Improvement of Aspergillus niger glucoamylase thermostability by directed evolution

dc.contributor.authorWang, Yue
dc.contributor.authorFuchs, Erica
dc.contributor.authorda Silva, Roberto
dc.contributor.authorMcDaniel, Allison
dc.contributor.authorSeibel, Janice
dc.contributor.authorFord, Clark
dc.contributor.institutionIowa State Univ
dc.contributor.institutionUniv Calif Berkeley
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.date.accessioned2014-05-20T15:25:56Z
dc.date.available2014-05-20T15:25:56Z
dc.date.issued2006-10-01
dc.description.abstractDirected evolution was used to improve the thermostability of Aspergillus niger glucoamylase (GA) expressed in Saccharomyces cerevisiae. A starch-plate assay developed to screen GA mutants for thermostability gave results consistent with those of irreversible thermoinactivation kinetic analysis. Several thermostable multiply-mutated GAs were isolated and characterized by DNA sequencing and kinetic analysis. Three new GA mutations, T62A, T290A and H391Y, have been identified that encode GAs that are more thermostable than wild-type GA, and that improve thermostability cumulatively. These individual mutations were combined with the previously constructed thermostable site-directed mutations D20C/A27C (forming a disulficle bond), S30P, and G137A to create a multiply-mutated GA designated THS8. THS8 GA is substantially more thermostable than wild-type GA at 8OoC, with a 5.1 kJ/mol increase in the free energy of therrnoinactivation, making it the most thermostable Aspergillus niger GA mutant characterized to date. THS8 GA and the singly-mutated GAs have specific activities and catalytic efficiencies (k(cat)/K-m) similar to those of wild-type GA.en
dc.description.affiliationIowa State Univ, Food Sci & Human Nutr Dept, Ames, IA 50011 USA
dc.description.affiliationUniv Calif Berkeley, Dept Bioengn, Berkeley, CA 94720 USA
dc.description.affiliationUNESP, Lab Bioquim & Microbiol Aplicada, Sao Jose do Rio Preto, SP, Brazil
dc.description.affiliationUnespUNESP, Lab Bioquim & Microbiol Aplicada, Sao Jose do Rio Preto, SP, Brazil
dc.format.extent501-508
dc.identifierhttp://dx.doi.org/10.1002/star.200600493
dc.identifier.citationStarch-starke. Weinheim: Wiley-v C H Verlag Gmbh, v. 58, n. 10, p. 501-508, 2006.
dc.identifier.doi10.1002/star.200600493
dc.identifier.issn0038-9056
dc.identifier.lattes9424175688206545
dc.identifier.urihttp://hdl.handle.net/11449/36250
dc.identifier.wosWOS:000241483900001
dc.language.isoeng
dc.publisherWiley-Blackwell
dc.relation.ispartofStarch-starke
dc.relation.ispartofjcr2.173
dc.relation.ispartofsjr0,725
dc.rights.accessRightsAcesso restrito
dc.sourceWeb of Science
dc.subjectthermostabilitypt
dc.subjectdirected evolutionpt
dc.subjectglucoamylasept
dc.subjectenzymept
dc.subjectmutationpt
dc.titleImprovement of Aspergillus niger glucoamylase thermostability by directed evolutionen
dc.typeArtigo
dcterms.licensehttp://olabout.wiley.com/WileyCDA/Section/id-406071.html
dcterms.rightsHolderWiley-Blackwell
unesp.author.lattes9424175688206545
unesp.campusUniversidade Estadual Paulista (Unesp), Instituto de Biociências, Letras e Ciências Exatas, São José do Rio Pretopt
unesp.departmentQuímica e Ciências Ambientais - IBILCEpt

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