Biosynthetic studies of thiosugar-containing natural products, BE-7585A and Lincomycin A

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dc.contributor.advisorLiu, Hung-wen, 1952-en
dc.contributor.committeeMemberAnslyn, Eric Ven
dc.contributor.committeeMemberBielawski, Christopheren
dc.contributor.committeeMemberFast, Walter Len
dc.contributor.committeeMemberWhitman, Christian Pen
dc.creatorSasaki, Eitaen
dc.date.accessioned2016-05-12T16:19:15Z
dc.date.accessioned2018-01-22T22:29:52Z
dc.date.available2016-05-12T16:19:15Z
dc.date.available2018-01-22T22:29:52Z
dc.date.issued2011-05en
dc.date.submittedMay 2011
dc.date.updated2016-05-12T16:19:15Z
dc.description.abstractSulfur is an essential element found ubiquitously in living systems. However, there exist only a few sulfur-containing sugars in nature and their biosyntheses have not been well understood. On the other hand, a wide variety of sugar derivatives commonly found in natural products are often vital components for the efficacy and specificity of their parent molecules. Elucidation of such unusual sugar biosyntheses is important both for understanding their intriguing chemical mechanisms and creating unnatural compounds by altering their biosynthetic machineries, which could potentially exhibit enhanced or novel biological activities. This dissertation describes biosynthetic studies of two thiosugar-containing natural products, BE-7585A and lincomycin A, produced by Amycolatopsis orientalis and Streptomyces lincolnensis, respectively. While the former possess a C-2-thiosugar-containing disaccharide moiety, the latter contains a C-1-thio substituent on a characteristic eight-carbon backbone sugar. The focus of this research is to characterize the biological pathways and mechanisms responsible for the sulfur incorporation and the unique sugar scaffolds. BE-7585A, an angucycline-type natural product, contains the rare C-2-thiosugar moiety. PCR-based screening of a cosmid library constructed from the genomic DNA of A. orientalis led to the identification of the BE-7585A biosynthetic gene cluster. A gene, bexX, was found to be a candidate for a thiosugar synthase with moderate sequence similarity to a thiazole synthase. The gene, bexX, and a glycosyltransferase homologue, bexG2, were heterologously expressed in Escherichia coli. A variety of biochemical experiments provided a wealth of evidence supporting the proposed biosynthetic pathway for the C-2-thiodisaccharide moiety. Finally, whole genome sequencing and a genome mining approach led to the identification of a sulfur carrier protein to accomplish the in vitro enzymatic synthesis of the C-2-thiosugar for the first time. Lincomycin A is a lincosamide antimicrobial natural product with a C-1 methylthio substituent. Although the lincomycin A biosynthetic gene cluster has been reported, biochemical verification of the biosynthetic pathway has remained elusive. In this dissertation, the complete methlthiolincosamide biosynthetic pathway including the potential C-1 sulfur incorporation mechanism was proposed. Furthermore, two early intermediates of the pathway were characterized for the first time by demonstrating the LmbR (transaldolase) and LmbN (isomerase) reactions in vitro.en
dc.description.departmentChemistryen
dc.format.mimetypeapplication/pdfen
dc.identifierdoi:10.15781/T2BZ5Qen
dc.identifier.urihttp://hdl.handle.net/2152/35412en
dc.subjectBiosynthesis
dc.subjectThiosugar
dc.subjectAngucycline
dc.subjectLincosamide
dc.subjectBE-7585A
dc.subjectLincomycin
dc.subjectSulfur
dc.subjectAntibiotic
dc.subjectAnticancer
dc.subjectUnusual sugar
dc.subjectDeoxysugar
dc.subjectTrehalose
dc.subjectTransaldolase
dc.subjectOctulose
dc.subjectOctose
dc.subjectMechanism
dc.subjectPathway
dc.titleBiosynthetic studies of thiosugar-containing natural products, BE-7585A and Lincomycin Aen
dc.typeThesisen
dc.type.materialtexten

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