Purification and Characterization of a Cellulose-Binding (beta)-Glucosidase from Cellulose-Degrading Cultures of Phanerochaete chrysosporium.
Identifieur interne : 000C89 ( Main/Exploration ); précédent : 000C88; suivant : 000C90Purification and Characterization of a Cellulose-Binding (beta)-Glucosidase from Cellulose-Degrading Cultures of Phanerochaete chrysosporium.
Auteurs : E S Lymar ; B. Li ; V. RenganathanSource :
- Applied and environmental microbiology [ 0099-2240 ] ; 1995.
Abstract
Extracellular (beta)-glucosidase from cellulose-degrading cultures of Phanerochaete chrysosporium was purified by DEAE-Sephadex chromatography, by Sephacryl S-200 chromatography, and by fast protein liquid chromatography (FPLC) using a Mono Q anion-exchange column. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic (SDS-PAGE) analysis of FPLC-purified (beta)-glucosidase indicated the presence of three enzyme forms with molecular weights of 96,000, 98,000, and 114,000. On further fractionation with a microcrystalline cellulose column, the 114,000-molecular-weight (beta)-glucosidase, which had 82% of the (beta)-glucosidase activity, was bound to cellulose. The (beta)-glucosidases with molecular weights of 96,000 and 98,000 did not bind to cellulose. The cellulose-bound (beta)-glucosidase was eluted completely from the cellulose matrix with water. Cellulose-bound (beta)-glucosidase catalyzed p-nitrophenylglucoside hydrolysis, suggesting that the catalytic site is not involved in cellulose binding. When the cellulose-binding form was incubated with papain for 20 h, no decrease in the enzyme activity was observed; however, approximately 74% of the papain-treated glucosidase did not bind to microcrystalline cellulose. SDS-PAGE analysis of the nonbinding glucosidase produced by papain indicated the presence of three bands with molecular weights in the range of 95,000 to 97,000. On the basis of these results, we propose that the low-molecular-weight (96,000 and 98,000) non-cellulose-binding (beta)-glucosidase forms are most probably formed from the higher-molecular-weight (114,000) cellulose-binding (beta)-glucosidase via extracellular proteolytic hydrolysis. Also, it appears that the extracellular (beta)-glucosidase from P. chrysosporium might be organized into two domains, a cellulose-binding domain and a catalytic domain. Kinetic characterization of the cellulose-binding form is also presented.
DOI: 10.1128/AEM.61.8.2976-2980.1995
PubMed: 16535099
PubMed Central: PMC1388553
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Renganathan</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="1995">1995</date><idno type="RBID">pubmed:16535099</idno><idno type="pmid">16535099</idno><idno type="pmc">PMC1388553</idno><idno type="doi">10.1128/AEM.61.8.2976-2980.1995</idno><idno type="wicri:Area/Main/Corpus">000C91</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Corpus" wicri:corpus="PubMed">000C91</idno><idno type="wicri:Area/Main/Curation">000C91</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Curation">000C91</idno><idno type="wicri:Area/Main/Exploration">000C91</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Purification and Characterization of a Cellulose-Binding (beta)-Glucosidase from Cellulose-Degrading Cultures of Phanerochaete chrysosporium.</title><author><name sortKey="Lymar, E S" sort="Lymar, E S" uniqKey="Lymar E" first="E S" last="Lymar">E S Lymar</name></author><author><name sortKey="Li, B" sort="Li, B" uniqKey="Li B" first="B" last="Li">B. Li</name></author><author><name sortKey="Renganathan, V" sort="Renganathan, V" uniqKey="Renganathan V" first="V" last="Renganathan">V. Renganathan</name></author></analytic><series><title level="j">Applied and environmental microbiology</title><idno type="ISSN">0099-2240</idno><imprint><date when="1995" type="published">1995</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Extracellular (beta)-glucosidase from cellulose-degrading cultures of Phanerochaete chrysosporium was purified by DEAE-Sephadex chromatography, by Sephacryl S-200 chromatography, and by fast protein liquid chromatography (FPLC) using a Mono Q anion-exchange column. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic (SDS-PAGE) analysis of FPLC-purified (beta)-glucosidase indicated the presence of three enzyme forms with molecular weights of 96,000, 98,000, and 114,000. On further fractionation with a microcrystalline cellulose column, the 114,000-molecular-weight (beta)-glucosidase, which had 82% of the (beta)-glucosidase activity, was bound to cellulose. The (beta)-glucosidases with molecular weights of 96,000 and 98,000 did not bind to cellulose. The cellulose-bound (beta)-glucosidase was eluted completely from the cellulose matrix with water. Cellulose-bound (beta)-glucosidase catalyzed p-nitrophenylglucoside hydrolysis, suggesting that the catalytic site is not involved in cellulose binding. When the cellulose-binding form was incubated with papain for 20 h, no decrease in the enzyme activity was observed; however, approximately 74% of the papain-treated glucosidase did not bind to microcrystalline cellulose. SDS-PAGE analysis of the nonbinding glucosidase produced by papain indicated the presence of three bands with molecular weights in the range of 95,000 to 97,000. On the basis of these results, we propose that the low-molecular-weight (96,000 and 98,000) non-cellulose-binding (beta)-glucosidase forms are most probably formed from the higher-molecular-weight (114,000) cellulose-binding (beta)-glucosidase via extracellular proteolytic hydrolysis. Also, it appears that the extracellular (beta)-glucosidase from P. chrysosporium might be organized into two domains, a cellulose-binding domain and a catalytic domain. Kinetic characterization of the cellulose-binding form is also presented.</div></front></TEI><pubmed><MedlineCitation Status="PubMed-not-MEDLINE" Owner="NLM"><PMID Version="1">16535099</PMID><DateCompleted><Year>2010</Year><Month>06</Month><Day>25</Day></DateCompleted><DateRevised><Year>2020</Year><Month>07</Month><Day>27</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0099-2240</ISSN><JournalIssue CitedMedium="Print"><Volume>61</Volume><Issue>8</Issue><PubDate><Year>1995</Year><Month>Aug</Month></PubDate></JournalIssue><Title>Applied and environmental microbiology</Title><ISOAbbreviation>Appl Environ Microbiol</ISOAbbreviation></Journal><ArticleTitle>Purification and Characterization of a Cellulose-Binding (beta)-Glucosidase from Cellulose-Degrading Cultures of Phanerochaete chrysosporium.</ArticleTitle><Pagination><MedlinePgn>2976-80</MedlinePgn></Pagination><Abstract><AbstractText>Extracellular (beta)-glucosidase from cellulose-degrading cultures of Phanerochaete chrysosporium was purified by DEAE-Sephadex chromatography, by Sephacryl S-200 chromatography, and by fast protein liquid chromatography (FPLC) using a Mono Q anion-exchange column. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic (SDS-PAGE) analysis of FPLC-purified (beta)-glucosidase indicated the presence of three enzyme forms with molecular weights of 96,000, 98,000, and 114,000. On further fractionation with a microcrystalline cellulose column, the 114,000-molecular-weight (beta)-glucosidase, which had 82% of the (beta)-glucosidase activity, was bound to cellulose. The (beta)-glucosidases with molecular weights of 96,000 and 98,000 did not bind to cellulose. The cellulose-bound (beta)-glucosidase was eluted completely from the cellulose matrix with water. Cellulose-bound (beta)-glucosidase catalyzed p-nitrophenylglucoside hydrolysis, suggesting that the catalytic site is not involved in cellulose binding. When the cellulose-binding form was incubated with papain for 20 h, no decrease in the enzyme activity was observed; however, approximately 74% of the papain-treated glucosidase did not bind to microcrystalline cellulose. SDS-PAGE analysis of the nonbinding glucosidase produced by papain indicated the presence of three bands with molecular weights in the range of 95,000 to 97,000. On the basis of these results, we propose that the low-molecular-weight (96,000 and 98,000) non-cellulose-binding (beta)-glucosidase forms are most probably formed from the higher-molecular-weight (114,000) cellulose-binding (beta)-glucosidase via extracellular proteolytic hydrolysis. Also, it appears that the extracellular (beta)-glucosidase from P. chrysosporium might be organized into two domains, a cellulose-binding domain and a catalytic domain. Kinetic characterization of the cellulose-binding form is also presented.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Lymar</LastName><ForeName>E S</ForeName><Initials>ES</Initials></Author><Author ValidYN="Y"><LastName>Li</LastName><ForeName>B</ForeName><Initials>B</Initials></Author><Author ValidYN="Y"><LastName>Renganathan</LastName><ForeName>V</ForeName><Initials>V</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>Appl Environ Microbiol</MedlineTA><NlmUniqueID>7605801</NlmUniqueID><ISSNLinking>0099-2240</ISSNLinking></MedlineJournalInfo></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>1995</Year><Month>8</Month><Day>1</Day><Hour>0</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>1995</Year><Month>8</Month><Day>1</Day><Hour>0</Hour><Minute>1</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>1995</Year><Month>8</Month><Day>1</Day><Hour>0</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">16535099</ArticleId><ArticleId IdType="pmc">PMC1388553</ArticleId><ArticleId IdType="doi">10.1128/AEM.61.8.2976-2980.1995</ArticleId></ArticleIdList><ReferenceList><Reference><Citation>Eur J Biochem. 1978 Sep 15;90(1):191-8</Citation><ArticleIdList><ArticleId IdType="pubmed">101374</ArticleId></ArticleIdList></Reference><Reference><Citation>Appl Environ Microbiol. 1987 Jul;53(7):1464-9</Citation><ArticleIdList><ArticleId IdType="pubmed">16347375</ArticleId></ArticleIdList></Reference><Reference><Citation>Can J Microbiol. 1977 Feb;23(2):139-47</Citation><ArticleIdList><ArticleId IdType="pubmed">837251</ArticleId></ArticleIdList></Reference><Reference><Citation>FEMS Microbiol Lett. 1992 Oct 1;76(1-2):149-53</Citation><ArticleIdList><ArticleId IdType="pubmed">1426998</ArticleId></ArticleIdList></Reference><Reference><Citation>FEBS Lett. 1992 May 4;302(1):77-80</Citation><ArticleIdList><ArticleId IdType="pubmed">1587358</ArticleId></ArticleIdList></Reference><Reference><Citation>Appl Environ Microbiol. 1992 Jul;58(7):2168-75</Citation><ArticleIdList><ArticleId IdType="pubmed">1637155</ArticleId></ArticleIdList></Reference><Reference><Citation>Microbiol Rev. 1991 Jun;55(2):303-15</Citation><ArticleIdList><ArticleId IdType="pubmed">1886523</ArticleId></ArticleIdList></Reference><Reference><Citation>Eur J Biochem. 1987 Jun 1;165(2):333-41</Citation><ArticleIdList><ArticleId IdType="pubmed">3109900</ArticleId></ArticleIdList></Reference><Reference><Citation>Annu Rev Microbiol. 1987;41:465-505</Citation><ArticleIdList><ArticleId IdType="pubmed">3318677</ArticleId></ArticleIdList></Reference><Reference><Citation>Anal Biochem. 1985 Oct;150(1):76-85</Citation><ArticleIdList><ArticleId IdType="pubmed">3843705</ArticleId></ArticleIdList></Reference><Reference><Citation>Nature. 1970 Aug 15;227(5259):680-5</Citation><ArticleIdList><ArticleId IdType="pubmed">5432063</ArticleId></ArticleIdList></Reference><Reference><Citation>Arch Biochem Biophys. 1981 Mar;207(1):185-96</Citation><ArticleIdList><ArticleId IdType="pubmed">6786226</ArticleId></ArticleIdList></Reference><Reference><Citation>Eur J Biochem. 1982 Jun;124(3):635-42</Citation><ArticleIdList><ArticleId IdType="pubmed">7049699</ArticleId></ArticleIdList></Reference><Reference><Citation>Arch Biochem Biophys. 1993 Feb 1;300(2):705-13</Citation><ArticleIdList><ArticleId IdType="pubmed">8434950</ArticleId></ArticleIdList></Reference><Reference><Citation>Appl Environ Microbiol. 1979 May;37(5):938-42</Citation><ArticleIdList><ArticleId IdType="pubmed">16345389</ArticleId></ArticleIdList></Reference><Reference><Citation>Eur J Biochem. 1978 Sep 15;90(1):171-81</Citation><ArticleIdList><ArticleId IdType="pubmed">710416</ArticleId></ArticleIdList></Reference></ReferenceList></PubmedData></pubmed><affiliations><list></list><tree><noCountry><name sortKey="Li, B" sort="Li, B" uniqKey="Li B" first="B" last="Li">B. Li</name><name sortKey="Lymar, E S" sort="Lymar, E S" uniqKey="Lymar E" first="E S" last="Lymar">E S Lymar</name><name sortKey="Renganathan, V" sort="Renganathan, V" uniqKey="Renganathan V" first="V" last="Renganathan">V. Renganathan</name></noCountry></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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