Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain.
Identifieur interne : 000066 ( PubMed/Corpus ); précédent : 000065; suivant : 000067Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain.
Auteurs : Moez Rhimi ; Samir BejarSource :
- Biochimica et biophysica acta [ 0006-3002 ] ; 2006.
English descriptors
- KwdEn :
- Aldose-Ketose Isomerases (genetics), Aldose-Ketose Isomerases (isolation & purification), Amino Acid Sequence, Base Sequence, Chromatography, High Pressure Liquid, Cloning, Molecular, Cobalt (pharmacology), Enzyme Stability, Geobacillus stearothermophilus (enzymology), Geobacillus stearothermophilus (genetics), Hot Temperature, Hydrogen-Ion Concentration, Kinetics, Manganese (pharmacology), Molecular Sequence Data, Recombinant Proteins (isolation & purification).
- MESH :
- chemical , genetics : Aldose-Ketose Isomerases.
- chemical , isolation & purification : Aldose-Ketose Isomerases, Recombinant Proteins.
- chemical , pharmacology : Cobalt, Manganese.
- enzymology : Geobacillus stearothermophilus.
- genetics : Geobacillus stearothermophilus.
- Amino Acid Sequence, Base Sequence, Chromatography, High Pressure Liquid, Cloning, Molecular, Enzyme Stability, Hot Temperature, Hydrogen-Ion Concentration, Kinetics, Molecular Sequence Data.
Abstract
The araA gene encoding L-arabinose isomerase from Bacillus stearothermophilus US100 strain was cloned, sequenced and over-expressed in E. coli. This gene encodes a 496-amino acid protein with a calculated molecular weight of 56.161 kDa. Its amino acid sequence displays the highest identity with L-AI from Thermus sp. IM6501 (98%) and that of Geobacillus stearothermophilus T6 (97%). According to SDS-PAGE analysis, under reducing and non-reducing conditions, the recombinant enzyme has an apparent molecular weight of nearly 225 kDa, composed of four identical 56-kDa subunits. The L-AI US100 was optimally active at pH 7.5 and 80 degrees C. It was distinguishable by its behavior towards divalent ions. Indeed, the L-AI US100 activity and thermostability were totally independent for metallic ions until 65 degrees C. At temperatures above 65 degrees C, the enzyme was also independent for metallic ions for its activity but its thermostability was obviously improved in presence of only 0.2 mM Co2+ and 1 mM Mn2+. The V(max) values were calculated to be 41.3 U/mg for L-arabinose and 8.9 U/mg for D-galactose. Their catalytic efficiencies (k(cat)/K(m)) for l-arabinose and D-galactose were, respectively, 71.4 and 8.46 mM(-1) min(-1). L-AI US100 converted the d-galactose into D-tagatose with a high conversion rate of 48% after 7 h at 70 degrees C.
DOI: 10.1016/j.bbagen.2005.11.007
PubMed: 16386851
Links to Exploration step
pubmed:16386851Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain.</title><author><name sortKey="Rhimi, Moez" sort="Rhimi, Moez" uniqKey="Rhimi M" first="Moez" last="Rhimi">Moez Rhimi</name><affiliation><nlm:affiliation>Laboratoire d'Enzymes et de Métabolites des Procaryotes, Centre de Biotechnologie de Sfax BP K 3038 Sfax, Tunisie.</nlm:affiliation></affiliation></author><author><name sortKey="Bejar, Samir" sort="Bejar, Samir" uniqKey="Bejar S" first="Samir" last="Bejar">Samir Bejar</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2006">2006</date><idno type="RBID">pubmed:16386851</idno><idno type="pmid">16386851</idno><idno type="doi">10.1016/j.bbagen.2005.11.007</idno><idno type="wicri:Area/PubMed/Corpus">000066</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">000066</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain.</title><author><name sortKey="Rhimi, Moez" sort="Rhimi, Moez" uniqKey="Rhimi M" first="Moez" last="Rhimi">Moez Rhimi</name><affiliation><nlm:affiliation>Laboratoire d'Enzymes et de Métabolites des Procaryotes, Centre de Biotechnologie de Sfax BP K 3038 Sfax, Tunisie.</nlm:affiliation></affiliation></author><author><name sortKey="Bejar, Samir" sort="Bejar, Samir" uniqKey="Bejar S" first="Samir" last="Bejar">Samir Bejar</name></author></analytic><series><title level="j">Biochimica et biophysica acta</title><idno type="ISSN">0006-3002</idno><imprint><date when="2006" type="published">2006</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Aldose-Ketose Isomerases (genetics)</term><term>Aldose-Ketose Isomerases (isolation & purification)</term><term>Amino Acid Sequence</term><term>Base Sequence</term><term>Chromatography, High Pressure Liquid</term><term>Cloning, Molecular</term><term>Cobalt (pharmacology)</term><term>Enzyme Stability</term><term>Geobacillus stearothermophilus (enzymology)</term><term>Geobacillus stearothermophilus (genetics)</term><term>Hot Temperature</term><term>Hydrogen-Ion Concentration</term><term>Kinetics</term><term>Manganese (pharmacology)</term><term>Molecular Sequence Data</term><term>Recombinant Proteins (isolation & purification)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Aldose-Ketose Isomerases</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>Aldose-Ketose Isomerases</term><term>Recombinant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Cobalt</term><term>Manganese</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>Geobacillus stearothermophilus</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Geobacillus stearothermophilus</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Amino Acid Sequence</term><term>Base Sequence</term><term>Chromatography, High Pressure Liquid</term><term>Cloning, Molecular</term><term>Enzyme Stability</term><term>Hot Temperature</term><term>Hydrogen-Ion Concentration</term><term>Kinetics</term><term>Molecular Sequence Data</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The araA gene encoding L-arabinose isomerase from Bacillus stearothermophilus US100 strain was cloned, sequenced and over-expressed in E. coli. This gene encodes a 496-amino acid protein with a calculated molecular weight of 56.161 kDa. Its amino acid sequence displays the highest identity with L-AI from Thermus sp. IM6501 (98%) and that of Geobacillus stearothermophilus T6 (97%). According to SDS-PAGE analysis, under reducing and non-reducing conditions, the recombinant enzyme has an apparent molecular weight of nearly 225 kDa, composed of four identical 56-kDa subunits. The L-AI US100 was optimally active at pH 7.5 and 80 degrees C. It was distinguishable by its behavior towards divalent ions. Indeed, the L-AI US100 activity and thermostability were totally independent for metallic ions until 65 degrees C. At temperatures above 65 degrees C, the enzyme was also independent for metallic ions for its activity but its thermostability was obviously improved in presence of only 0.2 mM Co2+ and 1 mM Mn2+. The V(max) values were calculated to be 41.3 U/mg for L-arabinose and 8.9 U/mg for D-galactose. Their catalytic efficiencies (k(cat)/K(m)) for l-arabinose and D-galactose were, respectively, 71.4 and 8.46 mM(-1) min(-1). L-AI US100 converted the d-galactose into D-tagatose with a high conversion rate of 48% after 7 h at 70 degrees C.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">16386851</PMID><DateCreated><Year>2006</Year><Month>3</Month><Day>6</Day></DateCreated><DateCompleted><Year>2006</Year><Month>04</Month><Day>27</Day></DateCompleted><DateRevised><Year>2013</Year><Month>11</Month><Day>21</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Print">0006-3002</ISSN><JournalIssue CitedMedium="Print"><Volume>1760</Volume><Issue>2</Issue><PubDate><Year>2006</Year><Month>Feb</Month></PubDate></JournalIssue><Title>Biochimica et biophysica acta</Title><ISOAbbreviation>Biochim. Biophys. Acta</ISOAbbreviation></Journal><ArticleTitle>Cloning, purification and biochemical characterization of metallic-ions independent and thermoactive l-arabinose isomerase from the Bacillus stearothermophilus US100 strain.</ArticleTitle><Pagination><MedlinePgn>191-9</MedlinePgn></Pagination><Abstract><AbstractText>The araA gene encoding L-arabinose isomerase from Bacillus stearothermophilus US100 strain was cloned, sequenced and over-expressed in E. coli. This gene encodes a 496-amino acid protein with a calculated molecular weight of 56.161 kDa. Its amino acid sequence displays the highest identity with L-AI from Thermus sp. IM6501 (98%) and that of Geobacillus stearothermophilus T6 (97%). According to SDS-PAGE analysis, under reducing and non-reducing conditions, the recombinant enzyme has an apparent molecular weight of nearly 225 kDa, composed of four identical 56-kDa subunits. The L-AI US100 was optimally active at pH 7.5 and 80 degrees C. It was distinguishable by its behavior towards divalent ions. Indeed, the L-AI US100 activity and thermostability were totally independent for metallic ions until 65 degrees C. At temperatures above 65 degrees C, the enzyme was also independent for metallic ions for its activity but its thermostability was obviously improved in presence of only 0.2 mM Co2+ and 1 mM Mn2+. The V(max) values were calculated to be 41.3 U/mg for L-arabinose and 8.9 U/mg for D-galactose. Their catalytic efficiencies (k(cat)/K(m)) for l-arabinose and D-galactose were, respectively, 71.4 and 8.46 mM(-1) min(-1). L-AI US100 converted the d-galactose into D-tagatose with a high conversion rate of 48% after 7 h at 70 degrees C.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Rhimi</LastName><ForeName>Moez</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Laboratoire d'Enzymes et de Métabolites des Procaryotes, Centre de Biotechnologie de Sfax BP K 3038 Sfax, Tunisie.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Bejar</LastName><ForeName>Samir</ForeName><Initials>S</Initials></Author></AuthorList><Language>ENG</Language><DataBankList CompleteYN="Y"><DataBank><DataBankName>GENBANK</DataBankName><AccessionNumberList><AccessionNumber>AJ866972</AccessionNumber></AccessionNumberList></DataBank></DataBankList><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2005</Year><Month>Dec</Month><Day>13</Day></ArticleDate></Article><MedlineJournalInfo><Country>Netherlands</Country><MedlineTA>Biochim Biophys Acta</MedlineTA><NlmUniqueID>0217513</NlmUniqueID><ISSNLinking>0006-3002</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D011994">Recombinant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>3G0H8C9362</RegistryNumber><NameOfSubstance UI="D003035">Cobalt</NameOfSubstance></Chemical><Chemical><RegistryNumber>42Z2K6ZL8P</RegistryNumber><NameOfSubstance UI="D008345">Manganese</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 5.3.1.-</RegistryNumber><NameOfSubstance UI="D019747">Aldose-Ketose Isomerases</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 5.3.1.4</RegistryNumber><NameOfSubstance UI="C019715">L-arabinose isomerase</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D019747" MajorTopicYN="N">Aldose-Ketose Isomerases</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="Y">genetics</QualifierName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D000595" MajorTopicYN="N">Amino Acid Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D001483" MajorTopicYN="N">Base Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D002851" MajorTopicYN="N">Chromatography, High Pressure Liquid</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D003001" MajorTopicYN="N">Cloning, Molecular</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D003035" MajorTopicYN="N">Cobalt</DescriptorName><QualifierName UI="Q000494" MajorTopicYN="Y">pharmacology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D004795" MajorTopicYN="N">Enzyme Stability</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D001411" MajorTopicYN="N">Geobacillus stearothermophilus</DescriptorName><QualifierName UI="Q000201" MajorTopicYN="Y">enzymology</QualifierName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D006358" MajorTopicYN="N">Hot Temperature</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D006863" MajorTopicYN="N">Hydrogen-Ion Concentration</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D007700" MajorTopicYN="N">Kinetics</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008345" MajorTopicYN="N">Manganese</DescriptorName><QualifierName UI="Q000494" MajorTopicYN="Y">pharmacology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D008969" MajorTopicYN="N">Molecular Sequence Data</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D011994" MajorTopicYN="N">Recombinant Proteins</DescriptorName><QualifierName UI="Q000302" MajorTopicYN="N">isolation & purification</QualifierName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2005</Year><Month>5</Month><Day>17</Day></PubMedPubDate><PubMedPubDate PubStatus="revised"><Year>2005</Year><Month>11</Month><Day>9</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2005</Year><Month>11</Month><Day>9</Day></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2006</Year><Month>1</Month><Day>3</Day><Hour>9</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2006</Year><Month>4</Month><Day>28</Day><Hour>9</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2006</Year><Month>1</Month><Day>3</Day><Hour>9</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">16386851</ArticleId><ArticleId IdType="pii">S0304-4165(05)00358-2</ArticleId><ArticleId IdType="doi">10.1016/j.bbagen.2005.11.007</ArticleId></ArticleIdList></PubmedData></pubmed></record>Pour manipuler ce document sous Unix (Dilib)
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