Cloning, expression and characterization of Novel Lipase and Esterases from Burkholderia multivorans UWC10

dc.contributor.advisorCowan, Don A
dc.contributor.authorRashamuse, Konanani J
dc.date.accessioned2021-10-28T07:34:24Z
dc.date.accessioned2024-05-09T08:19:08Z
dc.date.available2021-10-28T07:34:24Z
dc.date.available2024-05-09T08:19:08Z
dc.date.issued2005
dc.descriptionDoctor Scientiaeen_US
dc.description.abstractAn esterase and lipase producing Burkholderia multivorans strain was isolated by culture enrichment strategies. A shotgun library of Burkholderia multivorans genomic DNA (prepared in E. coli/pUC18) was screened for lipase and esterase activities. Three positive recombinant clones, pTEND5, pHOLA6 and pRASHI4, conferring esterolytic and lipolytic phenotypes respectively, were identified. Full-length sequencing of DNA inserts was performed using subeloning and "primer-walking" strategies. Nucleotide sequence analysis revealed that the pRASH14 plasmid DNA consisted of two open reading frames (ORPI and ORP2) encoding 356 and 350 amino acids, respectively. Database searches revealed that ORPI and ORP2 were homologous to lipases and chaperones from subfamily I.2. In the pTEND5 sequence, an open reading frame consisting of 978 bp, encoding 326 amino acids, was identified. Database searches revealed that this open reading frame was homologous to family Vesterases. Nucleotide sequence analysis revealed that pHOLA6, plasmid DNA consisted of 1194 bp encoding 398 amino acids and showed homology to family VIII esterases. The primary structures of LipA, EstEFH5 and EstBL from pRASHI4, pTEND5 and pHOLA6, respectively, showed a classical GxSxG motif, which is conserved in many serine hydrolases. In addition, EstBL also showed a consensus SxxK motif, the serine of which acts as a catalytic nucleophile in class C B-lactames and some peptidases.en_US
dc.identifier.urihttps://hdl.handle.net/10566/13471
dc.language.isoenen_US
dc.subjectBurkholderia multivoransen_US
dc.subjectEsterasesen_US
dc.subjectLipasesen_US
dc.subjectaB-hydrolase folden_US
dc.subjectGene cloningen_US
dc.subjectProtein expressionen_US
dc.subjectProtein purificationen_US
dc.titleCloning, expression and characterization of Novel Lipase and Esterases from Burkholderia multivorans UWC10en_US

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