Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1
A psychrophilic bacterium, Cytophaga sp. strain KUC-1, that abundantly produces a NAD+-dependent l-threonine dehydrogenase was isolated from Antarctic seawater, and the enzyme was purified. The molecular weight of the enzyme was estimated to be 139,000, and that of the subunit was determined to be 3...
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American Society for Microbiology
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ftpubmed:oai:pubmedcentral.nih.gov:165783 2023-05-15T13:32:01+02:00 Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 Kazuoka, Takayuki Takigawa, Shouhei Arakawa, Noriaki Hizukuri, Yoshiyuki Muraoka, Ikuo Oikawa, Tadao Soda, Kenji 2003-08 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC165783 http://www.ncbi.nlm.nih.gov/pubmed/12867457 https://doi.org/10.1128/JB.185.15.4483-4489.2003 en eng American Society for Microbiology http://www.ncbi.nlm.nih.gov/pmc/articles/PMC165783 http://www.ncbi.nlm.nih.gov/pubmed/12867457 http://dx.doi.org/10.1128/JB.185.15.4483-4489.2003 Copyright © 2003, American Society for Microbiology Enzymes and Proteins Text 2003 ftpubmed https://doi.org/10.1128/JB.185.15.4483-4489.2003 2013-08-29T12:33:19Z A psychrophilic bacterium, Cytophaga sp. strain KUC-1, that abundantly produces a NAD+-dependent l-threonine dehydrogenase was isolated from Antarctic seawater, and the enzyme was purified. The molecular weight of the enzyme was estimated to be 139,000, and that of the subunit was determined to be 35,000. The enzyme is a homotetramer. Atomic absorption analysis showed that the enzyme contains no metals. In these respects, the Cytophaga enzyme is distinct from other l-threonine dehydrogenases that have thus far been studied. l-Threonine and dl-threo-3-hydroxynorvaline were the substrates, and NAD+ and some of its analogs served as coenzymes. The enzyme showed maximum activity at pH 9.5 and at 45°C. The kinetic parameters of the enzyme are highly influenced by temperatures. The Km for l-threonine was lowest at 20°C. Dead-end inhibition studies with pyruvate and adenosine-5′-diphosphoribose showed that the enzyme reaction proceeds via the ordered Bi Bi mechanism in which NAD+ binds to an enzyme prior to l-threonine and 2-amino-3-oxobutyrate is released from the enzyme prior to NADH. The enzyme gene was cloned into Escherichia coli, and its nucleotides were sequenced. The enzyme gene contains an open reading frame of 939 bp encoding a protein of 312 amino acid residues. The amino acid sequence of the enzyme showed a significant similarity to that of UDP-glucose 4-epimerase from Staphylococcus aureus and belongs to the short-chain dehydrogenase-reductase superfamily. In contrast, l-threonine dehydrogenase from E. coli belongs to the medium-chain alcohol dehydrogenase family, and its amino acid sequence is not at all similar to that of the Cytophaga enzyme. l-Threonine dehydrogenase is significantly similar to an epimerase, which was shown for the first time. The amino acid residues playing an important role in the catalysis of the E. coli and human UDP-glucose 4-epimerases are highly conserved in the Cytophaga enzyme, except for the residues participating in the substrate binding. Text Antarc* Antarctic PubMed Central (PMC) Antarctic Journal of Bacteriology 185 15 4483 4489 |
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Enzymes and Proteins |
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Enzymes and Proteins Kazuoka, Takayuki Takigawa, Shouhei Arakawa, Noriaki Hizukuri, Yoshiyuki Muraoka, Ikuo Oikawa, Tadao Soda, Kenji Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
topic_facet |
Enzymes and Proteins |
description |
A psychrophilic bacterium, Cytophaga sp. strain KUC-1, that abundantly produces a NAD+-dependent l-threonine dehydrogenase was isolated from Antarctic seawater, and the enzyme was purified. The molecular weight of the enzyme was estimated to be 139,000, and that of the subunit was determined to be 35,000. The enzyme is a homotetramer. Atomic absorption analysis showed that the enzyme contains no metals. In these respects, the Cytophaga enzyme is distinct from other l-threonine dehydrogenases that have thus far been studied. l-Threonine and dl-threo-3-hydroxynorvaline were the substrates, and NAD+ and some of its analogs served as coenzymes. The enzyme showed maximum activity at pH 9.5 and at 45°C. The kinetic parameters of the enzyme are highly influenced by temperatures. The Km for l-threonine was lowest at 20°C. Dead-end inhibition studies with pyruvate and adenosine-5′-diphosphoribose showed that the enzyme reaction proceeds via the ordered Bi Bi mechanism in which NAD+ binds to an enzyme prior to l-threonine and 2-amino-3-oxobutyrate is released from the enzyme prior to NADH. The enzyme gene was cloned into Escherichia coli, and its nucleotides were sequenced. The enzyme gene contains an open reading frame of 939 bp encoding a protein of 312 amino acid residues. The amino acid sequence of the enzyme showed a significant similarity to that of UDP-glucose 4-epimerase from Staphylococcus aureus and belongs to the short-chain dehydrogenase-reductase superfamily. In contrast, l-threonine dehydrogenase from E. coli belongs to the medium-chain alcohol dehydrogenase family, and its amino acid sequence is not at all similar to that of the Cytophaga enzyme. l-Threonine dehydrogenase is significantly similar to an epimerase, which was shown for the first time. The amino acid residues playing an important role in the catalysis of the E. coli and human UDP-glucose 4-epimerases are highly conserved in the Cytophaga enzyme, except for the residues participating in the substrate binding. |
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Text |
author |
Kazuoka, Takayuki Takigawa, Shouhei Arakawa, Noriaki Hizukuri, Yoshiyuki Muraoka, Ikuo Oikawa, Tadao Soda, Kenji |
author_facet |
Kazuoka, Takayuki Takigawa, Shouhei Arakawa, Noriaki Hizukuri, Yoshiyuki Muraoka, Ikuo Oikawa, Tadao Soda, Kenji |
author_sort |
Kazuoka, Takayuki |
title |
Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
title_short |
Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
title_full |
Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
title_fullStr |
Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
title_full_unstemmed |
Novel Psychrophilic and Thermolabile l-Threonine Dehydrogenase from Psychrophilic Cytophaga sp. Strain KUC-1 |
title_sort |
novel psychrophilic and thermolabile l-threonine dehydrogenase from psychrophilic cytophaga sp. strain kuc-1 |
publisher |
American Society for Microbiology |
publishDate |
2003 |
url |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC165783 http://www.ncbi.nlm.nih.gov/pubmed/12867457 https://doi.org/10.1128/JB.185.15.4483-4489.2003 |
geographic |
Antarctic |
geographic_facet |
Antarctic |
genre |
Antarc* Antarctic |
genre_facet |
Antarc* Antarctic |
op_relation |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC165783 http://www.ncbi.nlm.nih.gov/pubmed/12867457 http://dx.doi.org/10.1128/JB.185.15.4483-4489.2003 |
op_rights |
Copyright © 2003, American Society for Microbiology |
op_doi |
https://doi.org/10.1128/JB.185.15.4483-4489.2003 |
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Journal of Bacteriology |
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185 |
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15 |
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4483 |
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4489 |
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1766023665616420864 |