Isolation and deduced amino acid sequence of the gene encoding gp115, a yeast glycophospholipid-anchored protein containing a serine-rich region

ABSTRACT Here, we report the identification of the gene encoding a novel enzyme, 3-(5-oxo-2-thioxoimidazolidin-4-yl) propionic acid desulfhydrase, in Burkholderia sp. HME13. The enzyme converts 3-(5-oxo-2-thioxoimidazolidin-4-yl) propionic acid and H2O to 3-(2,5-dioxoimidazolidin-4-yl) propionic acid and H2S. Amino acid sequence analysis of the enzyme indicates that it belongs to the DUF917 protein family, which consists of proteins of unknown function.

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Catalytic and Molecular Properties of the Quinohemoprotein Tetrahydrofurfuryl Alcohol Dehydrogenase from Ralstonia eutropha Strain Bo

Journal of Bacteriology ◽

10.1128/jb.183.6.1954-1960.2001 ◽

2001 ◽

Vol 183 (6) ◽

pp. 1954-1960 ◽

Cited By ~ 12

Author(s):

Grit Zarnt ◽

Thomas Schräder ◽

Jan R. Andreesen

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Alcohol Dehydrogenase ◽

Tertiary Structure ◽

Ralstonia Eutropha ◽

Signal Sequence ◽

Substrate Inhibition ◽

Catalytic Efficiency ◽

Gene Encoding

ABSTRACT The quinohemoprotein tetrahydrofurfuryl alcohol dehydrogenase (THFA-DH) from Ralstonia eutropha strain Bo was investigated for its catalytic properties. The apparentk cat/Km andK i values for several substrates were determined using ferricyanide as an artificial electron acceptor. The highest catalytic efficiency was obtained with n-pentanol exhibiting a k cat/Km value of 788 × 104 M−1 s−1. The enzyme showed substrate inhibition kinetics for most of the alcohols and aldehydes investigated. A stereoselective oxidation of chiral alcohols with a varying enantiomeric preference was observed. Initial rate studies using ethanol and acetaldehyde as substrates revealed that a ping-pong mechanism can be assumed for in vitro catalysis of THFA-DH. The gene encoding THFA-DH from R. eutropha strain Bo (tfaA) has been cloned and sequenced. The derived amino acid sequence showed an identity of up to 67% to the sequence of various quinoprotein and quinohemoprotein dehydrogenases. A comparison of the deduced sequence with the N-terminal amino acid sequence previously determined by Edman degradation analysis suggested the presence of a signal sequence of 27 residues. The primary structure of TfaA indicated that the protein has a tertiary structure quite similar to those of other quinoprotein dehydrogenases.

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Characterization of Gmhsp26-A, a stress gene encoding a divergent heat shock protein of soybean: heavy-metal-induced inhibition of intron processing.

Molecular and Cellular Biology ◽

10.1128/mcb.8.3.1113 ◽

1988 ◽

Vol 8 (3) ◽

pp. 1113-1122 ◽

Cited By ~ 91

Author(s):

E Czarnecka ◽

R T Nagao ◽

J L Key ◽

W B Gurley

Keyword(s):

Amino Acid ◽

Heat Shock ◽

Amino Acid Sequence ◽

Genomic Sequence ◽

Stress Protein ◽

Initiation Site ◽

Gene Encoding ◽

S1 Nuclease ◽

Soybean Seedlings ◽

Nuclease Mapping

We determined the DNA sequence and mapped the corresponding transcripts of a genomic clone containing the Gmhsp26-A gene of soybean. This gene is homologous to the previously characterized cDNA clone pCE54 (E. Czarnecka, L. Edelman, F. Schöffl, and J. L. Key, Plant Mol. Biol. 3:45-58, 1984) and is expressed in response to a wide variety of physiological stresses including heat shock (HS). S1 nuclease mapping of transcripts and a comparison of the cDNA sequence with the genomic sequence indicated the presence of a soybean seedlings with either CdCl2 or CuSO4. Analysis of the 5' termini of transcripts indicated the presence of one major and at least two minor start sites. In each case, initiation occurred 27 to 30 base pairs downstream from a TATA-like motif, and thus each initiation site appears to be promoted by the activity of a separate subpromoter. The three subpromoters are all associated with sequences showing low homology to the HS consensus element of Drosophila melanogaster HS genes and are differentially induced in response to various stresses. Within the carboxyl-terminal half of the protein, hydropathy analysis of the deduced amino acid sequence indicated a high degree of relatedness to the small HS proteins. A comparison of the primary amino acid sequence of hsp26-A with sequences of the small HS proteins suggested that this stress protein is highly diverged and may therefore be specialized for stress adaptation in soybean.

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Characterization of Gmhsp26-A, a stress gene encoding a divergent heat shock protein of soybean: heavy-metal-induced inhibition of intron processing

Molecular and Cellular Biology ◽

10.1128/mcb.8.3.1113-1122.1988 ◽

1988 ◽

Vol 8 (3) ◽

pp. 1113-1122

Author(s):

E Czarnecka ◽

R T Nagao ◽

J L Key ◽

W B Gurley

Keyword(s):

Amino Acid ◽

Heat Shock ◽

Amino Acid Sequence ◽

Genomic Sequence ◽

Stress Protein ◽

Initiation Site ◽

Gene Encoding ◽

S1 Nuclease ◽

Soybean Seedlings ◽

Nuclease Mapping

We determined the DNA sequence and mapped the corresponding transcripts of a genomic clone containing the Gmhsp26-A gene of soybean. This gene is homologous to the previously characterized cDNA clone pCE54 (E. Czarnecka, L. Edelman, F. Schöffl, and J. L. Key, Plant Mol. Biol. 3:45-58, 1984) and is expressed in response to a wide variety of physiological stresses including heat shock (HS). S1 nuclease mapping of transcripts and a comparison of the cDNA sequence with the genomic sequence indicated the presence of a soybean seedlings with either CdCl2 or CuSO4. Analysis of the 5' termini of transcripts indicated the presence of one major and at least two minor start sites. In each case, initiation occurred 27 to 30 base pairs downstream from a TATA-like motif, and thus each initiation site appears to be promoted by the activity of a separate subpromoter. The three subpromoters are all associated with sequences showing low homology to the HS consensus element of Drosophila melanogaster HS genes and are differentially induced in response to various stresses. Within the carboxyl-terminal half of the protein, hydropathy analysis of the deduced amino acid sequence indicated a high degree of relatedness to the small HS proteins. A comparison of the primary amino acid sequence of hsp26-A with sequences of the small HS proteins suggested that this stress protein is highly diverged and may therefore be specialized for stress adaptation in soybean.

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The EGD1 product, a yeast homolog of human BTF3, may be involved in GAL4 DNA binding.

Molecular and Cellular Biology ◽

10.1128/mcb.12.12.5683 ◽

1992 ◽

Vol 12 (12) ◽

pp. 5683-5689 ◽

Cited By ~ 29

Author(s):

M R Parthun ◽

D A Mangus ◽

J A Jaehning

Keyword(s):

Amino Acid ◽

Dna Binding ◽

Amino Acid Sequence ◽

Yeast Cells ◽

Stable Complex ◽

Affinity Column ◽

Gene Encoding ◽

Heat Stable ◽

Gel Retardation ◽

Final Purification Step

A variety of techniques, including filter binding, footprinting, and gel retardation, can be used to assay the transcriptional activator GAL4 (Gal4p) through the initial steps of its purification from yeast cells. Following DNA affinity chromatography, Gal4p still bound DNA selectively when assayed by filter binding or footprinting. However, the affinity-purified protein was no longer capable of forming a stable complex with DNA, as assayed by gel retardation. Mixing the purified Gal4p with the flowthrough fraction from the DNA affinity column restored gel retardation complex formation. Gel retardation assays were used to monitor the purification of a heat-stable Gal4p-DNA complex stabilization activity from the affinity column flowthrough. The activity coeluted from the final purification step with polypeptides of 21 and 27 kDa. The yeast gene encoding the 21-kDa protein was cloned on the basis of its N-terminal amino acid sequence. The gene, named EGD1 (enhancer of GAL4 DNA binding), encodes a highly basic protein (21% lysine and arginine) with a predicted molecular mass of 16.5 kDa. The amino acid sequence of the EGD1 product, Egd1p, is highly similar to that of the human protein BTF3 (X. M. Zheng, D. Black, P. Chambon, and J. M. Egly, Nature [London] 344:556-559, 1990). Although an egd1 null mutant was viable and Gal+, induction of the galactose-regulated genes in the egd1 mutant strain was significantly reduced when cells were shifted from glucose to galactose.

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Pathogenesis-Related Proteins in Trifolium subterraneum: A General Survey and Subsequent Characterisation of a Protein Inducible by Ethephon and Redlegged Earth Mite Attack

Functional Plant Biology ◽

10.1071/pp97022 ◽

1997 ◽

Vol 24 (6) ◽

pp. 819 ◽

Cited By ~ 14

Author(s):

Kevin Broderick ◽

Christopher Pittock ◽

Tony Arioli ◽

Ernest H. Creaser ◽

Jeremy J. Weinman ◽

...

Keyword(s):

Amino Acid ◽

Amino Acid Sequence ◽

Subterranean Clover ◽

Trifolium Subterraneum ◽

Chitinase Activity ◽

Pr Proteins ◽

Gene Encoding ◽

Pathogenesis Related ◽

Redlegged Earth Mite ◽

Ethephon Treatment

One of the chief predators of subterranean clover (Trifolium subterraneum) pastures is redlegged earth mite (Halotydeus destructor; RLEM). Subterranean clover pathogenesis-related (PR) proteins induced by RLEM attack and ethephon treatment were surveyed, and PR proteins with peroxidase, β-1,3-glucanase and chitinase activities were detected. A protein co-migrating with a chitinase activity, induced by RLEM predation and treatment with ethephon, was isolated. It was purified and the N-terminal amino acid sequence determined. Using a degenerate oligonucleotide primer designed from this sequence, a corresponding cDNA fragment was amplified by reverse transcriptase-PCR, then cloned, and used as a probe to screen a subterranean clover cv. Karridale genomic library. The cDNA and a 97% homologous genomic clone were sequenced and the deduced amino acid sequence revealed an open reading frame of 157 amino acids capable of encoding a peptide of 16 478 Da. Significant homology (80%) was found between this protein and an abscisic acid (ABA)-responsive protein from Pisum sativum of unknown function which is an intracellular pathogenesis-related (IPR) protein. The gene encoding this protein also has homology to pea ‘disease response resistance genes’ and to proteins from other plant species in the PR-10 family. The induced protein was designated TsPR-10a due to its homology to other PR-10 proteins. Genomic Southern analysis indicates that the gene encoding this protein, designated Ypr10a, is a member of a multigene family with at least three members. Northern blot analysis indicates that the subterranean clover Ypr10a mRNA, or homologous transcript, level is strongly induced by ethephon treatment in both root and aerial tissues of 3 week old plants. The rapid induction kinetics of Ypr10a mRNA under ethephon treatment, its correlation with a putative chitinase activity, and homology to other PR-protein genes, suggests a pathogenesis-related role for TsPR-10a protein in subterranean clover.

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A novel phytase from Citrobacter gillenii: characterization and expression in Pichia pastoris (Komagataella pastoris)

FEMS Microbiology Letters ◽

10.1093/femsle/fnaa217 ◽

2020 ◽

Author(s):

Artur A Tkachenko ◽

Anna N Kalinina ◽

Larisa N Borshchevskaya ◽

Sergey P Sineoky ◽

Tatiana L Gordeeva

Keyword(s):

Amino Acid ◽

Pichia Pastoris ◽

Amino Acid Sequence ◽

Specific Activity ◽

Gene Encoding ◽

Maximum Reaction Rate ◽

Maximum Reaction ◽

Wide Ph Range ◽

Recombinant Phytase ◽

Ph Range

Abstract The phyCg gene encoding a new phytase from C. gillenii was optimized, synthesized, cloned, and expressed in Pichia pastoris. Analysis of the amino acid sequence of the enzyme showed that it belongs to the histidine acid phosphatase family. The amino acid sequence of the PhyCg phytase has the highest homology (73.49%) with a phytase sequence from Citrobacter braakii. The main characteristics for the purified recombinant phytase were established. The optimum pH and temperature were 4.5 and 50°C, respectively. The specific activity of the enzyme was 1577 U/mg. The Michaelis constant (Km) and the maximum reaction rate (Vmax) for sodium phytate were 0,185 mM and 2185 U/mg, respectively. The enzyme showed the pH and trypsin stability and had a high activity over a wide pH range.

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Molecular Heterogeneity of the L-1 Metallo-β-Lactamase Family from Stenotrophomonas maltophilia

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.5.1245 ◽

1998 ◽

Vol 42 (5) ◽

pp. 1245-1248 ◽

Cited By ~ 28

Author(s):

François Sanschagrin ◽

Julien Dufresne ◽

Roger C. Levesque

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Nucleotide Sequence ◽

Amino Acid Sequence ◽

Stenotrophomonas Maltophilia ◽

Amino Acid Sequences ◽

Molecular Heterogeneity ◽

Gene Encoding ◽

Class B

ABSTRACT We have determined the nucleotide sequence of the blaSgene encoding the carbapenem-hydrolyzing L-1 β-lactamase fromStenotrophomonas maltophilia GN12873. Analysis of the DNA and deduced amino acid sequences identified a product of 290 amino acids. Comparisons of the L-1 amino acid sequence with those of other zinc β-lactamases showed 88.6% identity with the L-1 enzyme fromS. maltophilia IID1275 and less than 20% identity with other class B metalloenzymes.

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Cloning and characterization of a gene encoding pig epidermal growth factor

Journal of Molecular Endocrinology ◽

10.1677/jme.0.0060063 ◽

1991 ◽

Vol 6 (1) ◽

pp. 63-70 ◽

Cited By ~ 18

Author(s):

J. C. Pascall ◽

D. S. C. Jones ◽

S. M. Doel ◽

J. M. Clements ◽

M. Hunter ◽

...

Keyword(s):

Amino Acid ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Amino Acid Sequence ◽

Egf Receptor ◽

Predicted Amino Acid Sequence ◽

Recombinant Polypeptide ◽

Gene Encoding ◽

Epidermal Growth ◽

Direct Amplification

ABSTRACT A portion of the pig epidermal growth factor (EGF) gene has been isolated and characterized. The nucleotide sequencies of exons 20 and 21, which encode the EGF region of the precursor protein, show 85% similarity with the human EGF gene sequence. In addition, conservation of the intron—exon boundaries between the two species was generally observed. Although the pig exon 21 appeared to lack a single nucleotide at its 5′ end relative to the human gene, sequences obtained by direct amplification of the genomic DNA around the 5′ end of this exon using the polymerase chain reaction, and from a pig EGF cDNA recombinant isolated from a kidney library, indicated that the deletion was probably a cloning artifact. Comparison of the predicted amino acid sequence of pig EGF with that of EGF from other species, as well as with several other polypeptides which bind to the EGF receptor, indicated conservation of Gly18, Tyr37, Gly39 and Arg41 in addition to all six cysteine residues and Leu47, which are known to be critical for biological activity. A synthetic gene encoding the predicted amino acid sequence of pig EGF was expressed in yeast. The recombinant polypeptide was shown to compete with 125I-labelled mouse EGF for binding to cells and to stimulate DNA synthesis in quiescent monolayers of Swiss 3T3 cells.

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