation and electron transfer dissociation (ETD) were studied for 25 biological and model acidic peptides. For chain lengths of seven or more residues, even highly acidic peptides that can be difficult to protonate by electrospray ionization will metallate and undergo abundant ETD fragmentation. Peptides composed of predominantly acidic residues form only the deprotonated ion, [M + Pr - H](2+) ; this ion yields near complete ETD sequence coverage for larger peptides. Peptides with a mixture of acidic and neutral residues, generate [M + Pr](3+) , which cleaves between every residue for many peptides. Acidic peptides that contain at least one residue with a basic side chain also produce the protonated ion, [M + Pr + H](4+) ; this ion undergoes the most extensive sequence coverage by ETD. Primarily metallated and non-metallated c- and z-ions form for all peptides investigated. Metal adducted product ions are only present when at least half of the peptide sequence can be incorporated into the ion; this suggests that the metal ion simultaneously attaches to more than one acidic site. The only site consistently lacking dissociation is at the N-terminal side of a proline residue. Increasing peptide chain length generates more backbone cleavage for metal-peptide complexes with the same charge state. For acidic peptides with the same length, increasing the precursor ion charge state from 2+ to 3+ also leads to more cleavage. The results of this study indicate that highly acidic peptides can be sequenced by ETD of complexes formed with Pr(III).

  10. The Association between Marine n-3 Polyunsaturated Fatty Acid Levels and Survival after Renal Transplantation

    PubMed Central

    Jenssen, Trond; Hartmann, Anders; Diep, Lien M.; Dahle, Dag O.; Reisæter, Anna V.; Bjerve, Kristian S.; Christensen, Jeppe H.; Schmidt, Erik B.; Svensson, My

    2015-01-01

    Background and objectives Several studies have reported beneficial cardiovascular effects of marine n-3 polyunsaturated fatty acids. To date, no large studies have investigated the potential benefits of marine n-3 polyunsaturated fatty acids in recipients of renal transplants. Design, setting, participants, & measurements In this observational cohort study of 1990 Norwegian recipients of renal transplants transplanted between 1999 and 2011, associations between marine n-3 polyunsaturated fatty acid levels and mortality were investigated by stratified analysis and multivariable Cox proportional hazard regression analysis adjusting for traditional and transplant-specific mortality risk factors. Marine n-3 polyunsaturated fatty acid levels in plasma phospholipids were measured by gas chromatography in a stable phase 10 weeks after transplantation. Results There were 406 deaths (20.4%) during a median follow-up period of 6.8 years. Mortality rates were lower in patients with high marine n-3 polyunsaturated fatty acid levels (≥7.95 weight percentage) compared with low levels (<7.95 weight percentage) for all age categories (pooled mortality rate ratio estimate, 0.69; 95% confidence interval, 0.57 to 0.85). When divided into quartiles according to marine n-3 polyunsaturated fatty acid levels, patients in the upper quartile compared with the lower quartile had a 56% lower risk of death (adjusted hazard ratio, 0.44; 95% confidence interval, 0.26 to 0.75) using multivariable Cox proportional hazard regression analysis. There was a lower hazard ratio for death from cardiovascular disease with high levels of marine n-3 polyunsaturated fatty acid and a lower hazard ratio for death from infectious disease with high levels of the marine n-3 polyunsaturated fatty acid eicosapentaenoic acid, whereas there was no association between total or individual marine n-3 polyunsaturated fatty acid levels and cancer mortality. Conclusions Higher plasma phospholipid marine n-3

  11. Determination of the complete amino acid sequence for the coat protein of brome mosaic virus by time-of-flight mass spectrometry. Evidence for mutations associated with change of propagation host.

    PubMed

    She, Y M; Haber, S; Seifers, D L; Loboda, A; Chernushevich, I; Perreault, H; Ens, W; Standing, K G

    2001-06-08

    Time-of-flight mass spectrometry (TOFMS) has been applied to determine the complete coat protein amino acid sequences of a number of distinct brome mosaic virus (BMV) isolates. Ionization was carried out by both electrospray ionization and matrix-assisted laser desorption/ionization (MALDI). After determining overall coat protein masses, the proteins were digested with trypsin or Lys-C proteinases, and the digestion products were analyzed in a MALDI QqTOF mass spectrometer. The N terminus of the coat protein was found to be acetylated in each BMV isolate analyzed. In one isolate (BMV-Valverde), the amino acid sequence was identical to that predicted from the cDNA sequence of the "type" isolate, but deviations from the predicted amino acid sequence were observed for all the other isolates analyzed. When isolates were propagated in different host taxa, modified coat protein sequences were observed in some cases, along with the original sequence. Sequencing by TOFMS may therefore provide a basis for monitoring the effects of host passaging on a virus at the molecular level. Such TOFMS-based analyses assess the complete profiles of coat protein sequences actually present in infected tissues. They are therefore not subject to the selection biases inherent in deducing such sequences from reverse-transcribed viral RNA and cloning the resulting cDNA.

  12. Serotonin (5-HT) receptor 5A sequence variants affect human plasma triglyceride levels

    PubMed Central

    Zhang, Y.; Smith, E. M.; Baye, T. M.; Eckert, J. V.; Abraham, L. J.; Moses, E. K.; Kissebah, A. H.; Martin, L. J.

    2010-01-01

    Neurotransmitters such as serotonin (5-hydroxytryptamine, 5-HT) work closely with leptin and insulin to fine-tune the metabolic and neuroendocrine responses to dietary intake. Losing the sensitivity to excess food intake can lead to obesity, diabetes, and a multitude of behavioral disorders. It is largely unclear how different serotonin receptor subtypes respond to and integrate metabolic signals and which genetic variations in these receptor genes lead to individual differences in susceptibility to metabolic disorders. In an obese cohort of families of Northern European descent (n = 2,209), the serotonin type 5A receptor gene, HTR5A, was identified as a prominent factor affecting plasma levels of triglycerides (TG), supported by our data from both genome-wide linkage and targeted association analyses using 28 publicly available and 12 newly discovered single nucleotide polymorphisms (SNPs), of which 3 were strongly associated with plasma TG levels (P < 0.00125). Bayesian quantitative trait nucleotide (BQTN) analysis identified a putative causal promoter SNP (rs3734967) with substantial posterior probability (P = 0.59). Functional analysis of rs3734967 by electrophoretic mobility shift assay (EMSA) showed distinct binding patterns of the two alleles of this SNP with nuclear proteins from glioma cell lines. In conclusion, sequence variants in HTR5A are strongly associated with high plasma levels of TG in a Northern European population, suggesting a novel role of the serotonin receptor system in humans. This suggests a potential brain-specific regulation of plasma TG levels, possibly by alteration of the expression of HTR5A. PMID:20388841

  13. Strain-Level Discrimination of Shiga Toxin-Producing Escherichia coli in Spinach Using Metagenomic Sequencing

    PubMed Central

    Leonard, Susan R.; Mammel, Mark K.; Lacher, David W.; Elkins, Christopher A.

    2016-01-01

    Consumption of fresh bagged spinach contaminated with Shiga toxin-producing Escherichia coli (STEC) has led to severe illness and death; however current culture-based methods to detect foodborne STEC are time consuming. Since not all STEC strains are considered pathogenic to humans, it is crucial to incorporate virulence characterization of STEC in the detection method. In this study, we assess the comprehensiveness of utilizing a shotgun metagenomics approach for detection and strain-level identification by spiking spinach with a variety of genomically disparate STEC strains at a low contamination level of 0.1 CFU/g. Molecular serotyping, virulence gene characterization, microbial community analysis, and E. coli core gene single nucleotide polymorphism (SNP) analysis were performed on metagenomic sequence data from enriched samples. It was determined from bacterial community analysis that E. coli, which was classified at the phylogroup level, was a major component of the population in most samples. However, in over half the samples, molecular serotyping revealed the presence of indigenous E. coli which also contributed to the percent abundance of E. coli. Despite the presence of additional E. coli strains, the serotype and virulence genes of the spiked STEC, including correct Shiga toxin subtype, were detected in 94% of the samples with a total number of reads per sample averaging 2.4 million. Variation in STEC abundance and/or detection was observed in replicate spiked samples, indicating an effect from the indigenous microbiota during enrichment. SNP analysis of the metagenomic data correctly placed the spiked STEC in a phylogeny of related strains in cases where the indigenous E. coli did not predominate in the enriched sample. Also, for these samples, our analysis demonstrates that strain-level phylogenetic resolution is possible using shotgun metagenomic data for determining the genomic relatedness of a contaminating STEC strain to other closely related E

  14. Plasma ω-3 fatty acid levels negatively and ω-6 fatty acid levels positively associated with other cardiovascular risk factors including homocysteine in severe obese subjects.

    PubMed

    Mehmetoglu, Idris; Yerlikaya, F Hümeyra; Kurban, Sevil; Polat, Hakkı

    2012-01-01

    Obesity and homocysteine (tHcy) are important risk factors for cardiovascular diseases (CVD). Plasma omega-3 fatty acids (ω-3 FAs) and omega-6 fatty acids (ω-6 FAs) are essential fatty acids with diverse biological effects in human health and disease. We have investigated the relation of plasma ω-3 FAs and ω-6 FAs levels with other cardiovascular risk factors including tHcy in severe obese subjects. This study was performed on 96 severe obese and 65 normal weight subjects. Plasma fatty acid composition was measured by GC/MS and serum tHcy level was measured by HPLC methods. There were no differences between groups in terms of concentrations of serum tHcy, plasma ω-3 FAs, ω-6 FAs and ω-3/ω-6 ratio, whereas serum vitamin B-12 (p<0.01) and folic acid (p<0.05) levels were lower than those of the normal weight subjects. Homocysteine positively correlated with ω-6 FAs and negatively correlated with ω-3 FAs in severe obese and normal weight subjects. Serum vitamin B-12 positively correlated with ω-3 FAs (p<0.01) and ω-3/ω-6 ratio (p<0.01) and negatively correlated with ω-6 FAs (p<0.05) in severe obese subjects. Serum folic acid positively correlated with ω-3 FAs (p<0.01) in severe obese subjects. Our results suggest an association between the plasma ω-3 FAs and ω-6 FAs and serum tHcy concentrations in severe obese and normal weight subjects. Low levels vitamin B-12 and folic acid may have been responsible for the elevated tHcy levels in severe obese subjects, increasing the risk for future development of cardiovascular diseases.

  15. 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.

  16. The Effects of Lowering Uric Acid Levels Using Allopurinol on Components of Metabolic Syndrome

    PubMed Central

    Heimbach, Esther J.; Bowden, Rodney G.; Griggs, Jackson O.; Beaujean, A. Alexander; Doyle, Eva I.; Doyle, Robert D.

    2012-01-01

    Background Researchers have reported an independent direct relationship between lipid levels and hyperuricemia with MetS. The purpose of this study was to determine the relationship between serum uric acid levels and lipids among patients on allopurinol. Methods A retrospective secondary data analysis was conducted on 66 adult patients from a family health clinic in Central Texas. Medical records used were recorded during a nine year period (2002 - 2010) ascertaining the relationship between uric acid and lipids. Results Spearman correlations revealed a weak correlation between uric acid and total cholesterol, a weak correlation between uric acid and triglycerides and LDL-C. A weak inverse correlation was discovered between uric acid and HDL-C. A moderate correlation was discovered when all lipid variables combined were compared to uric acid. Conclusions We discovered LDL-C and triglycerides to be significant predictors of uric acid with weak correlations. Additionally, weak correlations existed between uric acid and total cholesterol and HDL-C with an inverse relationship discovered with HDL-C. These findings support the literature suggesting that uric acid is more likely to be associated with total cholesterol and triglycerides. In addition, new discoveries serve as an indication that LDL-C may also be associated with uric acids levels. The mechanism by which uric acid may regulate lipids is elusive but suggestions have included suppression of lipid peroxidase and decreases in critical lipase activity.

  17. Hippuric Acid Levels in Paint Workers at Steel Furniture Manufacturers in Thailand

    PubMed Central

    Decharat, Somsiri

    2014-01-01

    Background The aims of this study were to determine hippuric acid levels in urine samples, airborne toluene levels, acute and chronic neurological symptoms, and to describe any correlation between urinary hippuric acid and airborne toluene. Methods The hippuric acid concentration in the urine of 87 paint workers exposed to toluene at work (exposed group), and 87 nonexposed people (control group) was studied. Study participants were selected from similar factories in the same region. Urine samples were collected at the end of a shift and analyzed for hippuric acid by high performance liquid chromatography. Air samples for the estimation of toluene exposure were collected with diffusive personal samplers and the toluene quantified using gas–liquid chromatography. The two groups were also interviewed and observed about their work practices and health. Results The median of the 87 airborne toluene levels was 55 ppm (range, 12–198 ppm). The median urinary hippuric acid level was 800 mg/g creatinine (range, 90–2547 mg/g creatinine). A statistically significant positive correlation was found between airborne toluene exposure and urine hippuric acid levels (r = 0.548, p < 0.01). Workers with acute symptoms had significantly higher hippuric acid levels than those who did not (p < 0.05). It was concluded that there was a significant correlation between toluene exposure, hippuric acid levels, and health (p < 0.001). Conclusion There appears to be a significant correlation between workers exposure to toluene at work, their urine hippuric acid levels, and resulting symptoms of poor health. Improvements in working conditions and occupational health education are required at these workplaces. There was good correlation between urinary hippuric acid and airborne toluene levels. PMID:25516817

  18. Messenger ribonucleic acid levels in disrupted human anterior cruciate ligaments.

    PubMed

    Lo, Ian K Y; Marchuk, Linda; Hart, David A; Frank, Cyril B

    2003-02-01

    Thirty patients had anterior cruciate ligament reconstruction for ongoing instability. Two groups were defined according to gross morphologic features identified during reconstruction: anterior cruciate ligament disruptions with scars attached to a structure in the joint and disruptions without reattachments. Reverse transcription polymerase chain reaction for a subset of extracellular matrix molecules, proteinases, and proteinase inhibitors was done on samples of scarred anterior cruciate ligament tissue removed during reconstructive surgery. Results of the nonattached scar group showed significantly increased mRNA levels for Type I collagen, and an increased Type I to Type III collagen ratio compared with that for the attached scar group. In the first year after injury, decorin mRNA levels in the nonattached scar group also were significantly higher than in the attached scar group. Biglycan mRNA levels in the nonattached scar group correlated closely with Type I collagen mRNA levels. These results suggest differences in cellular expression in torn anterior cruciate ligaments that attach to structures in the joint versus those which do not. Although the molecular mechanisms responsible for these differences have not been delineated, different molecular signals may influence the gross morphologic features of anterior cruciate ligament disruptions or alternatively, differing gross morphologic features may be subject to different mechanical loads leading to altered molecular expression. However, the finding of endogenous cellular activity in injured anterior cruciate ligaments raises the possibility that this activity may be enhanced to improve outcomes.

  19. Molecular cloning and sequencing of a cDNA encoding the thioesterase domain of the rat fatty acid synthetase.

    PubMed

    Naggert, J; Witkowski, A; Mikkelsen, J; Smith, S

    1988-01-25

    A cloned cDNA containing the entire coding sequence for the long-chain S-acyl fatty acid synthetase thioester hydrolase (thioesterase I) component as well as the 3'-noncoding region of the fatty acid synthetase has been isolated using an expression vector and domain-specific antibodies. The coding region was assigned to the thioesterase I domain by identification of sequences coding for characterized peptide fragments, amino-terminal analysis of the isolated thioesterase I domain and the presence of the serine esterase active-site sequence motif. The thioesterase I domain is 306 amino acids long with a calculated molecular mass of 33,476 daltons; its DNA is flanked at the 5'-end by a region coding for the acyl carrier protein domain and at the 3'-end by a 1,537-base pairs-long noncoding sequence with a poly(A) tail. The thioesterase I domain exhibits a low, albeit discernible, homology with the discrete medium-chain S-acyl fatty acid synthetase thioester hydrolases (thioesterase II) from rat mammary gland and duck uropygial gland, suggesting a distant but common evolutionary ancestry for these proteins.

  20. Predicting subcellular localization of proteins based on their N-terminal amino acid sequence.

    PubMed

    Emanuelsson, O; Nielsen, H; Brunak, S; von Heijne, G

    2000-07-21

    A neural network-based tool, TargetP, for large-scale subcellular location prediction of newly identified proteins has been developed. Using N-terminal sequence information only, it discriminates between proteins destined for the mitochondrion, the chloroplast, the secretory pathway, and "other" localizations with a success rate of 85% (plant) or 90% (non-plant) on redundancy-reduced test sets. From a TargetP analysis of the recently sequenced Arabidopsis thaliana chromosomes 2 and 4 and the Ensembl Homo sapiens protein set, we estimate that 10% of all plant proteins are mitochondrial and 14% chloroplastic, and that the abundance of secretory proteins, in both Arabidopsis and Homo, is around 10%. TargetP also predicts cleavage sites with levels of correctly predicted sites ranging from approximately 40% to 50% (chloroplastic and mitochondrial presequences) to above 70% (secretory signal peptides). TargetP is available as a web-server at http://www.cbs.dtu.dk/services/TargetP/.

  1. Purification, properties, and partial amino acid sequences of thermostable xylanases from Streptomyces thermoviolaceus OPC-520

    SciTech Connect

    Tsujibo, Hiroshi; Miyamoto, Katsushiro; Kuda, Takashi; Minami, Kazushi; Sakamoto, Takashi; Inamori, Yoshihiko ); Hasegawa, Toru )

    1992-01-01

    Two types of xylanases (1,4-{beta}-D-xylan xylanohydrolase, EC 3.2.1.8) were isolated from the culture filtrate of a thermophilic actinomycete, Streptomyces thermoviolaceus OPC-520. The enzymes (STX-I and STX-II) were purified by chromatography with DEAE-Toyopearl 650 M, CM-Toyopearl 650 M, Sephadex G-75, Phenyl-Toyopearl 650 M, and Mono Q HR. The purified enzymes showed single bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The molecular weights of STX-I and STX-II were 54,000 and 33,000, respectively. The pIs were 4.2 (STX-I) and 8.0 (STX-II). The optimum pH levels for the activity of STX-I and STX-II were pH 7.0. The optimum temperature for the activity of STX-I was 70C, and that for the activity of STX-II was 60C. The enzymes were completely inhibited by N-bromosuccinimide. The enzymes degraded xylan, producing xylose and xylobiose as the predominant products, indicating that they were endoxylanases. STX-I showed high sequence homology with the exoglucanase from Cellulomonas fimi (47% homology), and STX-II showed high sequence homology with the xylanase from Bacillus pumilus (46% homology).

  2. Plasma D-lactic acid level: a useful marker to distinguish perforated from acute simple appendicitis.

    PubMed

    Demircan, Mehmet; Cetin, Selma; Uguralp, Sema; Sezgin, Nurzen; Karaman, Abdurrahman; Gozukara, Engin M

    2004-10-01

    Early diagnosis of perforated appendicitis is important for reducing morbidity rates. The aim of this study was to determine the value and utility of plasma D-lactic acid levels in identifying the type of appendicitis. In this clinical study, plasma D-lactic acid levels were assessed in 44 consecutive paediatric patients (23 with acute appendicitis, 21 with perforated appendicitis) before laparotomy. D-lactic acid levels were determined by an enzymatic spectrophotometric technique using a D-lactic acid dehydrogenase kit. Patients with perforated appendicitis had higher D-lactic acid levels (3.970 +/- 0.687 mg/dL) than patients in the control group (0.478 +/- 0.149 mg/dL) and patients with acute appendicitis (1.409 +/- 0.324 mg/dL; p < 0.05). For a plasma D-lactic acid level greater than 2.5 mg/dL, the sensitivity and specificity of the D-lactic acid assay were 96% and 87%, respectively. The positive predictive value was 87%, the negative predictive value was 96%, and the diagnostic value was 91%. These results suggest that the measurement of plasma D-lactic acid levels may be a useful adjunct to clinical and radiological findings in distinguishing perforated from acute non-perforated appendicitis in children.

  3. 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

  4. Inter-level Scaffolding and Sequences of Representational Activities in Teaching a Chemical System with Graphical Simulations

    NASA Astrophysics Data System (ADS)

    Li, Na; Black, John B.

    2016-10-01

    Chemistry knowledge can be represented at macro-, micro- and symbolic levels, and learning a chemistry topic requires students to engage in multiple representational activities. This study focused on scaffolding for inter-level connection-making in learning chemistry knowledge with graphical simulations. We also tested whether different sequences of representational activities produced different student learning outcomes in learning a chemistry topic. A sample of 129 seventh graders participated in this study. In a simulation-based environment, participants completed three representational activities to learn several ideal gas law concepts. We conducted a 2 × 3 factorial design experiment. We compared two scaffolding conditions: (1) the inter- level scaffolding condition in which participants received inter-level questions and experienced the dynamic link function in the simulation-based environment and (2) the intra- level scaffolding condition in which participants received intra-level questions and did not experience the dynamic link function. We also compared three different sequences of representational activities: macro-symbolic-micro, micro-symbolic-macro and symbolic-micro-macro. For the scaffolding variable, we found that the inter- level scaffolding condition produced significantly better performance in both knowledge comprehension and application, compared to the intra- level scaffolding condition. For the sequence variable, we found that the macro-symbolic-micro sequence produced significantly better knowledge comprehension performance than the other two sequences; however, it did not benefit knowledge application performance. There was a trend that the treatment group who experienced inter- level scaffolding and the micro-symbolic-macro sequence achieved the best knowledge application performance.

  5. 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.

  6. Solubility Challenges in High Concentration Monoclonal Antibody Formulations: Relationship with Amino Acid Sequence and Intermolecular Interactions.

    PubMed

    Pindrus, Mariya; Shire, Steven J; Kelley, Robert F; Demeule, Barthélemy; Wong, Rita; Xu, Yiren; Yadav, Sandeep

    2015-11-02

    The purpose of this work was to elucidate the molecular interactions leading to monoclonal antibody self-association and precipitation and utilize biophysical measurements to predict solubility behavior at high protein concentration. Two monoclonal antibodies (mAb-G and mAb-R) binding to overlapping epitopes were investigated. Precipitation of mAb-G solutions was most prominent at high ionic strength conditions and demonstrated strong dependence on ionic strength, as well as slight dependence on solution pH. At similar conditions no precipitation was observed for mAb-R solutions. Intermolecular interactions (interaction parameter, kD) related well with high concentration solubility behavior of both antibodies. Upon increasing buffer ionic strength, interactions of mAb-R tended to weaken, while those of mAb-G became more attractive. To investigate the role of amino acid sequence on precipitation behavior, mutants were designed by substituting the CDR of mAb-R into the mAb-G framework (GM-1) or deleting two hydrophobic residues in the CDR of mAb-G (GM-2). No precipitation was observed at high ionic strength for either mutant. The molecular interactions of mutants were similar in magnitude to those of mAb-R. The results suggest that presence of hydrophobic groups in the CDR of mAb-G may be responsible for compromising its solubility at high ionic strength conditions since deleting these residues mitigated the solubility issue.

  7. Genome level analysis of bacteriocins of lactic acid bacteria.

    PubMed

    Singh, Neetigyata Pratap; Tiwari, Abhay; Bansal, Ankiti; Thakur, Shruti; Sharma, Garima; Gabrani, Reema

    2015-06-01

    Bacteriocins are antimicrobial peptides which are ribosomally synthesized by mainly all bacterial species. LABs (lactic acid bacteria) are a diverse group of bacteria that include around 20 genera of various species. Though LABs have a tremendous potential for production of anti-microbial peptides, this group of bacteria is still underexplored for bacteriocins. To study the diversity among bacteriocin encoding clusters and the putative bacteriocin precursors, genome mining was performed on 20 different species of LAB not reported to be bacteriocin producers. The phylogenetic tree of gyrB, rpoB, and 16S rRNA were constructed using MEGA6 software to analyze the diversity among strains. Putative bacteriocins operons identified were found to be diverse and were further characterized on the basis of physiochemical properties and the secondary structure. The presence of at least two cysteine residues in most of the observed putative bacteriocins leads to disulphide bond formation and provide stability. Our data suggests that LABs are prolific source of low molecular weight non modified peptides.

  8. Frequencies of amino acid strings in globular protein sequences indicate suppression of blocks of consecutive hydrophobic residues

    PubMed Central

    Schwartz, Russell; Istrail, Sorin; King, Jonathan

    2001-01-01

    Patterns of hydrophobic and hydrophilic residues play a major role in protein folding and function. Long, predominantly hydrophobic strings of 20–22 amino acids each are associated with transmembrane helices and have been used to identify such sequences. Much less attention has been paid to hydrophobic sequences within globular proteins. In prior work on computer simulations of the competition between on-pathway folding and off-pathway aggregate formation, we found that long sequences of consecutive hydrophobic residues promoted aggregation within the model, even controlling for overall hydrophobic content. We report here on an analysis of the frequencies of different lengths of contiguous blocks of hydrophobic residues in a database of amino acid sequences of proteins of known structure. Sequences of three or more consecutive hydrophobic residues are found to be significantly less common in actual globular proteins than would be predicted if residues were selected independently. The result may reflect selection against long blocks of hydrophobic residues within globular proteins relative to what would be expected if residue hydrophobicities were independent of those of nearby residues in the sequence. PMID:11316883

  9. 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

  10. Compared with stearic acid, palmitic acid increased the yield of milk fat and improved feed efficiency across production level of cows.

    PubMed

    Rico, J E; Allen, M S; Lock, A L

    2014-02-01

    The effects of dietary palmitic and stearic acids on feed intake, yields of milk and milk components, and feed efficiency of dairy cows were evaluated in an experiment with a crossover arrangement of treatments with a covariate period. Cows with a wide range of milk production (38 to 65 kg/d) were used to determine if response to fat supplementation varied according to production level. Thirty-two Holstein cows (143 ± 61 d in milk) were assigned randomly to a treatment sequence within level of milk production. Treatments were diets supplemented (2% of diet dry matter) with palmitic acid (PA; 97.9% C16:0) or stearic acid (SA; 97.4% C18:0). Treatment periods were 21 d and cows were fed a nonfat supplemented diet for 14 d immediately before the first treatment period. The final 4d of each period were used for sample and data collection. Milk production measured during the covariate period (preliminary milk yield) was used as the covariate. No interactions were detected between treatment and preliminary milk yield for the production response variables measured. Compared with SA, the PA treatment increased milk fat concentration (3.66 vs. 3.55%) and yield (1.68 vs. 1.59 kg/d), and 3.5% fat-corrected milk yield (47.5 vs. 45.6 kg/d). Treatment did not affect dry matter intake, milk yield, milk protein yield, body weight, or body condition score. Milk protein concentration was lower for PA compared with SA treatment (3.24 vs. 3.29%). The PA treatment increased feed efficiency (3.5% fat-corrected milk yield/dry matter intake) compared with SA (1.48 vs. 1.40). The increase in milk fat yield by PA was entirely accounted for by a 24% increase in 16-carbon fatty acid output into milk. Yields of de novo (3.2%) and preformed fatty acids (2.9%) were only slightly decreased by PA relative to SA. The PA treatment increased plasma concentration of nonesterified fatty acids (96.3 vs. 88.2 μEq/L) and glucose (56.6 vs. 55.7 mg/dL) compared with SA, but insulin and

  11. 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.

  12. Chemical interactions between amino acid and RNA: multiplicity of the levels of specificity explains origin of the genetic code

    NASA Astrophysics Data System (ADS)

    Seligmann, Hervé; Amzallag, Nissim

    2002-11-01

    The emergence of the genetic code remains an enigma. Proposed mechanisms are based on random, historical, thermodynamic and natural selection. However, they introduce chance as a key factor for overcoming the difficulties encountered by the model. We propose here a model in which three successive levels of chemical specificity generated the nucleotide assignments of amino acids in the genetic code. The first level results from hydrophobic and stereospecific interactions between amino acids and short oligonucleotides (termed oligons). The second and third levels of specificity are determined by conditions of energy transfer from loaded oligons (amino acid-oligomer covalently linked) to formation of phosphodiester bond (second level of specificity) and peptidic bond (third level of specificity), while these reactions are catalyzed by RNA templates. This model is sustained by the relationships observed between dipole moments of the nucleotides (forming the anticodon) and reactivity of the amino acyl linkage of the loaded oligon. Moreover, analysis of modern tRNAs reveals that they were probably generated by loose duplication of the nucleotide sequence forming the oligons, after emergence of the 'genetic code.' Indeed, the similarity of nucleotide composition with that of the anticodon decreases with the tRNA domain's distance from the anticodon, but the acceptor stem is relatively more similar to the anticodon than other stems closer to it. This would be because energy transfer constraints that existed between anticodon and amino acid in prebiotic loaded oligonucleotides still affect the structures of modern tRNA acceptor stems. In the model presented, the genetic code is inherent to the most archaic 'molecular physiology' in protolife, even before emergence of a functional 'protein world.' Simple physical processes, in which a level of specificity is integrated in an emerging meta-structure expressing new properties, generate a parsimonious and realistic explanation

  13. delta-Aminolevulinic acid dehydratase activity, urinary delta-aminolevulinic acid concentration and zinc protoporphyrin level among people with low level of lead exposure.

    PubMed

    Wang, Qi; Zhao, Huan-hu; Chen, Jian-wei; Hao, Qiao-ling; Gu, Kang-ding; Zhu, Ye-xiang; Zhou, Yi-kai; Ye, Lin-xiang

    2010-01-01

    To evaluate the relationship of delta-aminolevulinic acid dehydratase (ALAD) activity, urinary delta-aminolevulinic acid (ALAU) level and blood zinc protoporphyrin (ZPP) concentration to low blood lead (PbB) levels, these biomarkers were determined for all subjects enrolled from a rural area of southeast China where people had low levels of exposure to lead. The mean values of PbB, ALAD, ALAU and ZPP were 67.11 microg/L (SD: 1.654, range: 10.90-514.04), 339.66 nmol ml(-1)h(-1) (1.419, 78.33-793.13), 20.64 microg/L (1.603, 2.00-326.00), and 0.14 micromol/L (3.437, 0.01-2.26), respectively. ALAD was inversely associated with low levels of PbB. ZPP was inversely related to low levels of PbB but positively related to relatively higher levels of PbB. Alcohol drinking contributed to low ALAD in men. Women had higher ZPP than men. ALAU had no significant association with PbB. In conclusion, ALAD possibly has a non-linear relation with low to moderate levels of PbB. At moderate levels of PbB, ZPP increases with increasing levels of PbB. ALAU is not suitable as an indicator for low levels of lead exposure.

  14. Limiting amino acids in bengal gram (Cicer arietinum) as determined from blood amino acid levels and amino acid supplementation studies in the rat.

    PubMed

    Khader, V; Rao, S V

    1982-01-01

    The limiting amino acids of Bengal gram (Cicer arietinum) were determined from plasma amino acid score and ratio and growth response of weanling rats to supplements of amino acids. The results indicated that methionine, threonine and tryptophan are the most limiting amino acids. Protein efficiency ratio of raw and cooked Bengal gram fed at a dietary level of 10% protein increased from 2.7 to 3.7 and 2.4 to 3.4, respectively, on supplementing the diets with methionine, threonine and tryptophan. Plasma levels of lysine, methionine, threonine and tryptophan were similar in rats fed raw or cooked Bengal gram, indicating that the trypsin or other inhibitors that may be present in the raw gram do not affect the biological availability of these amino acids.

  15. Molecular characterization of the level of sialic acids N-acetylneuraminic acid, N-glycolylneuraminic acid, and ketodeoxynonulosonic acid in porcine milk during lactation.

    PubMed

    Jahan, M; Wynn, P C; Wang, B

    2016-10-01

    Sialic acids (Sia) are key monosaccharide constituents of sialylated glycoproteins (Sia-GP), human sialylated milk oligosaccharide (Sia-MOS), and gangliosides. Human milk sialylated glycoconjugates (Sia-GC) are bioactive compounds known to act as prebiotics and promote neurodevelopment, immune function, and gut maturation in newborns. Only limited data are available on the Sia content of porcine milk. The objective of this study was to quantitatively determine the total level of Sia N-acetylneuraminic acid (Neu5Ac), N-glycolylneuraminic acid (Neu5Gc), and ketodeoxynonulosonic acid (KDN) in porcine milk and to compare these levels in gilt and sow milk during lactation. Milk from 8 gilts and 22 sows was collected at 3 stages of lactation (colostrum, transition, and mature milk). Standard and experimental samples were derivatized using 1,2-diamino-4,5-methylenedioxy-benzene and analyzed by ultra-high-performance liquid chromatography using a fluorescence detector. The following new findings are reported: (1) Gilt and sow milk contained significant levels of total Sia, with the highest concentration in colostrum (1,238.5 mg/L), followed by transition milk (778.3 mg/L) and mature milk (347.2 mg/L); (2) during lactation, the majority of Sia was conjugated to Sia-GP (41-46%), followed by Sia-MOS (31-42%) and a smaller proportion in gangliosides (12-28%); (3) Neu5Ac was the major form of Sia (93-96%), followed by Neu5Gc (3-6%) and then KDN (1-2%), irrespective of milk fraction or stage of lactation; (4) the concentration of Sia in Sia-GP and Sia-MOS showed a significant decline during lactation, but the level of ganglioside Sia remained relatively constant; (5) mature gilt milk contained a significantly higher concentration of Sia-GP than sow milk. The high concentration of total Sia in porcine milk suggests that Sia-GC are important nutrients that contribute to the optimization of neurodevelopment, immune function, and growth and development in piglets. These findings

  16. Two-Level Adaptive Algebraic Multigrid for a Sequence of Problems with Slowly Varying Random Coefficients [Adaptive Algebraic Multigrid for Sequence of Problems with Slowly Varying Random Coefficients

    SciTech Connect

    Kalchev, D.; Ketelsen, C.; Vassilevski, P. S.

    2013-11-07

    Our paper proposes an adaptive strategy for reusing a previously constructed coarse space by algebraic multigrid to construct a two-level solver for a problem with nearby characteristics. Furthermore, a main target application is the solution of the linear problems that appear throughout a sequence of Markov chain Monte Carlo simulations of subsurface flow with uncertain permeability field. We demonstrate the efficacy of the method with extensive set of numerical experiments.

  17. Draft Genome Sequence of a High-Level Colistin-Resistant Clinical Strain of the Enterobacter cloacae Complex

    PubMed Central

    Lin, Ji; Zhao, Feifei; Feng, Yu

    2017-01-01

    ABSTRACT Strain WCHECl-C4 of the Enterobacter cloacae complex, recovered from the blood of a patient with peritonitis, was high-level resistant to colistin. Here, we report its 5.1-Mb draft genome sequence, comprising 92 contigs with an average 55.74% G+C content. The genome contained 4,783 coding sequences and 68 tRNA genes. PMID:28385849

  18. Classifying nucleic acid sub-sequences as introns or exons using genetic programming

    SciTech Connect

    Handley, S.

    1995-12-31

    An evolutionary computation technique, genetic programming, created programs that classify messenger RNA sequences into one of two classes: (1) the sequence is expressed as (part of) a protein (an exon), or (2) not expressed as protein (an intron).

  19. 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.

  20. Clickable Nucleic Acids: Sequence-Controlled Periodic Copolymer/Oligomer Synthesis by Orthogonal Thiol-X Reactions.

    PubMed

    Xi, Weixian; Pattanayak, Sankha; Wang, Chen; Fairbanks, Benjamin; Gong, Tao; Wagner, Justine; Kloxin, Christopher J; Bowman, Christopher N

    2015-11-23

    Synthetic polymer approaches generally lack the ability to control the primary sequence, with sequence control referred to as the holy grail. Two click chemistry reactions were now combined to form nucleobase-containing sequence-controlled polymers in simple polymerization reactions. Two distinct approaches are used to form these click nucleic acid (CNA) polymers. These approaches employ thiol-ene and thiol-Michael reactions to form homopolymers of a single nucleobase (e.g., poly(A)n ) or homopolymers of specific repeating nucleobase sequences (e.g., poly(ATC)n). Furthermore, the incorporation of monofunctional thiol-terminated polymers into the polymerization system enables the preparation of multiblock copolymers in a single reaction vessel; the length of the diblock copolymer can be tuned by the stoichiometric ratio and/or the monomer functionality. These polymers are also used for organogel formation where complementary CNA-based polymers form reversible crosslinks.

  1. Blood folic acid, vitamin B12, and homocysteine levels in pregnant women with fetal growth restriction.

    PubMed

    Jiang, H L; Cao, L Q; Chen, H Y

    2016-12-19

    Deficiencies in nutrients such as folic acid and vitamin B12 may play a role in fetal growth restriction (FGR). However, whether folic acid, vitamin B12, or homocysteine is associated with FGR in Chinese populations remains unclear. This study investigated the relationship between these nutrient deficiencies and FGR in pregnant Chinese women. We selected 116 mother and infant pairs, and categorized the neonates into the FGR, appropriate for gestational age, and large for gestational age groups. Birth weight, body length, head circumference, body mass index (BMI), and Rohrer's body index of the newborns were measured. Serum folic acid, vitamin B12, and homocysteine levels were measured in mothers during the first three days of their hospital stay. Results showed that the FGR group exhibited reduced folic acid and vitamin B12 levels and elevated homocysteine levels than those in the other two groups. Folic acid and vitamin B12 levels were positively correlated with birth weight, head circumference, and BMI, whereas homocysteine level was negatively correlated with these variables. The FGR ratio in the folic acid and vitamin B12 deficiency group was higher than that in the sufficiency group (χ(2) = 4.717 and 4.437, P = 0.029 and 0.035, respectively). In addition, elevated homocysteine was associated with FGR (χ(2) = 5.366, P = 0.021). In conclusion, we found that folic acid and vitamin B12 deficiency was associated with elevated homocysteine levels, which may increase susceptibility to FGR.

  2. In vivo blockade of thalamic GABA(B) receptors increases excitatory amino-acid levels.

    PubMed

    Nyitrai, G; Emri, Z; Crunelli, V; Kékesi, K A; Dobolyi, A; Juhász, G

    1996-12-30

    The effect of intrathalamic application of GABA(B) receptor antagonists on the basal excitatory amino-acid levels was studied using microdialysis probes implanted in the dorsal lateral geniculate nucleus and in the ventrobasal complex. In both nuclei, continuous perfusion of the GABA(B) receptor antagonist 3-aminopropyl-(diethoxymethyl)-phosphinic acid (CGP 35348) produced an increase in the extracellular concentration of aspartate and (to a lesser extent) glutamate, but no change was observed in the level of taurine, the main amino acid involved in the regulation of brain osmolarity processes. In contrast, 3-amino-2-hydroxy-2-(4-chlorophenyl)-propanesulphonic acid (2-hydroxy-saclofen), another GABA(B) receptor antagonist, failed to affect the extracellular concentration of aspartate, glutamate and taurine. Thus, the basal level of excitatory amino acids in the thalamus in vivo is under the control of CGP 35348-sensitive GABA(B) receptors.

  3. Unusually high levels of n-6 polyunsaturated fatty acids in whale sharks and reef manta rays.

    PubMed

    Couturier, L I E; Rohner, C A; Richardson, A J; Pierce, S J; Marshall, A D; Jaine, F R A; Townsend, K A; Bennett, M B; Weeks, S J; Nichols, P D

    2013-10-01

    Fatty acid (FA) signature analysis has been increasingly used to assess dietary preferences and trophodynamics in marine animals. We investigated FA signatures of connective tissue of the whale shark Rhincodon typus and muscle tissue of the reef manta ray Manta alfredi. We found high levels of n-6 polyunsaturated fatty acids (PUFA), dominated by arachidonic acid (20:4n-6; 12-17 % of total FA), and comparatively lower levels of the essential n-3 PUFA-eicosapentaenoic acid (20:5n-3; ~1 %) and docosahexaenoic acid (22:6n-3; 3-10 %). Whale sharks and reef manta rays are regularly observed feeding on surface aggregations of coastal crustacean zooplankton during the day, which generally have FA profiles dominated by n-3 PUFA. The high levels of n-6 PUFA in both giant elasmobranchs raise new questions about the origin of their main food source.

  4. Effects of toxic work environments on sperm quality and ascorbic acid levels

    SciTech Connect

    Dawson, E.B.; Harris, W.A.; Powell, L.C. )

    1990-02-26

    Surveys have shown that toxic work environments lower sperm quality, and controlled studies indicate that ascorbic acid supplementation improves sperm viability and agglutination. The sperm quality of 50 subjects each from: (1) office workers, (2) a lead smelter, (3) petroleum refineries, and (4) a herbicide plant were compared with serum and semen ascorbic acid levels. The sperm characteristics studied were: count as million/ml and as percent; viability, motility, clumping, and abnormal morphology. The serum ascorbic acid levels were directly proportional to sperm viability and inversely correlated to clumping of all groups. Moreover, serum ascorbic acid levels were also inversely correlated to twin tail and amorphous forms of abnormal sperm morphology. The results of the study indicate that toxic environments depress sperm quality and suggest that ascorbic acid supplementation will improve sperm quality and fertility.

  5. Classification of protein motifs based on subcellular localization uncovers evolutionary relationships at both sequence and functional levels

    PubMed Central

    2013-01-01

    Background Most proteins have evolved in specific cellular compartments that limit their functions and potential interactions. On the other hand, motifs define amino acid arrangements conserved between protein family members and represent powerful tools for assigning function to protein sequences. The ideal motif would identify all members of a protein family but in practice many motifs identify both family members and unrelated proteins, referred to as True Positive (TP) and False Positive (FP) sequences, respectively. Results To address the relationship between protein motifs, protein function and cellular localization, we systematically assigned subcellular localization data to motif sequences from the comprehensive PROSITE sequence motif database. Using this data we analyzed relationships between localization and function. We find that TPs and FPs have a strong tendency to localize in different compartments. When multiple localizations are considered, TPs are usually distributed between related cellular compartments. We also identified cases where FPs are concentrated in particular subcellular regions, indicating possible functional or evolutionary relationships with TP sequences of the same motif. Conclusions Our findings suggest that the systematic examination of subcellular localization has the potential to uncover evolutionary and functional relationships between motif-containing sequences. We believe that this type of analysis complements existing motif annotations and could aid in their interpretation. Our results shed light on the evolution of cellular organelles and potentially establish the basis for new subcellular localization and function prediction algorithms. PMID:23865897

  6. Fe(II) oxidation during acid mine drainage neutralization in a pilot-scale Sequencing Batch Reactor.

    PubMed

    Zvimba, J N; Mathye, M; Vadapalli, V R K; Swanepoel, H; Bologo, L

    2013-01-01

    This study investigated Fe(II) oxidation during acid mine drainage (AMD) neutralization using CaCO3 in a pilot-scale Sequencing Batch Reactor (SBR) of hydraulic retention time (HRT) of 90 min and sludge retention time (SRT) of 360 min in the presence of air. The removal kinetics of Fe(II), of initial concentration 1,033 ± 0 mg/L, from AMD through oxidation to Fe(III) was observed to depend on both pH and suspended solids, resulting in Fe(II) levels of 679 ± 32, 242 ± 64, 46 ± 16 and 28 ± 0 mg/L recorded after cycles 1, 2, 3 and 4 respectively, with complete Fe(II) oxidation only achieved after complete neutralization of AMD. Generally, it takes 30 min to completely oxidize Fe(II) during cycle 4, suggesting that further optimization of SBR operation based on both pH and suspended solids manipulation can result in significant reduction of the number of cycles required to achieve acceptable Fe(II) oxidation for removal as ferric hydroxide. Overall, complete removal of Fe(II) during AMD neutralization is attractive as it promotes recovery of better quality waste gypsum, key to downstream gypsum beneficiation for recovery of valuables, thereby enabling some treatment-cost recovery and prevention of environmental pollution from dumping of sludge into landfills.

  7. 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.

  8. Differentiation of acetic acid bacteria based on sequence analysis of 16S-23S rRNA gene internal transcribed spacer sequences.

    PubMed

    González, Angel; Mas, Albert

    2011-06-30

    The 16S-23S gene internal transcribed spacer sequence of sixty-four strains belonging to different acetic acid bacteria genera were analyzed, and phylogenetic trees were generated for each genera. The topologies of the different trees were in accordance with the 16S rRNA gene trees, although the similarity percentages obtained between the species was shown to be much lower. These values suggest the usefulness of including the 16S-23S gene internal transcribed spacer region as a part of the polyphasic approach required for the further classification of acetic acid bacteria. Furthermore, the region could be a good target for primer and probe design. It has also been validated for use in the identification of unknown samples of this bacterial group from wine vinegar and fruit condiments.

  9. A rapid method for manual or automated purification of fluorescently labeled nucleic acids for sequencing, genotyping, and microarrays.

    PubMed

    Springer, Amy L; Booth, Lisa R; Braid, Michael D; Houde, Christiane M; Hughes, Karin A; Kaiser, Robert J; Pedrak, Casandra; Spicer, Douglas A; Stolyar, Sergey

    2003-03-01

    Fluorescent dyes provide specific, sensitive, and multiplexed detection of nucleic acids. To maximize sensitivity, fluorescently labeled reaction products (e.g., cycle sequencing or primer extension products) must be purified away from residual dye-labeled precursors. Successful high-throughput analyses require that this purification be reliable, rapid, and amenable to automation. Common methods for purifying reaction products involve several steps and require processes that are not easily automated. Prolinx, Inc. has devel oped RapXtract superparamagnetic separation technology affording rapid and easy-to-perform methods that yield high-quality product and are easily automated. The technology uses superparamagnetic particles that specifically remove unincorporated dye-labeled precursors. These particles are efficiently pelleted in the presence of a magnetic field, making them ideal for purification because of the rapid separations that they allow. RapXtract-purified sequencing reactions yield data with good signal and high Phred quality scores, and they work with various sequencing dye chemistries, including BigDye and near-infrared fluorescence IRDyes. RapXtract technology can also be used to purify dye primer sequencing reactions, primer extension reactions for genotyping analysis, and nucleic acid labeling reactions for microarray hybridization. The ease of use and versatility of RapXtract technology makes it a good choice for manual or automated purification of fluorescently labeled nucleic acids.

  10. Gene structure and amino acid sequence of Latimeria chalumnae (coelacanth) myelin DM20: phylogenetic relation of the fish.

    PubMed

    Tohyama, Y; Kasama-Yoshida, H; Sakuma, M; Kobayashi, Y; Cao, Y; Hasegawa, M; Kojima, H; Tamai, Y; Tanokura, M; Kurihara, T

    1999-07-01

    The structure of Latimeria chalumnae (coelacanth) proteolipid protein/DM20 gene excluding exon 1 was determined, and the amino acid sequence of Latimeria DM20 corresponding to exons 2-7 was deduced. The nucleotide sequence of exon 3 suggests that only DM20 isoform is expressed in Latimeria. The structure of proteolipid protein/DM20 gene is well preserved among human, dog, mouse, and Latimeria. Southern blot analysis indicates that Latimeria DM20 gene is a single-copy gene. When the amino acid sequences of DM20 were compared among various species, Latimeria was more similar to tetrapods than other fishes including lungfish, confirming the previous finding by immunoreactivity (Waehneldt and Malotka 1989 J. Neurochem. 52:1941-1943). However, when phylogenetic trees were constructed from the DM20 sequences, lungfish was clearly the closest to tetrapods. Latimeria was situated outside of lungfish by the maximum likelihood method. The apparent similarity of Latimeria DM20 to tetrapod proteolipid protein/DM20 is explained by the slow amino acid substitution rate of Latimeria DM20.

  11. Pyrazinoic acid decreases the proton motive force, respiratory ATP synthesis activity, and cellular ATP levels.

    PubMed

    Lu, Ping; Haagsma, Anna C; Pham, Hoang; Maaskant, Janneke J; Mol, Selena; Lill, Holger; Bald, Dirk

    2011-11-01

    Pyrazinoic acid, the active form of the first-line antituberculosis drug pyrazinamide, decreased the proton motive force and respiratory ATP synthesis rates in subcellular mycobacterial membrane assays. Pyrazinoic acid also significantly lowered cellular ATP levels in Mycobacterium bovis BCG. These results indicate that the predominant mechanism of killing by this drug may operate by depletion of cellular ATP reserves.

  12. [Effect of ribosome binding site sequence on the expression level of HBcAg in E. coli].

    PubMed

    Li, M; Ma, X

    1992-01-01

    The recombinant (pKL series) Plasmids showing different levels of HBcAg antigenicity in ELISA were constructed by inserting the HBcAg gene which was randomly deleted at the non-coding region by Bal-31 exonuclease into plasmid vector pKK223-3 containing tac promoter and SD sequence. SDS-PAGE and Western blot experiments indicated that molecular weight of the HBcAg protein generated by these positive clones was 21000D. Three plasmids with high, moderate and low HBcAg expression level were sequenced and the distance between SD sequence and HBcAg gene ATG codon was 12, 13 and 19bp respectively. Computer analysis of secondary structure of the ribosome binding sites on RNA transcripts also revealed energy and structure differences between the low and high level expression plasmids, suggesting the importance of this distance and the mRNA structure to gene expression.

  13. Cloning and sequencing of the Bet v 1-homologous allergen Fra a 1 in strawberry (Fragaria ananassa) shows the presence of an intron and little variability in amino acid sequence.

    PubMed

    Musidlowska-Persson, Anna; Alm, Rikard; Emanuelsson, Cecilia

    2007-02-01

    The Fra a 1 allergen in strawberry (Fragaria ananassa) is homologous to the major birch pollen allergen Bet v 1, which has numerous isoforms differing in terms of amino acid sequence and immunological impact. To map the extent of sequence differences in the Fra a 1 allergen, PCR cloning and sequencing was applied. Several genomic sequences of Fra a 1, with a length of either 584, 591 or 594 nucleotides, were obtained from three different strawberry varieties. All contained one intron, with the length of either 101 or 110 nucleotides. By sequencing 30 different clones, eight different DNA sequences were obtained, giving in total five potential Fra a 1 protein isoforms, with high sequence similarity (>97% sequence identity) and only seven positions of amino acid variability, which were largely confirmed by mass spectrometry of expressed proteins. We conclude that the sequence variability in the strawberry allergen Fra a 1 is small, within and between strawberry varieties, and that multiple spots, previously detected in 2DE, are presumably due to differences in post-translational modification rather than differences in amino acid sequence. The most abundant Fra a 1 isoform sequence, recombinantly expressed in Escherichia coli after removal of the intron, was recognized by IgE from strawberry allergic patients. It cross-reacted with antibodies to Bet v 1 and the homologous apple allergen Mal d 1 (61 and 78% sequence identity, respectively), and will be used in further analyses of variation in Fra a 1-expression.

  14. Purification of a marsupial insulin: amino-acid sequence of insulin from the eastern grey kangaroo Macropus giganteus.

    PubMed

    Treacy, G B; Shaw, D C; Griffiths, M E; Jeffrey, P D

    1989-03-24

    Insulin has been purified from kangaroo pancreas by acidic ethanol extraction, diethyl ether precipitation and gel filtration. The amino-acid sequence of this, the first marsupial insulin to be studied, is reported. It differs from human insulin by only four amino-acid substitutions, all in regions of the molecule previously known to be variable. However, it should be noted that one of these, asparagine for threonine at A8, has not been reported before. Computer comparisons of all 43 insulin sequences reported to date with kangaroo insulin show it to be most closely related to a group of mammalian insulins (dog, pig, cow, human) known to be of high biological potency. The measurement of blood glucose lowering in the rabbit by kangaroo insulin is consistent with this conclusion. Comparisons of amino-acid sequences of other proteins with their kangaroo counterparts show a greater difference, in line with the time of divergence of marsupials. The limited differences observed in insulin and cytochrome c suggest that their structures need to be closely conserved in order to maintain function.

  15. [A retrospective study analysis of urinary hippuric acid levels in occupational toxicology exams].

    PubMed

    Gonzalez, Kelly Cristina; Sagebin, Fernando Rodrigues; Oliveira, Paola Garcia; Glock, Luiz; Thiesen, Flavia Valladão

    2010-06-01

    Hippuric acid is the primary metabolite of toluene, a solvent widely used in industrial processes with considerable toxic effects, a fact which justifies regularly monitoring individuals with occupational exposure to this solvent. This work aims at evaluating urinary hippuric acid levels found in workers subject to biological monitoring. A retrospective study was carried out with data referring from 2002 to 2005, in which exams results and employment status were analyzed (periodic, post-employment, and pre-employment exams). Results indicate a significant reduction in hippuric acid levels for 2005. Periodic exams presented higher results than pre-employment and post-employment exams. No significant difference was found in individuals grouped according to their status in each of the established intervals, their reference numbers, and maximum biological levels allowed. Hippuric acid levels detected indicate low risk of toluene exposure for the population under evaluation, probably due to a growing concern with the deployment of measures regarding occupational hygiene.

  16. K-Pax2: Bayesian identification of cluster-defining amino acid positions in large sequence datasets

    PubMed Central

    Grad, Yonatan; Cobey, Sarah; Puranen, Juha Santeri; Corander, Jukka

    2015-01-01

    The recent growth in publicly available sequence data has introduced new opportunities for studying microbial evolution and spread. Because the pace of sequence accumulation tends to exceed the pace of experimental studies of protein function and the roles of individual amino acids, statistical tools to identify meaningful patterns in protein diversity are essential. Large sequence alignments from fast-evolving micro-organisms are particularly challenging to dissect using standard tools from phylogenetics and multivariate statistics because biologically relevant functional signals are easily masked by neutral variation and noise. To meet this need, a novel computational method is introduced that is easily executed in parallel using a cluster environment and can handle thousands of sequences with minimal subjective input from the user. The usefulness of this kind of machine learning is demonstrated by applying it to nearly 5000 haemagglutinin sequences of influenza A/H3N2.Antigenic and 3D structural mapping of the results show that the method can recover the major jumps in antigenic phenotype that occurred between 1968 and 2013 and identify specific amino acids associated with these changes. The method is expected to provide a useful tool to uncover patterns of protein evolution. PMID:28348810

  17. The amino acid sequence of Ole e I, the major allergen from olive tree (Olea europaea) pollen.

    PubMed

    Villalba, M; Batanero, E; López-Otín, C; Sánchez, L M; Monsalve, R I; González de la Peña, M A; Lahoz, C; Rodríguez, R

    1993-09-15

    The complete primary structure of the major allergen from Olea europaea (olive tree) pollen, Ole e I (IUIS nomenclature), has been determined. The amino acid sequence was established by automated Edman degradation of the reduced and alkylated molecule as well as of selected fragments obtained by proteolytic digestions. Ole e I contains a single polypeptide chain of 145 amino acid residues with a calculated molecular mass of 16331 Da. No free sulfhydryl groups have been detected in the native protein. The molecule contains a putative glycosylation site. A high degree of microheterogeneity has been observed, mainly centered in the first 33% of the molecule. Comparison of Ole e I sequence with protein sequence databases showed no similarity with other known allergens. However, it has a 36% and 38% sequence identity with the putative polypeptide structures, deduced, respectively, from nucleotide sequences of genes isolated from tomato anthers and corn pollen, which have been suggested to be involved in the growing of the pollen tube. Therefore, the olive tree allergen may be a constitutive protein of the pollen involved in reproductive functions.

  18. Complete genome sequence of Enterococcus mundtii QU 25, an efficient L-(+)-lactic acid-producing bacterium.

    PubMed

    Shiwa, Yuh; Yanase, Hiroaki; Hirose, Yuu; Satomi, Shohei; Araya-Kojima, Tomoko; Watanabe, Satoru; Zendo, Takeshi; Chibazakura, Taku; Shimizu-Kadota, Mariko; Yoshikawa, Hirofumi; Sonomoto, Kenji

    2014-08-01

    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.

  19. 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.

  20. Isolation and a partial amino acid sequence of insulin from the islet tissue of cod (Gadus callarias)

    PubMed Central

    Grant, P. T.; Reid, K. B. M.

    1968-01-01

    1. Insulin has been isolated by gel filtration and ion-exchange chromatography from extracts of the discrete islet tissue of cod. The final preparation yielded a single band on electrophoresis at two pH values. The biological potency was 11·5 international units/mg. in mouse-convulsion and other assay procedures. 2. Glycine and methionine were shown to be the N-terminal amino acids of the A and B chains respectively. An estimate of the molecular weight together with amino acid analyses indicated that cod insulin, like the bovine hormone, consists of 51 amino acid residues. In contrast, the amino acid composition differs markedly from bovine insulin. 3. Oxidation of insulin with performic acid yielded the A and B peptide chains, which were separated by ion-exchange chromatography. Sequence studies on smaller peptides isolated from enzymic digests or from dilute acetic acid hydrolysates of the two chains have established the sequential order of 14 of the 21 amino acid residues of the A chain and 25 of the 30 amino acid residues of the B chain. PMID:4866431

  1. Oxygen affinity and amino acid sequence of myoglobins from endothermic and ectothermic fish.

    PubMed

    Marcinek, D J; Bonaventura, J; Wittenberg, J B; Block, B A

    2001-04-01

    Myoglobin (Mb) buffers intracellular O2 and facilitates diffusion of O2 through the cell. These functions of Mb will be most effective when intracellular PO2 is near the partial pressure of oxygen at which Mb is half saturated (P50) of the molecule. We test the hypothesis that Mb oxygen affinity has evolved such that it is conserved when adjusted for body temperature among closely related animals. We measure oxygen P50s tonometrically and oxygen dissociation rate constants with stopped flow and generate amino acid sequence from cDNA of Mbs from fish with different body temperatures. P50s for the endothermic bluefin tuna, skipjack tuna, and blue marlin at 20 degrees C were 0.62 +/- 0.02, 0.59 +/- 0.01, 0.58 +/- 0.04 mmHg, respectively, and were significantly lower than those for ectothermic bonito (1.03 +/- 0.07 mmHg) and mackerel (1.39 +/- 0.03 mmHg). Because the oxygen affinity of Mb decreases with increasing temperature, the above differences in oxygen affinity between endothermic and ectothermic fish are reduced when adjusted for the in vivo muscle temperature of the animal. Oxygen dissociation rate constants at 20 degrees C for the endothermic species ranged from 34.1 to 49.3 s(-1), whereas those for mackerel and bonito were 102 and 62 s(-1), respectively. Correlated with the low oxygen affinity and fast dissociation kinetics of mackerel Mb is a substitution of alanine for proline that would likely result in a more flexible mackerel protein.

  2. Molecular cloning, nucleotide sequence, and abscisic acid induction of a suberization-associated highly anionic peroxidase.

    PubMed

    Roberts, E; Kolattukudy, P E

    1989-06-01

    A highly anionic peroxidase induced in suberizing cells was suggested to be the key enzyme involved in polymerization of phenolic monomers to generate the aromatic matrix of suberin. The enzyme encoded by a potato cDNA was found to be highly homologous to the anionic peroxidase induced in suberizing tomato fruit. A tomato genomic library was screened using the potato anionic peroxidase cDNA and one genomic clone was isolated that contained two tandemly oriented anionic peroxidase genes. These genes were sequenced and were 96% and 87% identical to the mRNA for potato anionic peroxidase. Both genes consist of three exons with the relative positions of their two introns being conserved between the two genes. Primer extension analysis showed that only one of the genes is expressed in the periderm of 3 day wound-healed tomato fruits. Southern blot analyses suggested that there are two copies each of the two highly homologous genes per haploid genome in both potato and tomato. Abscisic acid (ABA) induced the accumulation of the anionic peroxidase transcripts in potato and tomato callus tissues. Northern blots showed that peroxidase mRNA was detectable at 2 days and was maximal at 8 days after transfer of potato callus to solid agar media containing 10(-4) M ABA. The transcripts induced by ABA in both potato and tomato callus were identical in size to those induced in wound-healing potato tuber and tomato fruit. The anionic peroxidase peptide was detected in extracts of potato callus grown on the ABA-containing media by western blot analysis. The results support the suggestion that stimulation of suberization by ABA involves the induction of the highly anionic peroxidase.

  3. Complete genome sequence of the probiotic lactic acid bacterium Lactobacillus acidophilus NCFM

    PubMed Central

    Altermann, Eric; Russell, W. Michael; Azcarate-Peril, M. Andrea; Barrangou, Rodolphe; Buck, B. Logan; McAuliffe, Olivia; Souther, Nicole; Dobson, Alleson; Duong, Tri; Callanan, Michael; Lick, Sonja; Hamrick, Alice; Cano, Raul; Klaenhammer, Todd R.

    2005-01-01

    Lactobacillus acidophilus NCFM is a probiotic bacterium that has been produced commercially since 1972. The complete genome is 1,993,564 nt and devoid of plasmids. The average GC content is 34.71% with 1,864 predicted ORFs, of which 72.5% were functionally classified. Nine phage-related integrases were predicted, but no complete prophages were found. However, three unique regions designated as potential autonomous units (PAUs) were identified. These units resemble a unique structure and bear characteristics of both plasmids and phages. Analysis of the three PAUs revealed the presence of two R/M systems and a prophage maintenance system killer protein. A spacers interspersed direct repeat locus containing 32 nearly perfect 29-bp repeats was discovered and may provide a unique molecular signature for this organism. In silico analyses predicted 17 transposase genes and a chromosomal locus for lactacin B, a class II bacteriocin. Several mucus- and fibronectin-binding proteins, implicated in adhesion to human intestinal cells, were also identified. Gene clusters for transport of a diverse group of carbohydrates, including fructooligosaccharides and raffinose, were present and often accompanied by transcriptional regulators of the lacI family. For protein degradation and peptide utilization, the organism encoded 20 putative peptidases, homologs for PrtP and PrtM, and two complete oligopeptide transport systems. Nine two-component regulatory systems were predicted, some associated with determinants implicated in bacteriocin production and acid tolerance. Collectively, these features within the genome sequence of L. acidophilus are likely to contribute to the organisms' gastric survival and promote interactions with the intestinal mucosa and microbiota. PMID:15671160

  4. Coumarin effects on amino acid levels in mice prefrontal cortex and hippocampus.

    PubMed

    Pereira, Elaine Cristina; Lucetti, Daniel Luna; Barbosa-Filho, José Maria; de Brito, Eliane Magalhães; Monteiro, Valdécio Silvano; Patrocínio, Manoel Cláudio Azevedo; de Moura, Rebeca Ribeiro; Leal, Luzia Kalyne Almeida Moreira; Macedo, Danielle Silveira; de Sousa, Francisca Cléa Florenço; de Barros Viana, Glauce Socorro; Vasconcelos, Silvânia Maria Mendes

    2009-04-24

    Coumarin is a compound known to be present in a wide variety of plants, microorganisms and animal species. Most of its effects were studied in organs and systems other than the central nervous system. The present work evaluated the effect of coumarin administration on the levels of gamma-aminobutyric acid (GABA), glutamate (GLU), glycine (GLY) and taurine (TAU) in the prefrontal cortex and hippocampus of mice. Male Swiss mice were treated with distilled water (controls), coumarin (20 or 40 mg/kg, i.p.) or diazepam (1 mg/kg, i.p.). Results showed that in the prefrontal cortex, coumarin at the lowest dose increased the levels of GLU and TAU, while GABA increased with both doses studied and GLY had its levels increased only at the dose of 40 mg/kg. Diazepam (DZP) increased the levels of GABA and TAU and decreased the levels of GLU and GLY in this area. In the hippocampus, only glutamate had its levels decreased after coumarin treatment, while diazepam increased the levels of GABA and TAU and decreased the levels of GLU in this brain region. We concluded that coumarin stimulates the release of endogenous amino acids, increasing the levels of inhibitory and excitatory amino acids in the prefrontal cortex, and decreasing glutamate levels in the hippocampus. Together, these results are of interest, considering that some neurodegenerative diseases and seizures are related to the imbalance of the amino acid levels in the CNS suggesting a perspective of a therapeutic use of coumarins in these disorders.

  5. Complete nucleotide and derived amino acid sequence of cDNA encoding the mitochondrial uncoupling protein of rat brown adipose tissue: lack of a mitochondrial targeting presequence.

    PubMed Central

    Ridley, R G; Patel, H V; Gerber, G E; Morton, R C; Freeman, K B

    1986-01-01

    A cDNA clone spanning the entire amino acid sequence of the nuclear-encoded uncoupling protein of rat brown adipose tissue mitochondria has been isolated and sequenced. With the exception of the N-terminal methionine the deduced N-terminus of the newly synthesized uncoupling protein is identical to the N-terminal 30 amino acids of the native uncoupling protein as determined by protein sequencing. This proves that the protein contains no N-terminal mitochondrial targeting prepiece and that a targeting region must reside within the amino acid sequence of the mature protein. Images PMID:3012461

  6. Peptide Mass Fingerprinting and N-Terminal Amino Acid Sequencing of Glycosylated Cysteine Protease of Euphorbia nivulia Buch.-Ham.

    PubMed Central

    Badgujar, Shamkant B.; Mahajan, Raghunath T.

    2013-01-01

    A new cysteine protease named Nivulian-II has been purified from the latex of Euphorbia nivulia Buch.-Ham. The apparent molecular mass of Nivulian-II is 43670.846 Da (MALDI TOF/MS). Peptide mass fingerprint analysis revealed peptide matches to Maturase K (Q52ZV1_9MAGN) of Banksia quercifolia. The N-terminal sequence (DFPPNTCCCICC) showed partial homology with those of other cysteine proteinases of biological origin. This is the first paper to characterize a Nivulian-II of E. nivulia latex with respect to amino acid sequencing. PMID:23476742

  7. Comparison of Theoretical and Experimental Level Values of the n = 2 Configurations in the Nitrogen Isoelectronic Sequence

    NASA Astrophysics Data System (ADS)

    Edlén, Bengt

    1984-08-01

    Observed level intervals within and between the configurations 2s2 2p3, 2s2p4 and 2p5 of the nitrogen sequence are compared with the corresponding theoretical values derived from the tables of Cheng, Kim and Desclaux. The differences are expressed by suitable functions of Z which give the systematic correction to the theoretical values and reveal the accidental errors in the experimental values. The technique is the same as was used in previous examinations of the Li, Be, B, O and F sequences. The results are summarized in tables of recommended level values (Z = 10-36) and recalculated wavelengths.

  8. Analysis of methylphosphonic acid, ethyl methylphosphonic acid and isopropyl methylphosphonic acid at low microgram per liter levels in groundwater.

    PubMed

    Sega, G A; Tomkins, B A; Griest, W H

    1997-11-28

    A method is described for determining methylphosphonic acid, ethyl methylphosphonic acid and isopropyl methylphosphonic acid, which are hydrolysis products of the nerve agents VX (S-2-diisopropylaminoethyl O-ethyl methylphosphonothiolate) and GB (sarin, isopropylmethyl phosphonofluoridate). The analytes are extracted from 50 ml groundwater using a solid-phase extraction column packed with 500 mg of silica with a bonded quaternary amine phase, and are eluted and derivatized with methanolic trimethylphenylammonium hydroxide. Separation and quantitation are achieved using a capillary column gas chromatograph equipped with a flame photometric detector operated in its phosphorus-selective mode. Two independent statistically-unbiased procedures were employed to determine the detection limits, which ranged between 3 and 9 micrograms/l, for the three analytes.

  9. High-level exogenous glutamic acid-independent production of poly-(γ-glutamic acid) with organic acid addition in a new isolated Bacillus subtilis C10.

    PubMed

    Zhang, Huili; Zhu, Jianzhong; Zhu, Xiangcheng; Cai, Jin; Zhang, Anyi; Hong, Yizhi; Huang, Jin; Huang, Lei; Xu, Zhinan

    2012-07-01

    A new exogenous glutamic acid-independent γ-PGA producing strain was isolated and characterized as Bacillus subtilis C10. The factors influencing the endogenous glutamic acid supply and the biosynthesis of γ-PGA in this strain were investigated. The results indicated that citric acid and oxalic acid showed the significant capability to support the overproduction of γ-PGA. This stimulated increase of γ-PGA biosynthesis by citric acid or oxalic acid was further proved in the 10 L fermentor. To understand the possible mechanism contributing to the improved γ-PGA production, the activities of four key intracellular enzymes were measured, and the possible carbon fluxes were proposed. The result indicated that the enhanced level of pyruvate dehydrogenase (PDH) activity caused by oxalic acid was important for glutamic acid synthesized de novo from glucose. Moreover, isocitrate dehydrogenase (ICDH) and glutamate dehydrogenase (GDH) were the positive regulators of glutamic acid biosynthesis, while 2-oxoglutarate dehydrogenase complex (ODHC) was the negative one.

  10. 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

  11. The genome sequence of the highly acetic acid-tolerant Zygosaccharomyces bailii-derived interspecies hybrid strain ISA1307, isolated from a sparkling wine plant.

    PubMed

    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

    2014-06-01

    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.

  12. Amino acid limitation induces down-regulation of WNT5a at transcriptional level

    SciTech Connect

    Wang Zuguang; Chen Hong

    2009-01-23

    An aberrant WNT signaling contributes to the development and progression of multiple cancers. WNT5a is one of the WNT signaling molecules. This study was designed to test the hypothesis that amino acid deprivation induces changes in the WNT signaling pathway in colon cancer cells. Results showed that targets of the amino acid response pathway, ATF3 and p21, were induced in the human colon cancer cell line SW480 during amino acid limitation. There was a significant decrease in the WNT5a mRNA level following amino acid deprivation. The down-regulation of WNT5a mRNA by amino acid deprivation is not due to mRNA destabilization. There is a reduction of nuclear {beta}-catenin protein level by amino acid limitation. Under amino acid limitation, phosphorylation of ERK1/2 was increased and the blockage of ERK1/2 by the inhibitor U0126 partially restored WNT5a mRNA level. In conclusion, amino acid limitation in colon cancer cells induces phosphorylation of ERK1/2, which then down-regulates WNT5a expression.

  13. Solvent extraction in the treatment of acidic high-level liquid waste : where do we stand?

    SciTech Connect

    Horwitz, E. P.; Schulz, W. W.

    1998-06-18

    During the last 15 years, a number of solvent extraction/recovery processes have been developed for the removal of the transuranic elements, {sup 90}Sr and {sup 137}Cs from acidic high-level liquid waste. These processes are based on the use of a variety of both acidic and neutral extractants. This chapter will present an overview and analysis of the various extractants and flowsheets developed to treat acidic high-level liquid waste streams. The advantages and disadvantages of each extractant along with comparisons of the individual systems are discussed.

  14. 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.

  15. Evaluation of circulating levels and renal clearance of natural amino acids in patients with Cushing's disease.

    PubMed

    Faggiano, A; Pivonello, R; Melis, D; Alfieri, R; Filippella, M; Spagnuolo, G; Salvatore, F; Lombardi, G; Colao, A

    2002-02-01

    Although the hypercortisolism-induced impairment of protein homeostasis is object of several studies, a detailed evaluation of the complete amino acid profile of patients with Cushing's syndrome (CS) has never been performed. The aim of the current open transversal controlled study was to evaluate serum and urinary concentrations as well as renal clearance of the complete series of natural amino acids and their relationship with glucose tolerance in patients with Cushing's disease (CD). Twenty patients with CD (10 active and 10 cured) and 20 sex- and age-matched healthy controls entered the study. Measurement of serum and urinary levels of the complete series of natural amino acids was performed in all patients analyzed by cationic exchange high performance liquid cromatography (HPLC) after 2 weeks of a standardized protein intake regimen. The renal clearance (renal excretion rate) of each amino acid was calculated on the basis of the serum and urinary concentrations of creatinine and the specific amino acid. Fasting glucose and insulin levels, glucose and insulin response to standard glucose load, insulinogenic and homeostasis model insulin resistance (Homa-R) indexes were also evaluated and correlated to the circulating levels and renal clearances of each amino acid. Significantly higher serum (p<0.01) and urinary (p<0.05) levels of alanine and cystine, lower serum and higher urinary levels of leucine, isoleucine and valine (p<0.05) and higher renal excretion rates of leucine, isoleucine and valine (p<0.01) were found in patients with active CD than in patients cured from the disease and in controls. No difference was found between cured patients and controls. Creatinine clearance was similar in active and cured patients and in controls. In patients with active CD, urinary cortisol levels were significantly correlated to urinary cystine levels (r=0.85; p<0.01) and renal excretion rate of leucine (r=-0.76; p<0.05), isoleucine (r=-0.76; p<0.05) and valine (r=-0

  16. Genetic loci associated with circulating levels of very long-chain saturated fatty acids[S

    PubMed Central

    Lemaitre, Rozenn N.; King, Irena B.; Kabagambe, Edmond K.; Wu, Jason H. Y.; McKnight, Barbara; Manichaikul, Ani; Guan, Weihua; Sun, Qi; Chasman, Daniel I.; Foy, Millennia; Wang, Lu; Zhu, Jingwen; Siscovick, David S.; Tsai, Michael Y.; Arnett, Donna K.; Psaty, Bruce M.; Djousse, Luc; Chen, Yii-Der I.; Tang, Weihong; Weng, Lu-Chen; Wu, Hongyu; Jensen, Majken K.; Chu, Audrey Y.; Jacobs, David R.; Rich, Stephen S.; Mozaffarian, Dariush; Steffen, Lyn; Rimm, Eric B.; Hu, Frank B.; Ridker, Paul M.; Fornage, Myriam; Friedlander, Yechiel

    2015-01-01

    Very long-chain saturated fatty acids (VLSFAs) are saturated fatty acids with 20 or more carbons. In contrast to the more abundant saturated fatty acids, such as palmitic acid, there is growing evidence that circulating VLSFAs may have beneficial biological properties. Whether genetic factors influence circulating levels of VLSFAs is not known. We investigated the association of common genetic variation with plasma phospholipid/erythrocyte levels of three VLSFAs by performing genome-wide association studies in seven population-based cohorts comprising 10,129 subjects of European ancestry. We observed associations of circulating VLSFA concentrations with common variants in two genes, serine palmitoyl-transferase long-chain base subunit 3 (SPTLC3), a gene involved in the rate-limiting step of de novo sphingolipid synthesis, and ceramide synthase 4 (CERS4). The SPTLC3 variant at rs680379 was associated with higher arachidic acid (20:0 , P = 5.81 × 10−13). The CERS4 variant at rs2100944 was associated with higher levels of 20:0 (P = 2.65 × 10−40) and in analyses that adjusted for 20:0, with lower levels of behenic acid (P = 4.22 × 10−26) and lignoceric acid (P = 3.20 × 10−21). These novel associations suggest an inter-relationship of circulating VLSFAs and sphingolipid synthesis. PMID:25378659

  17. Metformin impairs systemic bile acid homeostasis through regulating SIRT1 protein levels.

    PubMed

    Chen, Qi; Yang, Xiaoying; Zhang, Huabing; Kong, Xingxing; Yao, Lu; Cui, Xiaona; Zou, Yongkang; Fang, Fude; Yang, Jichun; Chang, Yongsheng

    2017-01-01

    Metformin is widely used to treat hyperglycemia. However, metformin treatment may induce intrahepatic cholestasis and liver injury in a few patients with type II diabetes through an unknown mechanism. Here we show that metformin decreases SIRT1 protein levels in primary hepatocytes and liver. Both metformin-treated wild-type C57 mice and hepatic SIRT1-mutant mice had increased hepatic and serum bile acid levels. However, metformin failed to change systemic bile acid levels in hepatic SIRT1-mutant mice. Molecular mechanism study indicates that SIRT1 directly interacts with and deacetylates Foxa2 to inhibit its transcriptional activity on expression of genes involved in bile acids synthesis and transport. Hepatic SIRT1 mutation elevates Foxa2 acetylation levels, which promotes Foxa2 binding to and activating genes involved in bile acids metabolism, impairing hepatic and systemic bile acid homeostasis. Our data clearly suggest that hepatic SIRT1 mediates metformin effects on systemic bile acid metabolism and modulation of SIRT1 activity in liver may be an attractive approach for treatment of bile acid-related diseases such as cholestasis.

  18. Genetic loci associated with circulating levels of very long-chain saturated fatty acids.

    PubMed

    Lemaitre, Rozenn N; King, Irena B; Kabagambe, Edmond K; Wu, Jason H Y; McKnight, Barbara; Manichaikul, Ani; Guan, Weihua; Sun, Qi; Chasman, Daniel I; Foy, Millennia; Wang, Lu; Zhu, Jingwen; Siscovick, David S; Tsai, Michael Y; Arnett, Donna K; Psaty, Bruce M; Djousse, Luc; Chen, Yii-Der I; Tang, Weihong; Weng, Lu-Chen; Wu, Hongyu; Jensen, Majken K; Chu, Audrey Y; Jacobs, David R; Rich, Stephen S; Mozaffarian, Dariush; Steffen, Lyn; Rimm, Eric B; Hu, Frank B; Ridker, Paul M; Fornage, Myriam; Friedlander, Yechiel

    2015-01-01

    Very long-chain saturated fatty acids (VLSFAs) are saturated fatty acids with 20 or more carbons. In contrast to the more abundant saturated fatty acids, such as palmitic acid, there is growing evidence that circulating VLSFAs may have beneficial biological properties. Whether genetic factors influence circulating levels of VLSFAs is not known. We investigated the association of common genetic variation with plasma phospholipid/erythrocyte levels of three VLSFAs by performing genome-wide association studies in seven population-based cohorts comprising 10,129 subjects of European ancestry. We observed associations of circulating VLSFA concentrations with common variants in two genes, serine palmitoyl-transferase long-chain base subunit 3 (SPTLC3), a gene involved in the rate-limiting step of de novo sphingolipid synthesis, and ceramide synthase 4 (CERS4). The SPTLC3 variant at rs680379 was associated with higher arachidic acid (20:0 , P = 5.81 × 10(-13)). The CERS4 variant at rs2100944 was associated with higher levels of 20:0 (P = 2.65 × 10(-40)) and in analyses that adjusted for 20:0, with lower levels of behenic acid (P = 4.22 × 10(-26)) and lignoceric acid (P = 3.20 × 10(-21)). These novel associations suggest an inter-relationship of circulating VLSFAs and sphingolipid synthesis.

  19. Snake venoms. The amino-acid sequence of protein S5C4 from Dendroaspis jamesoni kaimosae (Jameson's mamba) venom.

    PubMed

    Joubert, F J; Strydom, A J; Taljaard, N

    1978-06-01

    A major component (S5C4) was purified from Jameson's mamba by gel filtration on Sephadex G-50 and by ion-exchange chromotography on CM-cellulose. Protein S5C4 contains 60 amino acid residues and is cross-linked by four intrachain disulphide bridges. The complete primary structure of the protein has been elucidated. The toxicities, the immunochemical properties, the sequence and the invariant amino acid residues of protein S5C4 resemble subgroup II of the angusticeps-type proteins.

  20. 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 ...

  1. High-Throughput Multilocus Sequence Typing: Bringing Molecular Typing to the Next Level

    PubMed Central

    Boers, Stefan A.; van der Reijden, Wil A.; Jansen, Ruud

    2012-01-01

    Multilocus sequence typing (MLST) is a widely used system for typing microorganisms by sequence analysis of housekeeping genes. The main advantage of MLST in comparison to other typing techniques is the unambiguity and transferability of sequence data. However, a main disadvantage is the high cost of DNA sequencing. Here we introduce a high-throughput MLST (HiMLST) method that employs next-generation sequencing (NGS) technology (Roche 454), to generate large quantities of high-quality MLST data at low costs. The HiMLST protocol consists of two steps. In the first step MLST target genes are amplified by PCR in multi-well plates. During this PCR the amplicons of each bacterial isolate are provided with a unique DNA barcode, the multiplex identifier (MID). In the second step all amplicons are pooled and sequenced in a single NGS-run. The MLST profile of each individual isolate can be retrieved easily using its unique MID. With HiMLST we have profiled 575 isolates of Legionella pneumophila, Staphylococcus aureus, Pseudomonas aeruginosa and Streptococcus pneumoniae in mixed species HiMLST experiments. In conclusion, the introduction of HiMLST paves the way for a broad employment of the MLST as a high-quality and cost-effective method for typing microbial species. PMID:22815712

  2. Chronic Depression of Serum Sialic Acid Levels in Alloxan-Induced Diabetes

    DTIC Science & Technology

    1974-10-01

    Serum L- Fucose , Protein-Bound Hexose, and Total Protein Levels in Alloxan Diabetic and Control Rats at Various Time Intervals After Treatment 5...ABSTRACT This study was performed to determine whether alloxan treatment of rats alters levels of the terminal carbohydrate residues L- fucose and...occurs with no apparent alteration in the level of L- fucose . The depression in sialic acid level may be attributed in part to decreased activities of

  3. Nuclear hormone receptors involved in neoplasia: erb A exhibits a novel DNA sequence specificity determined by amino acids outside of the zinc-finger domain.

    PubMed Central

    Chen, H; Smit-McBride, Z; Lewis, S; Sharif, M; Privalsky, M L

    1993-01-01

    The erb A oncogene is a dominant negative allele of a thyroid hormone receptor gene and acts in the cancer cell by encoding a transcriptional repressor. We demonstrate here that the DNA sequence recognition properties of the oncogenic form of the erb A protein are significantly altered from those of the normal thyroid hormone receptors and more closely resemble those of the retinoic acid receptors; this alteration appears to play an important role in defining the targets of erb A action in neoplasia. Unexpectedly, the novel DNA recognition properties of erb A are encoded by an N-terminal region not previously implicated as playing this function in current models of receptor-DNA interaction. Two N-terminal erb A amino acids in particular, histidine 12 and cysteine 32, contribute to this phenomenon, acting in conjunction with amino acids in the zinc finger domain. The effects of the N-terminal domain can be observed at the level of both DNA binding and transcriptional modulation. Our results indicate that unanticipated determinants within the nuclear hormone receptors participate in DNA sequence recognition and may contribute to the differential target gene specificity displayed by different receptor forms. Images PMID:8096060

  4. Serum nitrite and nitrate levels in epileptic children using valproic acid or carbamazepine.

    PubMed

    Karabiber, Hamza; Yakinci, Cengiz; Durmaz, Yasar; Temel, Ismail; Mehmet, Nihayet

    2004-01-01

    In experimental epilepsy studies, nitric oxide was found to act as both proconvulsant and anticonvulsant. The objective of this study was to investigate the effects of valproic acid and carbamazepine on serum levels of nitrite and nitrate, which are the metabolites of nitric oxide. To achieve this goal, serum nitrite and nitrate levels were determined in active epileptic 34 children using valproic acid and 23 children using carbamazepine and in non-active epileptic 38 children (control group) not using any antiepileptic drug. In the valproic acid group serum nitrite and nitrate levels were 2.66 +/- 2.11 micromol/l and 69.35 +/- 23.20 micromol/l, 1.89 +/- 1.01 micromol/l and 49.39 +/- 10.61 micromol/l in the carbamazepine group, and 1.22 +/- 0.55 micromol/l, 29.53 +/- 10.05 micromol in the control group, respectively. Nitrite and nitrate levels were significantly high in both valproic acid and carbamazepine groups compared to the control group (P < 0.01). When valproic acid and carbamazepine groups were compared to each other, level of nitrate was found statistically higher in the valproic acid group in relation to the carbamazepine group (P < 0.01), however, there was no statistically significant difference in the levels of nitrite (P > 0.05). No relation could be found between serum drug levels and nitrite and nitrate levels. According to these results, it can be suggested that valproic acid and carbamazepine might have antiepileptic effects through nitric oxide.

  5. Comparative Indole-3-Acetic Acid Levels in the Slender Pea and Other Pea Phenotypes 1

    PubMed Central

    Law, David M.; Davies, Peter J.

    1990-01-01

    Free indole-3-acetic acid levels were measured by gas chromatography-mass spectrometry in three ultra-tall `slender' Pisum sativum L. lines differing in gibberellin content. Measurements were made for apices and stem elongation zones of light-grown plants and values were compared with wild-type, dwarf, and nana phenotypes in which internode length is genetically regulated, purportedly via the gibberellin level. Indole-3-acetic acid levels of growing stems paralleled growth rates in all lines, and were high in all three slender genotypes. Growth was inhibited by p-chlorophenoxyisobutyric acid, demonstrating the requirement of auxin activity for stem elongation, and also by the ethylene precursor 1-aminocyclopropane-1-carboxylic acid. It is concluded that the slender phenotype may arise from constant activation of a gibberellin receptor or transduction chain event leading directly or indirectly to elevated levels of indole-3-acetic acid, and that increased indole-3-acetic acid levels are a significant factor in the promotion of stem elongation. PMID:16667653

  6. Drilling and dating New Jersey oligocene-miocene sequences: Ice volume, global sea level, and Exxon records

    SciTech Connect

    Miller, K.G.; Mountain, G.S.

    1996-02-23

    Oligocene to middle Miocene sequence boundaries on the New Jersey coastal plain (Ocean Drilling Project Leg 150X) and continental slope (Ocean Drilling Project Leg 150) were dated by integrating strontium isotopic stratigraphy, magnetostratigraphy, and biostratigraphy (planktonic foraminifera, nannofossils, dinocysts, and diatoms). The ages of coastal plain unconformities and slope seismic reflectors (unconformities or stratal breaks with no discernible hiatuses) match the ages of global {delta}{sup 18}O increases (inferred glacioeustatic lowerings) measured in deep-sea sites. These correlations confirm a causal link between coastal plain and slope sequence boundaries: both formed during global sea-level lowerings. The ages of New Jersey sequence boundaries and global {delta}{sup 18}O increases also correlate well within the Exxon Production Research sea-level records of Haq et al. and Vail et al., validating and refining their compilations. 33 refs., 2 figs., 1 tab.

  7. Genome wide identification of microRNAs involved in fatty acid and lipid metabolism of Brassica napus by small RNA and degradome sequencing.

    PubMed

    Wang, Zhiwei; Qiao, Yan; Zhang, Jingjing; Shi, Wenhui; Zhang, Jinwen

    2017-04-01

    Rapeseed (Brassica napus) is an important cash crop considered as the third largest oil crop worldwide. Rapeseed oil contains various saturation or unsaturation fatty acids, these fatty acids, whose could incorporation with TAG form into lipids stored in seeds play various roles in the metabolic activity. The different fatty acids in B. napus seeds determine oil quality, define if the oil is edible or must be used as industrial material. miRNAs are kind of non-coding sRNAs that could regulate gene expressions through post-transcriptional modification to their target transcripts playing important roles in plant metabolic activities. We employed high-throughput sequencing to identify the miRNAs and their target transcripts involved in fatty acids and lipids metabolism in different development of B. napus seeds. As a result, we identified 826 miRNA sequences, including 523 conserved and 303 newly miRNAs. From the degradome sequencing, we found 589 mRNA could be targeted by 236 miRNAs, it includes 49 novel miRNAs and 187 conserved miRNAs. The miRNA-target couple suggests that bna-5p-163957_18, bna-5p-396192_7, miR9563a-p3, miR9563b-p5, miR838-p3, miR156e-p3, miR159c and miR1134 could target PDP, LACS9, MFPA, ADSL1, ACO32, C0401, GDL73, PlCD6, OLEO3 and WSD1. These target transcripts are involving in acetyl-CoA generate and carbon chain desaturase, regulating the levels of very long chain fatty acids, β-oxidation and lipids transport and metabolism process. At the same, we employed the q-PCR to valid the expression of miRNAs and their target transcripts that involve in fatty acid and lipid metabolism, the result suggested that the miRNA and their transcript expression are negative correlation, which in accord with the expression of miRNA and its target transcript. The study findings suggest that the identified miRNA may play important role in the fatty acids and lipids metabolism in seeds of B. napus.

  8. Complete amino acid sequence of an acidic, cardiotoxic phospholipase A2 from the venom of Ophiophagus hannah (King Cobra): a novel cobra venom enzyme with "pancreatic loop".

    PubMed

    Huang, M Z; Gopalakrishnakone, P; Chung, M C; Kini, R M

    1997-02-15

    A phospholipase A2 (OHV A-PLA2) from the venom of Ophiophagus hannah (King cobra) is an acidic protein exhibiting cardiotoxicity, myotoxicity, and antiplatelet activity. The complete amino acid sequence of OHV A-PLA2 has been determined using a combination of Edman degradation and mass spectrometric techniques. OHV A-PLA2 is composed of a single chain of 124 amino acid residues with 14 cysteines and a calculated molecular weight of 13719 Da. It contains the loop of residues (62-66) found in pancreatic PLA2s and hence belongs to class IB enzymes. This pancreatic loop is between two proline residues (Pro 59 and Pro 68) and contains several hydrophilic amino acids (Ser and Asp). This region has high degree of conformational flexibility and is on the surface of the molecule, and hence it may be a potential protein-protein interaction site. A relatively low sequence homology is found between OHV A-PLA2 and other known cardiotoxic PLA2s, and hence a contiguous segment could not be identified as a site responsible for the cardiotoxic activity.

  9. 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+.

  10. The amino acid sequence of the zinc-requiring beta-lactamase II from the bacterium Bacillus cereus 569.

    PubMed

    Ambler, R P; Daniel, M; Fleming, J; Hermoso, J M; Pang, C; Waley, S G

    1985-09-23

    The amino acid sequence of the zinc-requiring beta-lactamase II from Bacillus cereus strain 569 has been determined. It consists of a single polypeptide chain of 227 residues. It is the only example so far fully characterized of a class B beta-lactamase, and is structurally and mechanistically distinct from both the widely distributed class A beta-lactamases (such as the Escherichia coli RTEM enzyme) and from the chromosomally encoded class C enzymes from Gram-negative bacteria.

  11. Uric Acid Level and Erectile Dysfunction In Patients With Coronary Artery Disease

    PubMed Central

    Solak, Yalcin; Akilli, Hakan; Kayrak, Mehmet; Aribas, Alpay; Gaipov, Abduzhappar; Turk, Suleyman; Perez-Pozo, Santos E.; Covic, Adrian; McFann, Kim; Johnson, Richard J.; Kanbay, Mehmet

    2013-01-01

    Introduction Erectile dysfunction (ED) is a frequent complaint of elderly subjects, and is closely associated with endothelial dysfunction and cardiovascular disease. Uric acid is also associated with endothelial dysfunction, oxidative stress and cardiovascular disease, raising the hypothesis that an increased serum uric acid might predict erectile dysfunction in patients who are at risk for coronary artery disease. Aim To evaluate the association of serum uric acid levels with presence and severity of ED in patients presenting with chest pain of presumed cardiac origin. Methods This is a cross-sectional study of 312 adult male patients with suspected coronary artery disease who underwent exercise stress test (EST) for workup of chest pain and completed a sexual health inventory for men (SHIM) survey form to determine the presence and severity of ED. Routine serum biochemistry (and uric acid levels) were measured. Logistic regression analysis was used to assess risk factors for ED. Main Outcome Measures The short version of the international index of erectile function (IIEF-5) questionnaire diagnosed ED (cutoff score ≤21). Serum Uric acid levels were determined. Patients with chest pain of suspected cardiac origin underwent an exercise stress test. Results 149 of 312 (47.7%) male subjects had ED by survey criteria. Patients with ED were older and had more frequent CAD, hypertension, diabetes, and impaired renal function, and also had significantly higher levels of uric acid, fibrinogen, glucose, CRP, triglycerides compared with patients without ED. Uric acid levels were associated with ED by univariate analysis (OR = 1.36, p = 0.002); however, this association was not observed in multivariate analysis adjusted for eGFR. Conclusion Subjects presenting with chest pain of presumed cardiac origin are more likely to have ED if they have elevated uric acid levels. PMID:24433559

  12. Polyunsaturated Fatty Acid Levels in Maternal Erythrocytes of Japanese Women during Pregnancy and after Childbirth

    PubMed Central

    Kawabata, Terue; Kagawa, Yasuo; Kimura, Fumiko; Miyazawa, Teruo; Saito, Shoji; Arima, Takahiro; Nakai, Kunihiko; Yaegashi, Nobuo

    2017-01-01

    Background: The transport of polyunsaturated fatty acids (PUFAs), such as arachidonic acid (ARA, 20:4n-6) and docosahexaenoic acid (DHA, 22:6n-3), to the fetus from maternal stores increases depending on the fetal requirements for PUFA during the last trimester of pregnancy. Therefore, maternal blood PUFA changes physiologically with gestational age. However, the changes in PUFA levels in maternal blood erythrocytes during pregnancy and after childbirth have not been fully investigated in a fish-eating population. Objective: To examine the changes of ARA and DHA levels in maternal erythrocytes with the progress of pregnancy and the relationship between maternal and umbilical cord erythrocyte PUFA levels in pregnant Japanese women who habitually eat fish and shellfish. Design: This study was performed as a part of the adjunct study of the Japan Environment and Children’s Study (JECS). The participants were 74 pregnant women. The maternal blood samples were collected at 27, 30, and 36 weeks of pregnancy, and 2 days and 1 month after delivery, and umbilical cord blood was collected at delivery. The fatty acid levels of erythrocytes in these blood samples were determined. Results: ARA and DHA levels in maternal erythrocytes tended to decrease with the progress of pregnancy. While the DHA level decreased further after delivery, the ARA level returned to the value at 27 weeks of pregnancy within 1 month after delivery. The n-3 and n-6 PUFA levels in maternal erythrocytes at 27, 30, and 36 weeks of pregnancy were significantly positively correlated with the corresponding fatty acid levels in umbilical cord erythrocytes. Conclusion: The present findings showed a significant change in erythrocyte PUFA levels during pregnancy and after childbirth in a fish-eating population. The PUFA levels of maternal blood after the second trimester may be a reliable marker for predicting PUFA levels in infants’ circulating blood. PMID:28272345

  13. 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

  14. The isolation, purification and amino-acid sequence of insulin from the teleost fish Cottus scorpius (daddy sculpin).

    PubMed

    Cutfield, J F; Cutfield, S M; Carne, A; Emdin, S O; Falkmer, S

    1986-07-01

    Insulin from the principal islets of the teleost fish, Cottus scorpius (daddy sculpin), has been isolated and sequenced. Purification involved acid/alcohol extraction, gel filtration, and reverse-phase high-performance liquid chromatography to yield nearly 1 mg pure insulin/g wet weight islet tissue. Biological potency was estimated as 40% compared to porcine insulin. The sculpin insulin crystallised in the absence of zinc ions although zinc is known to be present in the islets in significant amounts. Two other hormones, glucagon and pancreatic polypeptide, were copurified with the insulin, and an N-terminal sequence for pancreatic polypeptide was determined. The primary structure of sculpin insulin shows a number of sequence changes unique so far amongst teleost fish. These changes occur at A14 (Arg), A15 (Val), and B2 (Asp). The B chain contains 29 amino acids and there is no N-terminal extension as seen with several other fish. Presumably as a result of the amino acid substitutions, sculpin insulin does not readily form crystals containing zinc-insulin hexamers, despite the presence of the coordinating B10 His.

  15. The amino acid sequence of a carbohydrate-containing immunoglobulin-light-chain-type amyloid-fibril protein.

    PubMed Central

    Tveteraas, T; Sletten, K; Westermark, P

    1985-01-01

    The amino acid sequence of an amyloid-fibril protein Es492 of immunoglobulin-lambda-light-chain origin (AL) was elucidated. The amyloid fibrils were obtained from the spleen of a patient who died from systemic amyloidosis. The amino acid sequence was elucidated from structural studies of peptides derived from digestion of the protein with trypsin, thermolysin, chymotrypsin and Staphylococcus aureus V8 proteinase and from cleavage of the protein with CNBr and BNPS-skatole. A heterogeneity in the length of the polypeptide was seen in the C-terminal region. The protein was by sequence homology to other lambda-chains shown to be of the V lambda II subgroup. Although an extensive homology was seen, some amino acid residues in positions 26, 31, 32, 40, 44, 93, 97, 98 and 99 have not previously been reported in these positions of V lambda II proteins. The significance of these residues in the fibril formation is unclear. The protein was found to contain carbohydrate, with glycosylation sites in two of the hypervariable regions. PMID:3936482

  16. The Effect of Nucleic Acid Extraction Platforms and Sample Storage on the Integrity of Viral RNA for Use in Whole Genome Sequencing.

    PubMed

    Lewandowski, Kuiama; Bell, Andrew; Miles, Rory; Carne, Simon; Wooldridge, David; Manso, Carmen; Hennessy, Nicola; Bailey, Daniel; Pullan, Steven T; Gharbia, Saheer; Vipond, Richard

    2017-03-01

    Extraction of viral RNA and the storage of sample material are extremely important factors in the detection and whole genome sequencing (WGS) of viral pathogens. Although PCR-based detection methods focus on small amplicons, viral WGS applications require RNA of high quality and integrity for adequate sequence coverage and depth. This study examined the fitness of one manual and four automated RNA extraction platforms commonly used in diagnostic laboratories for use in metagenomic sequencing, how the practice of storing sample material in Qiagen buffer AVL before extraction affected the integrity of viral RNA and its suitability for use in amplicon-based WGS methods, and how the addition of Triton X-100 to buffer AVL affected the capability of the extraction platforms and the integrity of viral RNA in stored samples. This study found that the EZ1 platform gave the best performance of the automated platforms and gave comparable results to the frequently used manual Qiagen extraction protocol when extracted viral RNA was used in metagenomics sequencing. To maintain high levels of viral RNA integrity suitable for amplicon-based WGS, nucleic acid should be extracted from samples immediately, because even short storage periods in buffer AVL have a severe effect on integrity, and the addition of Triton X-100 had little effect on the quality of viral material for WGS.

  17. Coronavirus genome: prediction of putative functional domains in the non-structural polyprotein by comparative amino acid sequence analysis.

    PubMed Central

    Gorbalenya, A E; Koonin, E V; Donchenko, A P; Blinov, V M

    1989-01-01

    Amino acid sequences of 2 giant non-structural polyproteins (F1 and F2) of infectious bronchitis virus (IBV), a member of Coronaviridae, were compared, by computer-assisted methods, to sequences of a number of other positive strand RNA viral and cellular proteins. By this approach, juxtaposed putative RNA-dependent RNA polymerase, nucleic acid binding ("finger"-like) and RNA helicase domains were identified in F2. Together, these domains might constitute the core of the protein complex involved in the primer-dependent transcription, replication and recombination of coronaviruses. In F1, two cysteine protease-like domains and a growth factor-like one were revealed. One of the putative proteases of IBV is similar to 3C proteases of picornaviruses and related enzymes of como- nepo- and potyviruses. Search of IBV F1 and F2 sequences for sites similar to those cleaved by the latter proteases and intercomparison of the surrounding sequence stretches revealed 13 dipeptides Q/S(G) which are probably cleaved by the coronavirus 3C-like protease. Based on these observations, a partial tentative scheme for the functional organization and expression strategy of the non-structural polyproteins of IBV was proposed. It implies that, despite the general similarity to other positive strand RNA viruses, and particularly to potyviruses, coronaviruses possess a number of unique structural and functional features. PMID:2526320

  18. Recognition of 5'-YpG-3' sequences by coupled stacking/hydrogen bonding interactions with amino acid residues.

    PubMed

    Lamoureux, Jason S; Maynes, Jason T; Glover, J N Mark

    2004-01-09

    The combined biochemical and structural study of hundreds of protein-DNA complexes has indicated that sequence-specific interactions are mediated by two mechanisms termed direct and indirect readout. Direct readout involves direct interactions between the protein and base-specific atoms exposed in the major and minor grooves of DNA. For indirect readout, the protein recognizes DNA by sensing conformational variations in the structure dependent on nucleotide sequence, typically through interactions with the phosphodiester backbone. Based on our recent structure of Ndt80 bound to DNA in conjunction with a search of the existing PDB database, we propose a new method of sequence-specific recognition that utilizes both direct and indirect readout. In this mode, a single amino acid side-chain recognizes two consecutive base-pairs. The 3'-base is recognized by canonical direct readout, while the 5'-base is recognized through a variation of indirect readout, whereby the conformational flexibility of the particular dinucleotide step, namely a 5'-pyrimidine-purine-3' step, facilitates its recognition by the amino acid via cation-pi interactions. In most cases, this mode of DNA recognition helps explain the sequence specificity of the protein for its target DNA.

  19. Prednisone lowers serum uric acid levels in patients with decompensated heart failure by increasing renal uric acid clearance.

    PubMed

    Liu, Chao; Zhen, Yuzhi; Zhao, Qingzhen; Zhai, Jian-Long; Liu, Kunshen; Zhang, Jian-Xin

    2016-07-01

    Clinical studies have shown that large doses of prednisone could lower serum uric acid (SUA) in patients with decompensated heart failure (HF); however, the optimal dose of prednisone and underlying mechanisms are unknown. Thirty-eight patients with decompensated HF were randomized to receive standard HF care alone (n = 10) or with low-dose (15 mg/day, n = 8), medium-dose (30 mg/day, n = 10), or high-dose prednisone (60 mg/day, n = 10), for 10 days. At the end of the study, only high-dose prednisone significantly reduced SUA, whereas low- and medium-dose prednisone and standard HF care had no effect on SUA. The reduction in SUA in high-dose prednisone groups was associated with a significant increase in renal uric acid clearance. In conclusion, prednisone can reduce SUA levels by increasing renal uric acid clearance in patients with decompensated HF.

  20. Deduced amino acid sequence, functional expression, and unique enzymatic properties of the form I and form II ribulose bisphosphate carboxylase/oxygenase from the chemoautotrophic bacterium Thiobacillus denitrificans.

    PubMed

    Hernandez, J M; Baker, S H; Lorbach, S C; Shively, J M; Tabita, F R

    1996-01-01

    The cbbL cbbS and cbbM genes of Thiobacillus denitrificans, encoding form I and form II ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO), respectively, were found to complement a RubisCO-negative mutant of Rhodobacter sphaeroides to autotrophic growth. Endogenous T. denitrificans promoters were shown to function in R. sphaeroides, resulting in high levels of cbbL cbbS and cbbM expression in the R. sphaeroides host. This expression system provided high levels of both T. denitrificans enzymes, each of which was highly purified. The deduced amino acid sequence of the form I enzyme indicated that the large subunit was closely homologous to previously sequenced form I RubisCO enzymes from sulfur-oxidizing bacteria. The form I T. denitrificans enzyme possessed a very low substrate specificity factor and did not exhibit fallover, and yet this enzyme showed a poor ability to recover from incubation with ribulose 1,5-bisphosphate. The deduced amino acid sequence of the form II T. denitrificans enzyme resembled those of other form II RubisCO enzymes. The substrate specificity factor was characteristically low, and the lack of fallover and the inhibition by ribulose 1,5-bisphosphate were similar to those of form II RubisCO obtained from nonsulfur purple bacteria. Both form I and form II RubisCO from T. denitrificans possessed high KCO2 values, suggesting that this organism might suffer in environments containing low levels of dissolved CO2. These studies present the initial description of the kinetic properties of form I and form II RubisCO from a chemoautotrophic bacterium that synthesizes both types of enzyme.

  1. Deduced amino acid sequence, functional expression, and unique enzymatic properties of the form I and form II ribulose bisphosphate carboxylase/oxygenase from the chemoautotrophic bacterium Thiobacillus denitrificans.

    PubMed Central

    Hernandez, J M; Baker, S H; Lorbach, S C; Shively, J M; Tabita, F R

    1996-01-01

    The cbbL cbbS and cbbM genes of Thiobacillus denitrificans, encoding form I and form II ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO), respectively, were found to complement a RubisCO-negative mutant of Rhodobacter sphaeroides to autotrophic growth. Endogenous T. denitrificans promoters were shown to function in R. sphaeroides, resulting in high levels of cbbL cbbS and cbbM expression in the R. sphaeroides host. This expression system provided high levels of both T. denitrificans enzymes, each of which was highly purified. The deduced amino acid sequence of the form I enzyme indicated that the large subunit was closely homologous to previously sequenced form I RubisCO enzymes from sulfur-oxidizing bacteria. The form I T. denitrificans enzyme possessed a very low substrate specificity factor and did not exhibit fallover, and yet this enzyme showed a poor ability to recover from incubation with ribulose 1,5-bisphosphate. The deduced amino acid sequence of the form II T. denitrificans enzyme resembled those of other form II RubisCO enzymes. The substrate specificity factor was characteristically low, and the lack of fallover and the inhibition by ribulose 1,5-bisphosphate were similar to those of form II RubisCO obtained from nonsulfur purple bacteria. Both form I and form II RubisCO from T. denitrificans possessed high KCO2 values, suggesting that this organism might suffer in environments containing low levels of dissolved CO2. These studies present the initial description of the kinetic properties of form I and form II RubisCO from a chemoautotrophic bacterium that synthesizes both types of enzyme. PMID:8550452

  2. Cenozoic global sea level, sequences, and the New Jersey transect: Results from coastal plain and continental slope drilling

    USGS Publications Warehouse

    Miller, K.G.; Mountain, Gregory S.; Browning, J.V.; Kominz, M.; Sugarman, P.J.; Christie-Blick, N.; Katz, M.E.; Wright, J.D.

    1998-01-01

    The New Jersey Sea Level Transect was designed to evaluate the relationships among global sea level (eustatic) change, unconformity-bounded sequences, and variations in subsidence, sediment supply, and climate on a passive continental margin. By sampling and dating Cenozoic strata from coastal plain and continental slope locations, we show that sequence boundaries correlate (within ??0.5 myr) regionally (onshore-offshore) and interregionally (New Jersey-Alabama-Bahamas), implicating a global cause. Sequence boundaries correlate with ??18O increases for at least the past 42 myr, consistent with an ice volume (glacioeustatic) control, although a causal relationship is not required because of uncertainties in ages and correlations. Evidence for a causal connection is provided by preliminary Miocene data from slope Site 904 that directly link ??18O increases with sequence boundaries. We conclude that variation in the size of ice sheets has been a primary control on the formation of sequence boundaries since ~42 Ma. We speculate that prior to this, the growth and decay of small ice sheets caused small-amplitude sea level changes (<20 m) in this supposedly ice-free world because Eocene sequence boundaries also appear to correlate with minor ??18O increases. Subsidence estimates (backstripping) indicate amplitudes of short-term (million-year scale) lowerings that are consistent with estimates derived from ??18O studies (25-50 m in the Oligocene-middle Miocene and 10-20 m in the Eocene) and a long-term lowering of 150-200 m over the past 65 myr, consistent with estimates derived from volume changes on mid-ocean ridges. Although our results are consistent with the general number and timing of Paleocene to middle Miocene sequences published by workers at Exxon Production Research Company, our estimates of sea level amplitudes are substantially lower than theirs. Lithofacies patterns within sequences follow repetitive, predictable patterns: (1) coastal plain sequences consist

  3. Serum homocysteine, vitamin B12, folic acid levels and methylenetetrahydrofolate reductase (MTHFR) gene polymorphism in vitiligo.

    PubMed

    Yasar, Ali; Gunduz, Kamer; Onur, Ece; Calkan, Mehmet

    2012-01-01

    The aim of this study was to determine serum vitamin B12, folic acid and homocysteine (Hcy) levels as well as MTHFR (C677, A1298C) gene polymorphisms in patients with vitiligo, and to compare the results with healthy controls. Forty patients with vitiligo and 40 age and sex matched healthy subjects were studied. Serum vitamin B12 and folate levels were determined by enzyme-linked immunosorbent assay. Plasma Hcy levels and MTHFR polymorphisms were determined by chemiluminescence and real time PCR methods, respectively. Mean serum vitamin B12 and Hcy levels were not significantly different while folic acid levels were significantly lower in the control group. There was no significant relationship between disease activity and vitamin B12, folic acid and homocystein levels. No significant difference in C677T gene polymorphism was detected. Heterozygote A1298C gene polymorphism in the patient group was statistically higher than the control group. There was no significant relationship between MTHFR gene polymorphisms and vitamin B12, folic acid and homocysteine levels. In conclusion, vitamin B12, folate and Hcy levels are not altered in vitiligo and MTHFR gene mutations (C677T and A1298C) do not seem to create susceptibility for vitiligo.

  4. Two Levels of Caffeine Ingestion on Blood Lactate and Free Fatty Acid Responses during Incremental Exercise.

    ERIC Educational Resources Information Center

    McNaughton, Lars

    1987-01-01

    Research was conducted to determine the effects of two doses of caffeine on the lactate threshold and also to examine the effects on substrate utilization during incremental cycle ergometry. Results found that caffeine increased heart rates and free fatty acid levels for all workloads and decreased blood lactate levels at some of the workloads.…

  5. Accurate calculation of the p Ka of trifluoroacetic acid using high-level ab initio calculations

    NASA Astrophysics Data System (ADS)

    Namazian, Mansoor; Zakery, Maryam; Noorbala, Mohammad R.; Coote, Michelle L.

    2008-01-01

    The p Ka value of trifluoroacetic acid has been successfully calculated using high-level ab initio methods such as G3 and CBS-QB3. Solvation energies have been calculated using CPCM continuum model of solvation at the HF and B3-LYP levels of theory with various basis sets. Excellent agreement with experiment (to within 0.4 p Ka units) was obtained using CPCM solvation energies at the B3-LYP/6-31+G(d) level (or larger) in conjunction with CBS-QB3 or G3 gas-phase energies of trifluoroacetic acid and its anion.

  6. Fasting levels of monoketonic bile acids in human peripheral and portal circulation.

    PubMed

    Björkhem, I; Angelin, B; Einarsson, K; Ewerth, S

    1982-09-01

    . Einarsson, and S. Ewerth. Fasting levels of monoketonic bile acids in human peripheral and portal circulation.

  7. Evaluation of serum sialic acid, fucose levels and their ratio in oral squamous cell carcinoma

    PubMed Central

    Chinnannavar, Sangamesh Ningappa; Ashok, Lingappa; Vidya, Kodige Chandrashekhar; Setty, Sunil Mysore Kantharaja; Narasimha, Guru Eraiah; Garg, Ranjana

    2015-01-01

    Background: Detection of cancer at the early stage is of utmost importance to decrease the morbidity and mortality of the disease. Apart from the conventional biopsy, minimally invasive methods like serum evaluation are used for screening large populations. Thus, this study aimed to estimate serum levels of sialic acid and fucose and their ratio in oral cancer patients and in healthy control group to evaluate their role in diagnosis. Materials and Methods: Serum samples were collected from 52 healthy controls (group I) and 52 squamous cell carcinoma patients (group II). Estimation of serum levels of sialic acid and fucose and their ratio was performed. This was correlated histopathologically with the grades of carcinoma. Statistical analysis was done by using analysis of variance (ANOVA) test and unpaired “t” test. Results: Results showed that serum levels of sialic acid and fucose were significantly higher in oral cancer patients compared to normal healthy controls (P < 0.001). The sialic acid to fucose ratio was significantly lower in cancer patients than in normal controls (P < 0.01). However, comparison with histological grading, habits, gender, and age group did not show any significant result. Conclusion: The mean serum sialic acid and fucose levels showed an increasing trend from controls to malignant group and their corresponding ratio showed decreasing trend from controls to malignant group. The ratio of sialic acid to fucose can be a useful diagnostic aid for oral cancer patients. PMID:26759796

  8. Antibacterial drug treatment increases intestinal bile acid absorption via elevated levels of ileal apical sodium-dependent bile acid transporter but not organic solute transporter α protein.

    PubMed

    Miyata, Masaaki; Hayashi, Kenjiro; Yamakawa, Hiroki; Yamazoe, Yasushi; Yoshinari, Kouichi

    2015-01-01

    Antibacterial drug treatment increases the bile acid pool size and hepatic bile acid concentration through the elevation of hepatic bile acid synthesis. However, the involvement of intestinal bile acid absorption in the increased bile acid pool size remains unclear. To determine whether intestinal bile acid absorption contributes to the increased bile acid pool in mice treated with antibacterial drugs, we evaluated the levels of bile acid transporter proteins and the capacity of intestinal bile acid absorption. Ileal apical sodium-dependent bile acid transporter (ASBT) mRNA and protein levels were significantly increased in ampicillin (ABPC)-treated mice, whereas organic solute transporter α (OSTα) mRNA levels, but not protein levels, significantly decreased in mice. Similar alterations in the expression levels of bile acid transporters were observed in mice treated with bacitracin/neomycin/streptomycin. The capacity for intestinal bile acid absorption was evaluated by an in situ loop method. Increased ileal absorption of taurochenodeoxycholic acid was observed in mice treated with ABPC. These results suggest that intestinal bile acid absorption is elevated in an ASBT-dependent manner in mice treated with antibacterial drugs.

  9. Amino acid sequence and some properties of phytolacain G, a cysteine protease from growing fruit of pokeweed, Phytolacca americana.

    PubMed

    Uchikoba, T; Arima, K; Yonezawa, H; Shimada, M; Kaneda, M

    2000-10-18

    A protease, phytolacain G, has been found to appear on CM-Sepharose ion-exchange chromatography of greenish small-size fruits of pokeweed, Phytolacca americana L, from ca. 2 weeks after flowering, and increases during fruit enlargement. Reddish ripe fruit of the pokeweed contained both phytolacain G and R. The molecular mass of phytolacain G was estimated to be 25.5 kDa by SDS-PAGE. Its amino acid sequence was reconstructed by automated sequence analysis of the peptides obtained after cleavage with Achromobacter protease I, chymotrypsin, and cyanogen bromide. The enzyme is composed of 216 amino acid residues, of which it shares 152 identical amino acid residues (70%) with phytolacain R, 126 (58%) with melain G, 108 (50%) with papain, 106 (49%) with actinidain, and 96 (44%) with stem bromelain. The amino acid residues forming the substrate binding S(2) pocket of papain, Tyr67, Pro68, Trp69, Val133, and Phe207, were predicted to be replaced by Trp, Met, His, Ala, and Ser in phytolacain G, respectively. As a consequence of these substitutions, the S(2) pocket is expected to be less hydrophobic in phytolacain G than in papain.

  10. Multiregion ultra-deep sequencing reveals early intermixing and variable levels of intratumoral heterogeneity in colorectal cancer.

    PubMed

    Suzuki, Yuka; Ng, Sarah Boonhsi; Chua, Clarinda; Leow, Wei Qiang; Chng, Jermain; Liu, Shi Yang; Ramnarayanan, Kalpana; Gan, Anna; Ho, Dan Liang; Ten, Rachel; Su, Yan; Lezhava, Alexandar; Lai, Jiunn Herng; Koh, Dennis; Lim, Kiat Hon; Tan, Patrick; Rozen, Steven G; Tan, Iain Beehuat

    2017-02-01

    Intratumor heterogeneity (ITH) contributes to cancer progression and chemoresistance. We sought to comprehensively describe ITH of somatic mutations, copy number, and transcriptomic alterations involving clinically and biologically relevant gene pathways in colorectal cancer (CRC). We performed multiregion, high-depth (384× on average) sequencing of 799 cancer-associated genes in 24 spatially separated primary tumor and nonmalignant tissues from four treatment-naïve CRC patients. We then used ultra-deep sequencing (17 075× on average) to accurately verify the presence or absence of identified somatic mutations in each sector. We also digitally measured gene expression and copy number alterations using NanoString assays. We identified the subclonal point mutations and determined the mutational timing and phylogenetic relationships among spatially separated sectors of each tumor. Truncal mutations, those shared by all sectors in the tumor, affected the well-described driver genes such as APC, TP53, and KRAS. With sequencing at 17 075×, we found that mutations first detected at a sequencing depth of 384× were in fact more widely shared among sectors than originally assessed. Interestingly, ultra-deep sequencing also revealed some mutations that were present in all spatially dispersed sectors, but at subclonal levels. Ultra-high-depth validation sequencing, copy number analysis, and gene expression profiling provided a comprehensive and accurate genomic landscape of spatial heterogeneity in CRC. Ultra-deep sequencing allowed more sensitive detection of somatic mutations and a more accurate assessment of ITH. By detecting the subclonal mutations with ultra-deep sequencing, we traced the genomic histories of each tumor and the relative timing of mutational events. We found evidence of early mixing, in which the subclonal ancestral mutations intermixed across the sectors before the acquisition of subsequent nontruncal mutations. Our findings also indicate that

  11. Decreased eicosapentaenoic acid levels in acne vulgaris reveals the presence of a proinflammatory state.

    PubMed

    Aslan, İbrahim; Özcan, Filiz; Karaarslan, Taner; Kıraç, Ebru; Aslan, Mutay

    2017-01-01

    This study aimed to determine circulating levels of polyunsaturated fatty acids (PUFAs), secretory phospholipase A2 (sPLA2), lipoprotein lipase (LPL) and measure circulating protein levels of angiopoietin-like protein 3 (ANGPTL3), ANGPTL4, cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) in patients with acne vulgaris. Serum from 21 control subjects and 31 acne vulgaris patients were evaluated for levels of arachidonic acid (AA, C20:4n- 6), dihomo-gamma-linolenic acid (DGLA, C20:3n-6), eicosapentaenoic acid (EPA, C20:5n-3) and docosahexaenoic acid (DHA, C22:6n-3). PUFA levels were determined by an optimized multiple reaction monitoring (MRM) method using ultra fast-liquid chromatography (UFLC) coupled with tandem mass spectrometry (MS/MS). Lipid profile, routine biochemical and hormone parameters were assayed by standard kit methods Serum EPA levels were significantly decreased while AA/EPA and DGLA/EPA ratio were significantly increased in acne vulgaris patients compared to controls. Serum levels of AA, DGLA and DHA showed no significant difference while activity of sPLA2 and LPL were significantly increased in acne vulgaris compared to controls. Results of this study reveal the presence of a proinflammatory state in acne vulgaris as shown by significantly decreased serum EPA levels and increased activity of sPLA2, AA/EPA and DGLA/EPA ratio. Increased LPL activity in the serum of acne vulgaris patients can be protective through its anti-dyslipidemic actions. This is the first study reporting altered EPA levels and increased sPLA2 activity in acne vulgaris and supports the use of omega-3 fatty acids as adjuvant treatment for acne patients.

  12. Childrens' learning and behaviour and the association with cheek cell polyunsaturated fatty acid levels.

    PubMed

    Kirby, A; Woodward, A; Jackson, S; Wang, Y; Crawford, M A

    2010-01-01

    Increasing interest in the role of omega-3 fatty acids in relation to neurodevelopmental disorders (e.g. ADHD, dyslexia, autism) has occurred as a consequence of some international studies highlighting this link. In particular, some studies have shown that children with ADHD may have lower concentrations of polyunsaturated fatty acids (PUFAs), particularly omega-3, in their red blood cells and plasma, and that supplementation with omega-3 fatty acids may alleviate behavioural symptoms in this population. However, in order to compare levels it seems appropriate to establish fatty acid levels in a mainstream school aged population and if levels relate to learning and behaviour. To date no study has established this. For this study, cheek cell samples from 411 typically developing school children were collected and analysed for PUFA content, in order to establish the range in this population. In addition, measures of general classroom attention and behaviour were assessed in these children by teachers and parents. Cognitive performance tests were also administered in order to explore whether an association between behaviour and/or cognitive performance and PUFA levels exists. Relationships between PUFA levels and socio-economic status were also explored. Measures of reading, spelling and intelligence did not show any association with PUFA levels, but some associations were noted with the level of omega-3 fatty acids and teacher and parental reports of behaviour, with some evidence that higher omega-3 levels were associated with decreased levels of inattention, hyperactivity, emotional and conduct difficulties and increased levels of prosocial behaviour. These findings are discussed in relation to previous findings from omega-3 supplementation studies with children.

  13. Acid mine drainage neutralization in a pilot sequencing batch reactor using limestone from a paper and pulp industry.

    PubMed

    Vadapalli, V R K; Zvimba, J N; Mathye, M; Fischer, H; Bologo, L

    2015-01-01

    This study investigated the implications of using two grades of limestone from a paper and pulp industry for neutralization of acid mine drainage (AMD) in a pilot sequencing batch reactor (SBR). In this regard, two grades of calcium carbonate were used to neutralize AMD in a SBR with a hydraulic retention time (including settling) of 100 min and a sludge retention time of 360 min, by simultaneously monitoring the Fe(II) removal kinetics and overall assessment of the AMD after treatment. The Fe(II) kinetics removal and overall AMD treatment were observed to be highly dependent on the limestone grade used, with Fe(II) completely removed to levels lower than 50 mg/L in cycle 1 after 30 min using high quality or pure paper and pulp limestone. On the contrary, the other grade limestone, namely waste limestone, could only achieve a similar Fe(II) removal efficiency after four cycles. It was also noticed that suspended solids concentration plays a significant role in Fe(II) removal kinetics. In this regard, using pure limestone from the paper and pulp industry will have advantages compared with waste limestone for AMD neutralization. It has significant process impacts for the SBR configuration as it allows one cycle treatment resulting in a significant reduction of the feed stock, with subsequent generation of less sludge during AMD neutralization. However, the use of waste calcium carbonate from the paper and pulp industry as a feed stock during AMD neutralization can achieve significant cost savings as it is cheaper than the pure limestone and can achieve the same removal efficiency after four cycles.

  14. Inverse Association Between Serum Uric Acid Levels and Alzheimer's Disease Risk.

    PubMed

    Du, Na; Xu, Donghua; Hou, Xu; Song, Xuejia; Liu, Cancan; Chen, Ying; Wang, Yangang; Li, Xin

    2016-05-01

    The association between Alzheimer's disease and uric acid levels had gained great interest in recent years, but there was still lack of definite evidence. A systematic review and meta-analysis of relevant studies was performed to comprehensively estimate the association. Relevant studies published before October 26, 2014, were searched in PubMed, Embase, and China Biology Medicine (CBM) databases. Study-specific data were combined using random-effects or fixed-effects models of meta-analysis according to between-study heterogeneity. Twenty-four studies (21 case-control and 3 cohort studies) were finally included into the meta-analysis. Those 21 case-control studies included a total of 1128 cases of Alzheimer's disease and 2498 controls without Alzheimer's disease. Those 3 cohort studies included a total of 7327 participants. Meta-analysis showed that patients with Alzheimer's disease had lower levels of uric acid than healthy controls (weighted mean difference (WMD) = -0.77 mg/dl, 95% CI -2.28 to -0.36, P = 0.0002). High serum uric acid levels were significantly associated with decreased risk of Alzheimer's disease (risk ratio (RR) = 0.66, 95% CI 0.52-0.85, P = 0.001). There was low risk of publication bias in the meta-analysis. There is an inverse association between serum uric acid levels and Alzheimer's disease. High serum uric acid level is a protective factor of Alzheimer's disease.

  15. Serum sialic acid and glycoprotein levels in some Libyan cancer patients.

    PubMed

    Balo, N N; Ishaq, M

    1991-01-01

    Sialic acid is a common conjugate of some serum glycoproteins and glycolipids. Elevated levels of serum sialic acid and alterations in serum glycoproteins have been observed in certain types of cancer. In this study sialic acid concentration in the sera of patients with various types of cancer was determined. In addition to this, serum glycoproteins were also analysed by electrophoretic method. Our results indicate that serum sialic acid levels are generally raised in all types of cancer studied. This increase was more pronounced in case of lung, bronchogenic, intestinal and breast cancer. Some alterations in the serum glycoprotein profiles were also observed, particularly in bronchogenic and gall bladder cancer where an additional band in the low molecular weight region was present and in lung, breast and lymphoma where a band in the middle molecular weight region was found missing when compared with normals.

  16. Ascorbic acid inhibits PMP22 expression by reducing cAMP levels.

    PubMed

    Kaya, Ferdinand; Belin, Sophie; Bourgeois, Patrice; Micaleff, Joelle; Blin, Olivier; Fontés, Michel

    2007-03-01

    Charcot-Marie-Tooth [CMT] syndrome is the most common hereditary peripheral neuropathy. CMT1A, which accounts for 50% of all CMT cases, usually results from triploidy of the PMP22 gene. Preclinical trials using an animal model show that disabled mice force-fed with high doses of ascorbic acid partially recover muscular strength after a few months of treatment, and suggest that high doses of ascorbic acid repress PMP22 expression. In this study, we demonstrated that ascorbic acid represses PMP22 gene expression by acting on intracellular cAMP levels and adenylate cyclase activity. This action is dose dependent and specific to ascorbic acid, since repression is not observed after treatment with other antioxidants. The new properties of ascorbic acid are discussed, along with the implications of these findings for CMT disease treatment.

  17. Different Levels of Learning Interact to Shape the Congruency Sequence Effect

    ERIC Educational Resources Information Center

    Weissman, Daniel H.; Hawks, Zoë W.; Egner, Tobias

    2016-01-01

    The congruency effect in distracter interference tasks is often reduced after incongruent relative to congruent trials. Moreover, this "congruency sequence effect" (CSE) is influenced by learning related to concrete stimulus and response features as well as by learning related to abstract cognitive control processes. There is an ongoing…

  18. Draft Genome Sequences of Streptococcus pneumoniae with High-Level Resistance to Respiratory Fluoroquinolones.

    PubMed

    Keness, Yoram; Bisharat, Naiel

    2016-03-31

    Streptococcus pneumoniaeis the leading cause of community-acquired pneumonia. Levofloxacin is a fluoroquinolone used for treatment of severe community-acquired pneumonia. Here, we describe the draft genome sequences ofS. pneumoniaewith emerging resistance to levofloxacin, resulting in failure of treatment of pneumococcal pneumonia.

  19. High-throughput amplicon sequencing and stream benthic bacteria: identifying the best taxonomic level for multiple-stressor research

    PubMed Central

    Salis, R. K.; Bruder, A.; Piggott, J. J.; Summerfield, T. C.; Matthaei, C. D.

    2017-01-01

    Disentangling the individual and interactive effects of multiple stressors on microbial communities is a key challenge to our understanding and management of ecosystems. Advances in molecular techniques allow studying microbial communities in situ and with high taxonomic resolution. However, the taxonomic level which provides the best trade-off between our ability to detect multiple-stressor effects versus the goal of studying entire communities remains unknown. We used outdoor mesocosms simulating small streams to investigate the effects of four agricultural stressors (nutrient enrichment, the nitrification inhibitor dicyandiamide (DCD), fine sediment and flow velocity reduction) on stream bacteria (phyla, orders, genera, and species represented by Operational Taxonomic Units with 97% sequence similarity). Community composition was assessed using amplicon sequencing (16S rRNA gene, V3-V4 region). DCD was the most pervasive stressor, affecting evenness and most abundant taxa, followed by sediment and flow velocity. Stressor pervasiveness was similar across taxonomic levels and lower levels did not perform better in detecting stressor effects. Community coverage decreased from 96% of all sequences for abundant phyla to 28% for species. Order-level responses were generally representative of responses of corresponding genera and species, suggesting that this level may represent the best compromise between stressor sensitivity and coverage of bacterial communities. PMID:28327636

  20. High-throughput amplicon sequencing and stream benthic bacteria: identifying the best taxonomic level for multiple-stressor research

    NASA Astrophysics Data System (ADS)

    Salis, R. K.; Bruder, A.; Piggott, J. J.; Summerfield, T. C.; Matthaei, C. D.

    2017-03-01

    Disentangling the individual and interactive effects of multiple stressors on microbial communities is a key challenge to our understanding and management of ecosystems. Advances in molecular techniques allow studying microbial communities in situ and with high taxonomic resolution. However, the taxonomic level which provides the best trade-off between our ability to detect multiple-stressor effects versus the goal of studying entire communities remains unknown. We used outdoor mesocosms simulating small streams to investigate the effects of four agricultural stressors (nutrient enrichment, the nitrification inhibitor dicyandiamide (DCD), fine sediment and flow velocity reduction) on stream bacteria (phyla, orders, genera, and species represented by Operational Taxonomic Units with 97% sequence similarity). Community composition was assessed using amplicon sequencing (16S rRNA gene, V3-V4 region). DCD was the most pervasive stressor, affecting evenness and most abundant taxa, followed by sediment and flow velocity. Stressor pervasiveness was similar across taxonomic levels and lower levels did not perform better in detecting stressor effects. Community coverage decreased from 96% of all sequences for abundant phyla to 28% for species. Order-level responses were generally representative of responses of corresponding genera and species, suggesting that this level may represent the best compromise between stressor sensitivity and coverage of bacterial communities.

  1. Beta-alanine and beta-aminoisobutyric acid levels in two siblings with dihydropyrimidinase deficiency.

    PubMed

    van Kuilenburg, A B P; Stroomer, A E M; Bosch, A M; Duran, M

    2008-06-01

    Dihydropyrimidinase (DHP) deficiency is an inborn error of the pyrimidine degradation pathway, affecting the hydrolytic ring opening of the dihydropyrimidines. In two siblings with a complete DHP deficiency and a variable clinical presentation, a normal concentration of beta-alanine and strongly decreased levels of beta-aminoisobutyric acid were observed in plasma, urine and CSF. No major differences were observed for the concentrations of the beta-amino acids in plasma and urine between the symptomatic and asymptomatic sibling. Thus, the relevance of the shortage of beta-aminoisobutyric acid for the onset of a clinical phenotype in patients with DHP deficiency remains to be established.

  2. Spectra, energy levels, and energy transition of lanthanide complexes with cinnamic acid and its derivatives

    NASA Astrophysics Data System (ADS)

    Zhou, Kaining; Feng, Zhongshan; Shen, Jun; Wu, Bing; Luo, Xiaobing; Jiang, Sha; Li, Li; Zhou, Xianju

    2016-04-01

    High resolution spectra and luminescent lifetimes of 6 europium(III)-cinnamic acid complex {[Eu2L6(DMF)(H2O)]·nDMF·H2O}m (L = cinnamic acid I, 4-methyl-cinnamic acid II, 4-chloro-cinnamic acid III, 4-methoxy-cinnamic acid IV, 4-hydroxy-cinnamic acid V, 4-nitro-cinnamic acid VI; DMF = N, N-dimethylformamide, C3H7NO) were recorded from 8 K to room temperature. The energy levels of Eu3 + in these 6 complexes are obtained from the spectra analysis. It is found that the energy levels of the central Eu3 + ions are influenced by the nephelauxetic effect, while the triplet state of ligand is lowered by the p-π conjugation effect of the para-substituted functional groups. The best energy matching between the ligand triplet state and the central ion excited state is found in complex I. While the other complexes show poorer matching because the gap of 5D0 and triplet state contracts.

  3. Spectra, energy levels, and energy transition of lanthanide complexes with cinnamic acid and its derivatives.

    PubMed

    Zhou, Kaining; Feng, Zhongshan; Shen, Jun; Wu, Bing; Luo, Xiaobing; Jiang, Sha; Li, Li; Zhou, Xianju

    2016-04-05

    High resolution spectra and luminescent lifetimes of 6 europium(III)-cinnamic acid complex {[Eu2L6(DMF)(H2O)]·nDMF·H2O}m (L=cinnamic acid I, 4-methyl-cinnamic acid II, 4-chloro-cinnamic acid III, 4-methoxy-cinnamic acid IV, 4-hydroxy-cinnamic acid V, 4-nitro-cinnamic acid VI; DMF=N, N-dimethylformamide, C3H7NO) were recorded from 8 K to room temperature. The energy levels of Eu(3+) in these 6 complexes are obtained from the spectra analysis. It is found that the energy levels of the central Eu(3+) ions are influenced by the nephelauxetic effect, while the triplet state of ligand is lowered by the p-π conjugation effect of the para-substituted functional groups. The best energy matching between the ligand triplet state and the central ion excited state is found in complex I. While the other complexes show poorer matching because the gap of (5)D0 and triplet state contracts.

  4. Serum iron, vitamin B12 and folic acid levels in Parkinson's disease.

    PubMed

    Madenci, Gulizar; Bilen, Sule; Arli, Berna; Saka, Mustafa; Ak, Fikri

    2012-07-01

    We aimed to investigate possible associations between systemic iron metabolism deficiency and Parkinson's disease, and also to research any possible correlations between stage of the disease and vitamin B12 and folic acid levels. 33 male and 27 female patients diagnosed with idiopathic Parkinson's disease and 22 male and 20 female age- and sex-matched controls were enrolled in the study. Having the diagnosis of secondary Parkinsonism or Parkinson plus syndromes, and for the females, not being in the menopausal stage were considered as exclusion criteria. Recordings of blood samples of both groups collected after 8 h fasts were assessed in terms of serum iron, ferritin levels and iron-binding capacity, vitamin B12 and folic acid levels. The Hoehn and Yahr scale was used to determine the stage of the disease. No statistically significant difference was found with respect to mean serum iron, median serum ferritin levels and median serum iron-binding capacity between the groups. A statistically significant but inverse correlation was found between symptoms' duration and serum iron and ferritin levels. There was no statistically significant difference between the groups with respect to vitamin B12 and folic acid levels. However, a statistically significant but inverse correlation was determined between the patients' vitamin B12 levels and the Hoehn and Yahr scores. As Parkinson's disease progresses, serum iron, ferritin and vitamin B12 levels may decrease. The lower levels of these parameters may be the cause of the progression or may be the result of it.

  5. Autistic Children Exhibit Decreased Levels of Essential Fatty Acids in Red Blood Cells

    PubMed Central

    Brigandi, Sarah A.; Shao, Hong; Qian, Steven Y.; Shen, Yiping; Wu, Bai-Lin; Kang, Jing X.

    2015-01-01

    Omega-6 (n-6) and omega-3 (n-3) polyunsaturated fatty acids (PUFA) are essential nutrients for brain development and function. However, whether or not the levels of these fatty acids are altered in individuals with autism remains debatable. In this study, we compared the fatty acid contents between 121 autistic patients and 110 non-autistic, non-developmentally delayed controls, aged 3–17. Analysis of the fatty acid composition of red blood cell (RBC) membrane phospholipids showed that the percentage of total PUFA was lower in autistic patients than in controls; levels of n-6 arachidonic acid (AA) and n-3 docosahexaenoic acid (DHA) were particularly decreased (p < 0.001). In addition, plasma levels of the pro-inflammatory AA metabolite prostaglandin E2 (PGE2) were higher in a subset of the autistic participants (n = 20) compared to controls. Our study demonstrates an alteration in the PUFA profile and increased production of a PUFA-derived metabolite in autistic patients, supporting the hypothesis that abnormal lipid metabolism is implicated in autism. PMID:25946342

  6. Infantile Refsum Disease: Influence of Dietary Treatment on Plasma Phytanic Acid Levels.

    PubMed

    Sá, Maria João Nabais; Rocha, Júlio C; Almeida, Manuela F; Carmona, Carla; Martins, Esmeralda; Miranda, Vasco; Coutinho, Miguel; Ferreira, Rita; Pacheco, Sara; Laranjeira, Francisco; Ribeiro, Isaura; Fortuna, Ana Maria; Lacerda, Lúcia

    2016-01-01

    Infantile Refsum disease (IRD) is one of the less severe of Zellweger spectrum disorders (ZSDs), a group of peroxisomal biogenesis disorders resulting from a generalized peroxisomal function impairment. Increased plasma levels of very long chain fatty acids (VLCFA) and phytanic acid are biomarkers used in IRD diagnosis. Furthermore, an increased plasma level of phytanic acid is known to be associated with neurologic damage. Treatment of IRD is symptomatic and multidisciplinary.The authors report a 3-year-old child, born from consanguineous parents, who presented with developmental delay, retinitis pigmentosa, sensorineural deafness and craniofacial dysmorphisms. While the relative level of plasma C26:0 was slightly increased, other VLCFA were normal. Thus, a detailed characterization of the phenotype was essential to point to a ZSD. Repeatedly increased levels of plasma VLCFA, along with phytanic acid and pristanic acid, deficient dihydroxyacetone phosphate acyltransferase activity in fibroblasts and identification of the homozygous pathogenic mutation c.2528G>A (p.Gly843Asp) in the PEX1 gene, confirmed this diagnosis. Nutritional advice and follow-up was proposed aiming phytanic acid dietary intake reduction. During dietary treatment, plasma levels of phytanic acid decreased to normal, and the patient's development evaluation showed slow progressive acquisition of new competences.This case report highlights the relevance of considering a ZSD in any child with developmental delay who manifests hearing and visual impairment and of performing a systematic biochemical investigation, when plasma VLCFA are mildly increased. During dietary intervention, a biochemical improvement was observed, and the long-term clinical effect of this approach needs to be evaluated.

  7. Modulation of anti-endotoxin property of Temporin L by minor amino acid substitution in identified phenylalanine zipper sequence.

    PubMed

    Srivastava, Saurabh; Kumar, Amit; Tripathi, Amit Kumar; Tandon, Anshika; Ghosh, Jimut Kanti

    2016-11-01

    A 13-residue frog antimicrobial peptide Temporin L (TempL) possesses versatile antimicrobial activities and is considered a lead molecule for the development of new antimicrobial agents. To find out the amino acid sequences that influence the anti-microbial property of TempL, a phenylalanine zipper-like sequence was identified in it which was not reported earlier. Several alanine-substituted analogs and a scrambled peptide having the same composition of TempL were designed for evaluating the role of this motif. To investigate whether leucine residues instead of phenylalanine residues at 'a' and/or 'd' position(s) of the heptad repeat sequence could alter its antimicrobial property, several TempL analogs were synthesized after replacing these phenylalanine residues with leucine residues. Replacing phenylalanine residues with alanine residues in the phenylalanine zipper sequence significantly compromised the anti-endotoxin property of TempL. This is evident from the higher production of tumor necrosis factor-α and interleukin-6 in lipopolysaccharide (LPS)-stimulated rat bone-marrow-derived macrophage cells in the presence of its alanine-substituted analogs than TempL itself. However, replacement of these phenylalanine residues with leucine residues significantly augmented anti-endotoxin property of TempL. A single alanine-substituted TempL analog (F8A-TempL) showed significantly reduced cytotoxicity but retained the antibacterial activity of TempL, while the two single leucine-substituted analogs (F5L-TempL and F8L-TempL), although exhibiting lower cytotoxicity, were able to retain the antibacterial activity of the parent peptide. The results demonstrate how minor amino acid substitutions in the identified phenylalanine zipper sequence in TempL could yield analogs with better antibacterial and/or anti-endotoxin properties with their plausible mechanism of action.

  8. Evaluation of cooling strategies for pumping of milk - impact of fatty acid composition on free fatty acid levels.

    PubMed

    Wiking, Lars; Bertram, Hanne C; Björck, Lennart; Nielsen, Jacob H

    2005-11-01

    Cooling strategies for pumping of raw milk were evaluated. Milk was pumped for 450 s at 31 degrees C, or pumped after cooling to 4 degrees C and subsequently subjected to various incubation times. Two types of milk were used; i.e. milk from cows fed a diet high in saturated fat supplements resulting in significantly larger milk fat globules than the other type of milk which comes from cows fed a low-fat diet that stimulates high de novo fat synthesis. The content of liquid fat was determined by low-field 1H NMR, which showed that milk from cows given the saturated fat diet also contained less liquid fat at both 4 degrees and 31 degrees C than the other type of milk. This can be ascribed to the differences in the fatty acid composition of the milk as a result of the fatty acid composition of the diets. After pumping of the milk at 31 degrees C, measurement of fat globule size distribution revealed a significant coalescence of milk fat globules in the milk obtained from the saturated fat diet due to pumping. Pumping at 4 degrees C or pumping the other type of milk did not result in coalescence of milk fat globules. Formation of free fatty acids increased significantly in both types of milk by pumping at 31 degrees C. Cooling the milk to 4 degrees C immediately before pumping inhibited an increased content of free fatty acids. However, when the milk was incubated at 4 degrees C for 60 min after cooling and then subjected to pumping, a significant increase in the formation of free fatty acids was observed in both types of milk. It is suggested that this increase in free fatty acids is caused by transition of polymorphic crystal forms or higher level of attached lipoprotein lipases to the milk fat globule before pumping.

  9. Alterations in the levels of plasma amino acids in polycystic ovary syndrome- A pilot study

    PubMed Central

    Unni, C. Sumithra N.; Lakshman, Lakshmi R.; Vaidyanathan, Kannan; Subhakumari, K.N.; Menon, N. Leela

    2015-01-01

    Background & objectives: Plasma amino acid levels are known to be altered in conditions like sepsis and burns which are situations of metabolic stress. Polycystic ovary syndrome (PCOS), a condition which affects a woman throughout her life, is said to be associated with metabolic stress. This study was undertaken to assess if there were significant alterations in the levels of plasma amino acids in women with PCOS. Methods: Sixty five women with PCOS along with the similar number of age matched normal controls were included in this study. Levels of 14 amino acids were determined using reverse phase high performance liquid chromatography. Results: The levels of methionine, cystine, isoleucine, phenylalanine, valine, tyrosine, proline, glycine, lysine and histidine were found to be significantly (P<0.001) lower in cases than in controls. Arginine and alanine levels were found to be significantly (P<0.001) higher in cases compared with controls. Interpretation & conclusions: Our findings showed significant derangement in the levels of plasma amino acids in women with PCOS which might be due to the oxidative and metabolic stress associated with it. Further studies need to be done to confirm the findings. PMID:26658589

  10. The environmental light influences the circulatory levels of retinoic acid and associates with hepatic lipid metabolism.

    PubMed

    Pang, Wenqiang; Li, Chunying; Zhao, Yue; Wang, Shiming; Dong, Wei; Jiang, Pengjiu; Zhang, Jianfa

    2008-12-01

    Environmental light is involved in the regulation of photochemical reaction in mouse retina. It remains unclear whether light-mediated increase in all-trans retinoic acid (ATRA) synthesis in retina will result in altering the circulatory levels of ATRA and regulating downstream gene expression and physiological function. Here we showed circulatory levels of ATRA decreased in mice under constant darkness and elevated by light exposure. Fat gene pancreatic lipase-related protein 2 (mPlrp2) and its partner procolipase (mClps), but not hepatic lipase (mHl), activated in livers for responding to lack of light illuminating. Light-triggered alterations in circulatory ATRA levels regulated ecto-5'-nucleotidase gene expression by retinoic acid receptor retinoic acid receptor-alpha and modulated 5'-AMP levels in blood and were associated with mPlrp2 and mClps expression in the livers. Mice deficient in adenosine receptors displayed mPlrp2 and mClps expression in livers under 12-h light, 12-h dark cycles. Caffeine blocked adenosine receptors and induced hepatic mPlrp2 and mClps expression in wild-type mice. Mice activated in hepatic mPlrp2 and mClps expression lowered hepatic and serum lipid levels and markedly elevated circulatory levels of all-trans retinol. Our results suggest environmental light influence hepatic lipid homeostasis by light-modulated retinoic acid signaling associated with mPlrp2 and mClps gene expression in livers.

  11. Ursolic acid reduces prostate size and dihydrotestosterone level in a rat model of benign prostatic hyperplasia.

    PubMed

    Shin, In-Sik; Lee, Mee-Young; Jung, Da-Young; Seo, Chang-Seob; Ha, Hye-Kyung; Shin, Hyeun-Kyoo

    2012-03-01

    Benign prostatic hyperplasia (BPH) is characterized by hyperplasia of prostatic stromal and epithelial cells, which can lead to lower urinary tract symptoms. The prevalence of BPH increases in an age-dependent manner. We investigated the protective effect of ursolic acid in BPH development using a testosterone-induced BPH rat model. BPH was induced in experimental groups by daily subcutaneous injections of testosterone propionate (TP), for a period of four weeks. Ursolic acid was administrated daily by oral gavage at a dose level of 5mg/kg during the four weeks of TP injections. Animals were sacrificed on the scheduled termination, before prostates were weighed and subjected to histopathological examination. TP and dihydrotestosterone (DHT) levels in the serum and prostate were also measured. BPH-induced animals displayed an increase in prostate weight with increased testosterone and DHT levels in both the serum and prostate. However, ursolic acid treatment resulted in significant reductions in prostate weight and testosterone and DHT levels in both the serum and prostate, compared with BPH-induced animals. Histopathological examination also showed that ursolic acid treatment suppressed TP-induced prostatic hyperplasia. These findings indicate that ursolic acid may effectively inhibit the development of BPH and it may be a useful agent in BPH treatment.

  12. Fatty acid status and its relationship to cognitive decline and homocysteine levels in the elderly.

    PubMed

    Baierle, Marília; Vencato, Patrícia H; Oldenburg, Luiza; Bordignon, Suelen; Zibetti, Murilo; Trentini, Clarissa M; Duarte, Marta M M F; Veit, Juliana C; Somacal, Sabrina; Emanuelli, Tatiana; Grune, Tilman; Breusing, Nicolle; Garcia, Solange C

    2014-09-12

    Polyunsaturated fatty acids (PUFAs), especially the n-3 series, are known for their protective effects. Considering that cardiovascular diseases are risk factors for dementia, which is common at aging, the aim of this study was to evaluate whether fatty acid status in the elderly was associated with cognitive function and cardiovascular risk. Forty-five elderly persons (age ≥ 60 years) were included and divided into two groups based on their Mini-Mental Status Examination score adjusted for educational level: the case group (n = 12) and the control group (n = 33). Serum fatty acid composition, homocysteine (Hcy), hs-CRP, lipid profile and different cognitive domains were evaluated. The case group, characterized by reduced cognitive performance, showed higher levels of 14:0, 16:0, 16:1n-7 fatty acids and lower levels of 22:0, 24:1n-9, 22:6n-3 (DHA) and total PUFAs compared to the control group (p < 0.05). The n-6/n-3 ratio was elevated in both study groups, whereas alterations in Hcy, hs-CRP and lipid profile were observed in the case group. Cognitive function was positively associated with the 24:1n-9, DHA and total n-3 PUFAs, while 14:0, 16:0 and 16:1n-7 fatty acids, the n-6/n-3 ratio and Hcy were inversely associated. In addition, n-3 PUFAs, particularly DHA, were inversely associated with cardiovascular risk, assessed by Hcy levels in the elderly.

  13. 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

  14. 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/.

  15. IDP-ASE: haplotyping and quantifying allele-specific expression at the gene and gene isoform level by hybrid sequencing.

    PubMed

    Deonovic, Benjamin; Wang, Yunhao; Weirather, Jason; Wang, Xiu-Jie; Au, Kin Fai

    2016-11-28

    Allele-specific expression (ASE) is a fundamental problem in studying gene regulation and diploid transcriptome profiles, with two key challenges: (i) haplotyping and (ii) estimation of ASE at the gene isoform level. Existing ASE analysis methods are limited by a dependence on haplotyping from laborious experiments or extra genome/family trio data. In addition, there is a lack of methods for gene isoform level ASE analysis. We developed a tool, IDP-ASE, for full ASE analysis. By innovative integration of Third Generation Sequencing (TGS) long reads with Second Generation Sequencing (SGS) short reads, the accuracy of haplotyping and ASE quantification at the gene and gene isoform level was greatly improved as demonstrated by the gold standard data GM12878 data and semi-simulation data. In addition to methodology development, applications of IDP-ASE to human embryonic stem cells and breast cancer cells indicate that the imbalance of ASE and non-uniformity of gene isoform ASE is widespread, including tumorigenesis relevant genes and pluripotency markers. These results show that gene isoform expression and allele-specific expression cooperate to provide high diversity and complexity of gene regulation and expression, highlighting the importance of studying ASE at the gene isoform level. Our study provides a robust bioinformatics solution to understand ASE using RNA sequencing data only.

  16. Genotyping-by-Sequencing-Based Investigation of the Genetic Architecture Responsible for a ∼Sevenfold Increase in Soybean Seed Stearic Acid

    PubMed Central

    Heim, Crystal B.; Gillman, Jason D.

    2016-01-01

    Soybean oil is highly unsaturated but oxidatively unstable, rendering it nonideal for food applications. Until recently, the majority of soybean oil underwent partial chemical hydrogenation, which produces trans fats as an unavoidable consequence. Dietary intake of trans fats and most saturated fats are conclusively linked to negative impacts on cholesterol levels and cardiovascular health. Two major soybean oil breeding targets are: (1) to reduce or eliminate the need for chemical hydrogenation, and (2) to replace the functional properties of partially hydrogenated soybean oil. One potential solution is the elevation of seed stearic acid, a saturated fat which has no negative impacts on cardiovascular health, from 3 to 4% in typical cultivars to > 20% of the seed oil. We performed QTL analysis of a population developed by crossing two mutant lines, one with a missense mutation affecting a stearoyl-acyl-carrier protein desaturase gene resulting in ∼11% seed stearic acid crossed to another mutant, A6, which has 24–28% seed stearic acid. Genotyping-by-sequencing (GBS)-based QTL mapping identified 21 minor and major effect QTL for six seed oil related traits and plant height. The inheritance of a large genomic deletion affecting chromosome 14 is the basis for largest effect QTL, resulting in ∼18% seed stearic acid. This deletion contains SACPD-C and another gene(s); loss of both genes boosts seed stearic acid levels to ≥ 18%. Unfortunately, this genomic deletion has been shown in previous studies to be inextricably correlated with reduced seed yield. Our results will help inform and guide ongoing breeding efforts to improve soybean oil oxidative stability. PMID:27866151

  17. Genotyping-by-Sequencing-Based Investigation of the Genetic Architecture Responsible for a ∼Sevenfold Increase in Soybean Seed Stearic Acid.

    PubMed

    Heim, Crystal B; Gillman, Jason D

    2017-01-05

    Soybean oil is highly unsaturated but oxidatively unstable, rendering it nonideal for food applications. Until recently, the majority of soybean oil underwent partial chemical hydrogenation, which produces trans fats as an unavoidable consequence. Dietary intake of trans fats and most saturated fats are conclusively linked to negative impacts on cholesterol levels and cardiovascular health. Two major soybean oil breeding targets are: (1) to reduce or eliminate the need for chemical hydrogenation, and (2) to replace the functional properties of partially hydrogenated soybean oil. One potential solution is the elevation of seed stearic acid, a saturated fat which has no negative impacts on cardiovascular health, from 3 to 4% in typical cultivars to > 20% of the seed oil. We performed QTL analysis of a population developed by crossing two mutant lines, one with a missense mutation affecting a stearoyl-acyl-carrier protein desaturase gene resulting in ∼11% seed stearic acid crossed to another mutant, A6, which has 24-28% seed stearic acid. Genotyping-by-sequencing (GBS)-based QTL mapping identified 21 minor and major effect QTL for six seed oil related traits and plant height. The inheritance of a large genomic deletion affecting chromosome 14 is the basis for largest effect QTL, resulting in ∼18% seed stearic acid. This deletion contains SACPD-C and another gene(s); loss of both genes boosts seed stearic acid levels to ≥ 18%. Unfortunately, this genomic deletion has been shown in previous studies to be inextricably correlated with reduced seed yield. Our results will help inform and guide ongoing breeding efforts to improve soybean oil oxidative stability.

  18. The human erythrocyte anion-transport protein. Partial amino acid sequence, conformation and a possible molecular mechanism for anion exchange.

    PubMed Central

    Brock, C J; Tanner, M J; Kempf, C

    1983-01-01

    The N-terminal 72 residues of an integral membrane fragment, P5, of the human erythrocyte anion-transport protein, which is known to be directly involved in the anion-exchange process, was shown to have the following amino acid sequence: Met-Val-Pro-Lys-Pro-Gln-Gly-Pro-Leu-Pro-Asn-Thr-Ala-Leu-Leu-Ser-Leu-Val-Leu-Met -Ala-Gly-Thr-Phe-Phe-Phe-Ala-Met-Met-Leu-Arg-Lys-Phe-Lys-Asn-Ser-Ser-Tyr-Phe-Pro-Gly-Lys-Leu-Arg-Arg-Val-Ile-Gly-Asp-Phe-Gly-Val-Pro-Ile-Ser-Ile-Leu-Ile-Met-Val-Leu-Val-Asp-Phe-Phe-Ile-Gln-Asp-Thr-Tyr-Thr-Gln- The structure of this fragment was analysed, with account being taken of the constraints that apply to the folding of integral membrane proteins and the topographical locations of various sites in the sequence. It was concluded that this sequence forms two transmembrane alpha-helices. These are probably part of a cluster of amphipathic transmembrane alpha-helices, which could comprise that part of the protein responsible for transport activity. The presently available evidence relating to the anion-exchange process was considered with the structural features noted in this study and a possible molecular mechanism is proposed. In this model the rearrangement of a network of intramembranous charged pairs mediates the translocation of an anion between anion-binding regions at each surface of the membrane, which are composed of clusters of positively charged amino acids. This model imposes a sequential exchange mechanism on the system. Supplementary material, including Tables and Figures describing the compositions of peptides determined by amino acid analysis and sequence studies, quantitative and qualitative data that provide a residue-by-residue justification for the sequence assignment and a description of modifications to and use of the solid-phase sequencer has been deposited as Supplementary Publication SUP 50123 (12 pages) with the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be

  19. Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2230 from Karijini National Park, Australia

    PubMed Central

    Walker, Robert; Watkin, Elizabeth; Tian, Rui; Bräu, Lambert; O’Hara, Graham; Goodwin, Lynne; Han, James; Lobos, Elizabeth; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavromatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

    2013-01-01

    Burkholderia sp. strain WSM2230 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod isolated from acidic soil collected in 2001 from Karijini National Park, Western Australia, using Kennedia coccinea (Coral Vine) as a host. WSM2230 was initially effective in nitrogen-fixation with K. coccinea, but subsequently lost symbiotic competence. Here we describe the features of Burkholderia sp. strain WSM2230, together with genome sequence information and its annotation. The 6,309,801 bp high-quality-draft genome is arranged into 33 scaffolds of 33 contigs containing 5,590 protein-coding genes and 63 RNA-only encoding genes. The genome sequence of WSM2230 failed to identify nodulation genes and provides an explanation for the observed failure of the laboratory grown strain to nodulate. The genome of this strain 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:25197440

  20. Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2230 from Karijini National Park, Australia.

    PubMed

    Walker, Robert; Watkin, Elizabeth; Tian, Rui; Bräu, Lambert; O'Hara, Graham; Goodwin, Lynne; Han, James; Lobos, Elizabeth; Huntemann, Marcel; Pati, Amrita; Woyke, Tanja; Mavromatis, Konstantinos; Markowitz, Victor; Ivanova, Natalia; Kyrpides, Nikos; Reeve, Wayne

    2014-06-15

    Burkholderia sp. strain WSM2230 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod isolated from acidic soil collected in 2001 from Karijini National Park, Western Australia, using Kennedia coccinea (Coral Vine) as a host. WSM2230 was initially effective in nitrogen-fixation with K. coccinea, but subsequently lost symbiotic competence. Here we describe the features of Burkholderia sp. strain WSM2230, together with genome sequence information and its annotation. The 6,309,801 bp high-quality-draft genome is arranged into 33 scaffolds of 33 contigs containing 5,590 protein-coding genes and 63 RNA-only encoding genes. The genome sequence of WSM2230 failed to identify nodulation genes and provides an explanation for the observed failure of the laboratory grown strain to nodulate. The genome of this strain 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.

  1. Cloning, sequence analysis and expression of the F1F0-ATPase beta-subunit from wine lactic acid bacteria.

    PubMed

    Sievers, Martin; Uermösi, Christina; Fehlmann, Marc; Krieger, Sibylle

    2003-09-01

    The nucleotide sequences of the genes encoding the F1F0-ATPase beta-subunit from Oenococcus oeni, Leuconostoc mesenteroides subsp. mesenteroides, Pediococcus damnosus, Pediococcus parvulus, Lactobacillus brevis and Lactobacillus hilgardii were determined. Their deduced amino acid sequences showed homology values of 79-98%. Data from the alignment and ATPase tree indicated that O. oeni and L. mesenteroides subsp. mesenteroides formed a group well-separated from P. damnosus and P. parvulus and from the group comprises L. brevis and L. hilgardii. The N-terminus of the F1F0-ATPase beta-subunit of O. oeni contains a stretch of additional 38 amino acid residues. The catalytic site of the ATPase beta-subunit of the investigated strains is characterized by the two conserved motifs GGAGVGKT and GERTRE. The amplified atpD coding sequences were inserted into the pCRT7/CT-TOPO vector using TA-cloning strategy and transformed in Escherichia coli. SDS-PAGE and Western blot analyses confirmed that O. oeni has an ATPase beta-subunit protein which is larger in size than the corresponding molecules from the investigated strains.

  2. 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

  3. Modification of the repeated acquisition of response sequences in rats by low-level microwave exposure

    SciTech Connect

    Schrot, J.; Thomas, J.R.; Banvard, R.A.

    1980-01-01

    The acute effects of microwave exposure on a repeated acquisition baseline were investigated in three rats. Each session the animals acquired a different four-member response sequence. Each of the first three correct responses advanced the sequence to the next member, and the fourth correct response produced food reinforcement. Incorrect responses produced a three-second timeout. Baseline and control sessions were characterized by a decrease in errors within each session. The animals were acutely exposed to a 2.8 GHz pulsed-microwave field prior to test sessions, with average power densities ranging from 0.25 to to 10 mW/cm2. In comparison to control sessions, 1/2 hour of exposure to microwave radiation at power densities of 5 and 10 mW/cm2 increased errors and altered the pattern of within-session acquisition. Exposure to the 10 mW/cm2 power density decreased the rate of sequence completion in all animals. The results of exposures at 0.25, 0.5, and 1 mW/cm2 power densities were generally within the control range. The results are interpreted as indicating a disruption in the discriminative stimulus control of the repeated acquisition behavior.

  4. Mapping vaccinia virus DNA replication origins at nucleotide level by deep sequencing.

    PubMed

    Senkevich, Tatiana G; Bruno, Daniel; Martens, Craig; Porcella, Stephen F; Wolf, Yuri I; Moss, Bernard

    2015-09-01

    Poxviruses reproduce in the host cytoplasm and encode most or all of the enzymes and factors needed for expression and synthesis of their double-stranded DNA genomes. Nevertheless, the mode of poxvirus DNA replication and the nature and location of the replication origins remain unknown. A current but unsubstantiated model posits only leading strand synthesis starting at a nick near one covalently closed end of the genome and continuing around the other end to generate a concatemer that is subsequently resolved into unit genomes. The existence of specific origins has been questioned because any plasmid can replicate in cells infected by vaccinia virus (VACV), the prototype poxvirus. We applied directional deep sequencing of short single-stranded DNA fragments enriched for RNA-primed nascent strands isolated from the cytoplasm of VACV-infected cells to pinpoint replication origins. The origins were identified as the switching points of the fragment directions, which correspond to the transition from continuous to discontinuous DNA synthesis. Origins containing a prominent initiation point mapped to a sequence within the hairpin loop at one end of the VACV genome and to the same sequence within the concatemeric junction of replication intermediates. These findings support a model for poxvirus genome replication that involves leading and lagging strand synthesis and is consistent with the requirements for primase and ligase activities as well as earlier electron microscopic and biochemical studies implicating a replication origin at the end of the VACV genome.

  5. "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.

  6. High-level expression of alternative oxidase protein sequences enhances the spread of viral vectors in resistant and susceptible plants.

    PubMed

    Murphy, Alex M; Gilliland, Androulla; York, Caroline J; Hyman, Belinda; Carr, John P

    2004-12-01

    The alternative oxidase (AOX) is the terminal oxidase of the cyanide-resistant alternative respiratory pathway in plants and has been implicated in resistance to viruses. When tobacco mosaic virus (TMV) vectors were used to drive very high levels of expression of either AOX or AOX mutated in its active site (AOX-E), virus spread was enhanced. This was visualized as the induction of larger hypersensitive-response lesions after inoculation onto NN-genotype tobacco than those produced by vectors bearing sequences of comparable length [the green fluorescent protein (gfp) gene sequence or antisense aox] or the 'empty' viral vector. Also, in the highly susceptible host Nicotiana benthamiana, systemic movement of TMV vectors expressing AOX or AOX-E was faster than that of TMV constructs bearing gfp or antisense aox sequences. Notably, in N. benthamiana, TMV.AOX and TMV.AOX-E induced symptoms that were severe and ultimately included cell death, whereas the empty vector, TMV.GFP and the TMV vector expressing antisense aox sequences never induced necrosis. The results show that, if expressed at sufficiently high levels, active and inactive AOX proteins can affect virus spread and symptomology in plants.

  7. Defining sequence space and reaction products within the cyanuric acid hydrolase (AtzD)/barbiturase protein family.

    PubMed

    Seffernick, Jennifer L; Erickson, Jasmine S; Cameron, Stephan M; Cho, Seunghee; Dodge, Anthony G; Richman, Jack E; Sadowsky, Michael J; Wackett, Lawrence P

    2012-09-01

    Cyanuric acid hydrolases (AtzD) and barbiturases are homologous, found almost exclusively in bacteria, and comprise a rare protein family with no discernible linkage to other protein families or an X-ray structural class. There has been confusion in the literature and in genome projects regarding the reaction products, the assignment of individual sequences as either cyanuric acid hydrolases or barbiturases, and spurious connection of this family to another protein family. The present study has addressed those issues. First, the published enzyme reaction products of cyanuric acid hydrolase are incorrectly identified as biuret and carbon dioxide. The current study employed (13)C nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry to show that cyanuric acid hydrolase releases carboxybiuret, which spontaneously decarboxylates to biuret. This is significant because it revealed that homologous cyanuric acid hydrolases and barbiturases catalyze completely analogous reactions. Second, enzymes that had been annotated incorrectly in genome projects have been reassigned here by bioinformatics, gene cloning, and protein characterization studies. Third, the AtzD/barbiturase family has previously been suggested to consist of members of the amidohydrolase superfamily, a large class of metallohydrolases. Bioinformatics and the lack of bound metals both argue against a connection to the amidohydrolase superfamily. Lastly, steady-state kinetic measurements and observations of protein stability suggested that the AtzD/barbiturase family might be an undistinguished protein family that has undergone some resurgence with the recent introduction of industrial s-triazine compounds such as atrazine and melamine into the environment.

  8. 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

  9. The shikimate pathway: review of amino acid sequence, function and three-dimensional structures of the enzymes.

    PubMed

    Mir, Rafia; Jallu, Shais; Singh, T P

    2015-06-01

    The aromatic compounds such as aromatic amino acids, vitamin K and ubiquinone are important prerequisites for the metabolism of an organism. All organisms can synthesize these aromatic metabolites through shikimate pathway, except for mammals which are dependent on their diet for these compounds. The pathway converts phosphoenolpyruvate and erythrose 4-phosphate to chorismate through seven enzymatically catalyzed steps and chorismate serves as a precursor for the synthesis of variety of aromatic compounds. These enzymes have shown to play a vital role for the viability of microorganisms and thus are suggested to present attractive molecular targets for the design of novel antimicrobial drugs. This review focuses on the seven enzymes of the shikimate pathway, highlighting their primary sequences, functions and three-dimensional structures. The understanding of their active site amino acid maps, functions and three-dimensional structures will provide a framework on which the rational design of antimicrobial drugs would be based. Comparing the full length amino acid sequences and the X-ray crystal structures of these enzymes from bacteria, fungi and plant sources would contribute in designing a specific drug and/or in developing broad-spectrum compounds with efficacy against a variety of pathogens.

  10. Nucleotide sequence and spatial expression pattern of a drought- and abscisic Acid-induced gene of tomato.

    PubMed

    Plant, A L; Cohen, A; Moses, M S; Bray, E A

    1991-11-01

    The nucleotide sequence of le16, a tomato (Lycopersicon esculentum Mill.) gene induced by drought stress and regulated by abscisic acid specifically in aerial vegetative tissue, is presented. The single open reading frame contained within the gene has the capacity to encode a polypeptide of 12.7 kilodaltons and is interrupted by a small intron. The predicted polypeptide is rich in leucine, glycine, and alanine and has an isoelectric point of 8.7. The amino terminus is hydrophobic and characteristic of signal sequences that target polypeptides for export from the cytoplasm. There is homology (47.2% identity) between the amino terminus of the LE 16 polypeptide and the corresponding amino terminal domain of the maize phospholipid transfer protein. le16 was expressed in drought-stressed leaf, petiole, and stem tissue and to a much lower extent in the pericarp of mature green tomato fruit and developing seeds. No expression was detected in the pericarp of red fruit or in drought-stressed roots. Expression of le16 was also induced in leaf tissue by a variety of other abiotic stresses including polyethylene glycol-mediated water deficit, salinity, cold stress, and heat stress. None of these stresses or direct applications of abscisic acid induced the expression of le16 in the roots of the same plants. The unique expression characteristics of this gene indicates that novel regulatory mechanisms, in addition to endogenous abscisic acid, are involved in controlling gene expression.

  11. Eastern Mediterranean sea levels through the last interglacial from a coastal-marine sequence in northern Israel

    NASA Astrophysics Data System (ADS)

    Sivan, D.; Sisma-Ventura, G.; Greenbaum, N.; Bialik, O. M.; Williams, F. H.; Tamisiea, M. E.; Rohling, E. J.; Frumkin, A.; Avnaim-Katav, S.; Shtienberg, G.; Stein, M.

    2016-08-01

    A last interglacial (Marine Isotope Stage, MIS5e) marine-coastal sequence has been identified along the Galilee coast of Israel, with the type section located at Rosh Hanikra (RH). The microtidal regime and tectonic stability, along with the detailed stratigraphy of the RH shore, make the study region ideally suited for determining relative sea level (RSL) through the MIS5e interval in the eastern Mediterranean. The sequence contains fossilized microtidal subunits at a few meters above the current sea level. Unfortunately, all fossils were found to be altered, so that U-Th datings cannot be considered to represent initial deposition. We contend that U-Th dating of Strombus bubonius shells (recrystallized to calcite) suffices to indicate a lower limit of ∼110 ± 8 ka for the time sea level dropped below the RH sedimentary sequence. The RH-section comprises three main subunits of a previously determined member (the Yasaf Member): (a) a gravelly unit containing the diagnostic gastropod Strombus bubonius Lamarck (Persististrombus latus), which was deposited in the intertidal to super-tidal stormy zone; (b) Vermetidae reef domes indicating a shallow-water depositional environment; and (c) coarse to medium-sized, bioclastic sandstone, probably deposited in the shallow sub-tidal zone. The sequence overlies three abrasion platforms that are cut by tidal channels at elevations of +0.8 m, +2.6 m and +3.4 m, and which are filled with MIS5e sediments. We present a detailed study of the sequence, with emphasis on stratigraphic, sedimentological, and palaeontological characteristics that indicate sea-level changes. Although without precise absolute dating, the stratigraphic sequence of RH through MIS5e allows us to identify a time-series of RSL positions, using the elevations of three stratigraphic subunits. Reconstructed RSL values range from +1.0 m to +7 m (with uncertainly < 1 m), and most fall within a narrow range of +1.0 to +3.3 m. Toward the end of MIS5e, RSL exceeded

  12. 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...

  13. Circulating irisin levels are associated with lipid and uric acid metabolism in a Chinese population.

    PubMed

    Tang, Shanshan; Zhang, Rong; Jiang, Feng; Wang, Jie; Chen, Miao; Peng, Danfeng; Yan, Jing; Wang, Shiyun; Bao, Yuqian; Hu, Cheng; Jia, Weiping

    2015-06-26

    Irisin is a novel hormone secreted by skeletal muscle after exercise, which may ameliorate insulin resistance. In this study, we aimed to explore the relationship between circulating irisin levels and type 2 diabetes (T2DM) as well as related metabolic traits in a Chinese population. A total of 203 subjects were recruited. Of these, 68 subjects with NGT, 63 subjects with IGR and 72 subjects with new-onset T2DM. Circulating irisin levels were measured by ELISA. Detailed clinical investigations and biochemistry measurements were carried out in all of the subjects. Multivariate linear regression analysis was performed to assess the association between irisin levels and related metabolic characteristics. All subjects were classified into normal weight and overweight/obese subgroups according to body mass index (BMI). No significant differences in circulating irisin levels were identified among the three groups (p=0.9741). After adjusting for covariates, multiple linear regression analysis revealed that serum irisin level was independently and significantly associated with total cholesterol (p=0.0005), low-density lipoprotein cholesterol (p=0.0014), fasting fatty acids (p=0.0402) and uric acid (p=0.0062). By dividing the serum irisin levels into three tertile group, the values of total cholesterol, low-density lipoprotein cholesterol, fasting fatty acids and uric acid were all increased significantly with the increase of irisin (p<0.05) . Moreover, serum irisin levels remain closely related to total cholesterol in both normal weight and overweight/obese subgroups. Our study suggests that circulating irisin concentrations are significantly associated with lipid and uric acid metabolism in a Chinese population. This article is protected by copyright. All rights reserved.

  14. A reliable and sensitive bead-based fluorescence assay for identification of nucleic acid sequences

    NASA Astrophysics Data System (ADS)

    Klamp, Tobias; Yahiatène, Idir; Lampe, André; Schüttpelz, Mark; Sauer, Markus

    2011-03-01

    The sensitive and rapid detection of pathogenic DNA is of tremendous importance in the field of diagnostics. We demonstrate the ability of detecting and quantifying single- and double-stranded pathogenic DNA with picomolar sensitivity in a bead-based fluorescence assay. Selecting appropriate capturing and detection sequences enables rapid (2 h) and reliable DNA quantification. We show that synthetic sequences of S. pneumoniae and M. luteus can be quantified in very small sample volumes (20 μL) across a linear detection range over four orders of magnitude from 1 nM to 1 pM, using a miniaturized wide-field fluorescence microscope without amplification steps. The method offers single molecule detection sensitivity without using complex setups and thus volunteers as simple, robust, and reliable method for the sensitive detection of DNA and RNA sequences.

  15. Chromosomal-Level Assembly of the Asian Seabass Genome Using Long Sequence Reads and Multi-layered Scaffolding.

    PubMed

    Vij, Shubha; Kuhl, Heiner; Kuznetsova, Inna S; Komissarov, Aleksey; Yurchenko, Andrey A; Van Heusden, Peter; Singh, Siddharth; Thevasagayam, Natascha M; Prakki, Sai Rama Sridatta; Purushothaman, Kathiresan; Saju, Jolly M; Jiang, Junhui; Mbandi, Stanley Kimbung; Jonas, Mario; Hin Yan Tong, Amy; Mwangi, Sarah; Lau, Doreen; Ngoh, Si Yan; Liew, Woei Chang; Shen, Xueyan; Hon, Lawrence S; Drake, James P; Boitano, Matthew; Hall, Richard; Chin, Chen-Shan; Lachumanan, Ramkumar; Korlach, Jonas; Trifonov, Vladimir; Kabilov, Marsel; Tupikin, Alexey; Green, Darrell; Moxon, Simon; Garvin, Tyler; Sedlazeck, Fritz J; Vurture, Gregory W; Gopalapillai, Gopikrishna; Kumar Katneni, Vinaya; Noble, Tansyn H; Scaria, Vinod; Sivasubbu, Sridhar; Jerry, Dean R; O'Brien, Stephen J; Schatz, Michael C; Dalmay, Tamás; Turner, Stephen W; Lok, Si; Christoffels, Alan; Orbán, László

    2016-04-01

    We report here the ~670 Mb genome assembly of the Asian seabass (Lates calcarifer), a tropical marine teleost. We used long-read sequencing augmented by transcriptomics, optical and genetic mapping along with shared synteny from closely related fish species to derive a chromosome-level assembly with a contig N50 size over 1 Mb and scaffold N50 size over 25 Mb that span ~90% of the genome. The population structure of L. calcarifer species complex was analyzed by re-sequencing 61 individuals representing various regions across the species' native range. SNP analyses identified high levels of genetic diversity and confirmed earlier indications of a population stratification comprising three clades with signs of admixture apparent in the South-East Asian population. The quality of the Asian seabass genome assembly far exceeds that of any other fish species, and will serve as a new standard for fish genomics.

  16. Chromosomal-Level Assembly of the Asian Seabass Genome Using Long Sequence Reads and Multi-layered Scaffolding

    PubMed Central

    Vij, Shubha; Kuhl, Heiner; Kuznetsova, Inna S.; Komissarov, Aleksey; Yurchenko, Andrey A.; Van Heusden, Peter; Singh, Siddharth; Thevasagayam, Natascha M.; Prakki, Sai Rama Sridatta; Purushothaman, Kathiresan; Saju, Jolly M.; Jiang, Junhui; Mbandi, Stanley Kimbung; Jonas, Mario; Hin Yan Tong, Amy; Mwangi, Sarah; Lau, Doreen; Ngoh, Si Yan; Liew, Woei Chang; Shen, Xueyan; Hon, Lawrence S.; Drake, James P.; Boitano, Matthew; Hall, Richard; Chin, Chen-Shan; Lachumanan, Ramkumar; Korlach, Jonas; Trifonov, Vladimir; Kabilov, Marsel; Tupikin, Alexey; Green, Darrell; Moxon, Simon; Garvin, Tyler; Sedlazeck, Fritz J.; Vurture, Gregory W.; Gopalapillai, Gopikrishna; Kumar Katneni, Vinaya; Noble, Tansyn H.; Scaria, Vinod; Sivasubbu, Sridhar; Jerry, Dean R.; O'Brien, Stephen J.; Schatz, Michael C.; Dalmay, Tamás; Turner, Stephen W.; Lok, Si; Christoffels, Alan; Orbán, László

    2016-01-01

    We report here the ~670 Mb genome assembly of the Asian seabass (Lates calcarifer), a tropical marine teleost. We used long-read sequencing augmented by transcriptomics, optical and genetic mapping along with shared synteny from closely related fish species to derive a chromosome-level assembly with a contig N50 size over 1 Mb and scaffold N50 size over 25 Mb that span ~90% of the genome. The population structure of L. calcarifer species complex was analyzed by re-sequencing 61 individuals representing various regions across the species’ native range. SNP analyses identified high levels of genetic diversity and confirmed earlier indications of a population stratification comprising three clades with signs of admixture apparent in the South-East Asian population. The quality of the Asian seabass genome assembly far exceeds that of any other fish species, and will serve as a new standard for fish genomics. PMID:27082250

  17. Correlation of pncA sequence with pyrazinamide resistance level in BACTEC for 21 mycobacterium tuberculosis clinical isolates.

    PubMed

    Mestdagh, M; Realini, L; Fonteyne, P A; Rossau, R; Jannes, G; Mijs, W; DE Smet, K A; Portaels, F; Van den Eeckhout, E

    2000-01-01

    Mutations in the pncA gene, encoding pyrazinamidase, are considered the major mechanism of pyrazinamide (PZA) resistance in Mycobacterium tuberculosis, but resistant strains containing the wild-type gene have been described. The correlation of pncA sequence with PZA resistance level was examined for 21 M. tuberculosis clinical isolates. Susceptibility patterns were determined for 100, 300, and 900 microg/ml concentrations of the drug in BACTEC. Insertions and deletions and a substitution in the putative promoter region led to high-level resistance, whereas substitutions within the open reading frame seemed to confer variable levels of resistance. Variable resistance levels and PZase activities were also observed among isolates lacking pncA mutations. The high-level resistance (900 microg/ml) in pncA wild-type isolates highlights the clinical significance of these isolates. These data also suggest that there may still be more than one alternative mechanism leading to PZA resistance in M. tuberculosis isolates.

  18. Serum uric acid levels and long-term outcomes in chronic kidney disease.

    PubMed

    Miyaoka, Tokiko; Mochizuki, Toshio; Takei, Takashi; Tsuchiya, Ken; Nitta, Kosaku

    2014-07-01

    Hyperuricemia is common in chronic kidney disease (CKD), but data regarding the relationship between serum uric acid levels and the long-term outcomes of CKD patients have been limited. The present study evaluated the associations between baseline serum uric acid levels with mortality and end-stage renal disease (ESRD). The subjects of this study were 551 stage 2-4 CKD patients. Cox proportional hazards models were used to evaluate the relationship between serum uric acid tertiles and all-cause mortality, cardiovascular disease (CVD) mortality, 50 % reduction in estimated glomerular filtration rate (eGFR), and development of ESRD, initially without adjustment, and then after adjusting for several groups of covariates. The mean age of the study subjects was 58.5 years, 59.3 % were men, and 10.0 % had diabetes. The mean eGFR was 42.02 ± 18.52 ml/min/1.73 m(2). In all subjects, the mean serum uric acid level was 6.57 ± 1.35 mg/dl, and 52.2 % of study subjects were on hypouricemic therapy (allopurinol; 48.3 %) at baseline. Thirty-one patients (6.1 %) died during a follow-up period of approximately 6 years. There was no significant association between serum uric acid level and all-cause mortality, CVD mortality, development of ESRD and 50 % reduction in eGFR in the unadjusted Cox models. In the adjusted models, hyperuricemia was found to be associated with all-cause mortality and CVD mortality after adjustment with CVD risk factors, kidney disease factors, and allopurinol, but not associated with development of ESRD and 50 % reduction in eGFR. The results of this study showed that hyperuricemia but not serum uric acid levels were associated with all-cause mortality, CVD mortality after adjustments with CVD risk factors, kidney disease factors, and allopurinol in stage 2-4 CKD patients.

  19. Establishing tolerable dungeness crab (Cancer magister) and razor clam (Siliqua patula) domoic acid contaminant levels.

    PubMed

    Mariën, K

    1996-11-01

    Domoic acid has been found in razor clams (Siliqua patula) and dungeness crabs (Cancer magister) in Washington State and elsewhere on the West Coast of the United States. Due to toxic effects associated with domoic acid exposure, an effort has been made to establish tolerable domoic acid levels in crabs and clams obtained from commercial harvest and sale and from individual recreational harvesting. To accomplish this, the amount of clams and crabs consumed by populations of concern was determined, a tolerable daily intake (TDI) was developed for individuals most sensitive to effects of this compound, and the TDI was equated with consumption patterns to determine tolerable clam and crab domoic acid levels. Results indicate that the primary health effects associated with domoic acid toxicity can be averted in populations of concern and for others consuming crabs or clams less frequently (or in lesser quantity) if domoic acid contaminant concentration does not exceed 30 mg/kg in the hepatopancreas and viscera of dungeness crabs or 20 mg/kg in clams.

  20. Pyruvic acid levels in serum and saliva: A new course for oral cancer screening?

    PubMed Central

    Bhat, Manohara A; Prasad, KVV; Trivedi, Dheeraj; Rajeev, BR; Battur, Hemanth

    2016-01-01

    Objective: Cancerous cells show increased glycolysis rate. This will increase overall levels of pyruvate as it is one of the end products of glycolysis. The present on-going study is to estimate the levels of pyruvate in saliva and serum among healthy and oral cancer subjects. Settings and Design: Hospital-based cross-sectional comparative study. Methodology: A total of 50 subjects among healthy and oral cancer subjects were selected based on clinical and histological criteria. Saliva and serum samples were collected and subjected to pyruvate level estimation using biochemical analysis. Statistical Analysis: Descriptive analysis and Mann-Whitney test were used to find the statistical difference between the two independent groups. Results: Serum pyruvic acid levels of the healthy group were 1.09 ± 0.14 and for oral cancer, it was 2.95 ± 0.59 and salivary level were 3.49 ± 0.47 and 1.32 ± 0.10 respectively. Mann-Whitney test showed statistically significant difference in serum and salivary pyruvate level in between two groups (P < 0.000 respectively). Conclusion: The present study showed noticeable variation in the level of pyruvic acid among healthy and oral cancer subjects. This generates the hypothesis that estimation of the pyruvic acid can be a new tool to screening of the cancer. PMID:27194870

  1. Effect of folic acid and zinc sulphate on endocrine parameters and seminal antioxidant level after varicocelectomy.

    PubMed

    Nematollahi-Mahani, S N; Azizollahi, G H; Baneshi, M R; Safari, Z; Azizollahi, S

    2014-04-01

    Varicocele is among the most common problems which may lead to male infertility. Spermatogenesis is impaired as a consequence of this vascular defect, through mechanisms that are not well described. This study aimed to evaluate serum hormonal level (inhibin B, FSH and testosterone) and seminal plasma antioxidant defence levels after folic acid and zinc sulphate administration in varicocelectomised patients. Participants were randomly allocated to four experimental groups. Our randomisation schedule was as follows: zinc sulphate/folic acid, folic acid, zinc sulphate and placebo. The patients underwent varicocelectomy, before which a blood and semen sample were obtained and also three and six months after varicocelectomy for evaluation of blood hormonal level (FSH, testosterone, inhibin B) and seminal oxidative stress status (nitric oxide, superoxide dismutase, total antioxidant capacity). Patients in different groups took orally one capsule per day after dinner following varicocelectomy for 6 months. A significant rise in peripheral blood inhibin B and seminal plasma activity was detected in the zinc sulphate/folic acid group after 6 months. The present clinical trial indicates a change in the hormonal status of varicocelectomised patients following long-term administration of zinc sulphate and folic acid.

  2. Salicylic Acid Treatment Increases the Levels of Triterpene Glycosides in Black Cohosh (Actaea Racemosa) Rhizomes.

    PubMed

    De Capite, Annette; Lancaster, Tyler; Puthoff, David

    2016-01-01

    Black cohosh (Actaea racemosa) serves as the host plant for the Appalachian azure butterfly, Celastrina neglectamajor. Overharvesting of Black cohosh for the dietary supplement industry may result in its extirpation, and may also cause the elimination of the dependent butterfly. One way to increase or maintain the number of host plants in forested environments would be to reduce the number harvested, for example by increasing the levels of the desired metabolites in Black cohosh rhizomes. The secondary metabolites actein and deoxyactein are triterpene glycosides and are among the compounds associated with the putative activity of Black cohosh extracts. Acetein and deoxyacetein are used to standardize Black cohosh supplements. To gain an understanding of mechanisms that may control actein and deoxyactein accumulation, Black cohosh rhizomes were treated with exogenous salicylic acid, jasmonic acid, or ethylene, or were mechanically wounded. Salicylic acid treatment significantly increased the levels of actein and deoxyactein in the rhizome of Black cohosh, suggesting that the synthesis of triterpene glycosides is controlled in part by salicylic acid. Using salicylic acid or related chemicals to increase the levels of actein and deoxyactein in rhizomes may help supply the supplement industry and, simultaneously, help conserve Black cohosh and species dependent upon it.

  3. Lactose synthesis in a monotreme, the echidna (Tachyglossus aculeatus): isolation and amino acid sequence of echidna alpha-lactalbumin.

    PubMed

    Messer, M; Griffiths, M; Rismiller, P D; Shaw, D C

    1997-10-01

    alpha-Lactalbumin and lysozyme were each isolated from echidna (Tachyglossus aculeatus) milk by gel permeation and ion exchange chromatography. The alpha-lactalbumin modified the action of echidna milk galactosyltransferase to promote the synthesis of lactose but had very little effect on bovine galactosyltransferase. Echidna alpha-lactalbumin is a glycosylated protein with an apparent molecular weight of 20,000 (SDS-PAGE) whose concentration in the milk is very low compared with the concentrations of alpha-lactalbumin in the milk of other species. Its amino acid sequence is more similar to that of another monotreme, the platypus (Ornithorhynchus anatinus), than to the sequences of eutherian or marsupial alpha-lactalbumins. Echidna milk lysozyme, even at high concentrations, did not promote the synthesis of lactose by either echidna or bovine galactosyltransferase. We conclude that lactose synthesis in the echidna occurs by the same mechanism as that found in the platypus and other mammals.

  4. 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