Biochemical Studies of Polyketide Beta-Branching and Polyketide Synthase Module Architecture

Natural Products Biosynthesis;Bryostatin;Polyketide Synthase;Biological Chemistry;Health Sciences;Biological Chemistry

Slocum, SamuelTrievel, Raymond C ;

University of Michigan

关键词: Natural Products Biosynthesis;    Bryostatin;    Polyketide Synthase;    Biological Chemistry;    Health Sciences;    Biological Chemistry;   

Others  :  https://deepblue.lib.umich.edu/bitstream/handle/2027.42/140872/stslocum_1.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

Natural products are bioactive molecules produced across the tree of life and have been used medicinally by humans for millennia.Isolation and characterization of these molecules reveals an incredible diversity of structure, bioactivity, and biosynthetic mechanism.These biosynthetic systems have great potential for engineering to produce novel structures, however, to take advantage of these systems they must be studied to understand their natural behavior. Polyketide synthases (PKS) are a diverse class of enzymes that integrate distinct catalytic domains into modules that are repeated to form large molecular assembly lines.These PKS ;;megaenzymes” have intricate domain movements and protein-protein interactions that enable their functionality.Investigations of several key properties of these biosynthetic systems were carried out using diverse molecular techniques. Ion-mobility mass spectrometry (IM-MS) integrates gas-phase structural data with mass detection and was used to investigate the domain movements and structure of PikAIII, a single module from the pikromycin pathway.De novo modeling was successfully carried out using these data, and IM-MS was validated as an investigative tool for the study of PKS structure and behavior.PikAIII and associated substrates were also used to investigate the docking interactions that facilitate the transfer of polyketide intermediates through the bryostatin pathway.By engineering bryostatin docking sequences into the established PikAIII chemoenzymatic system, these interactions were investigated.Bryostatins are PKS-derived natural products that are potent PKC modulators and are under investigation for several clinical applications.Two pharmacologically important vinyl methyl ester moieties are installed in the bryostatin structure by a beta-branching cassette, but several components were missing from this uncommon expansion of PKS machinery.Because bryostatins are produced by an obligate symbiont (Candidatus Endobugula sertula), investigation of their biosynthesis is difficult.However, BryT and BryU were identified in a metagenomic sequencing effort, and biochemical investigations of their activities were carried out.It was found that BryU is the donor ACP for the beta-branching cassette, BryT is the ECH1 with an unexpected product, and the unusual carboxyl O-MT was also identified.These investigations bring the potential for leveraging biosynthetic enzymes to produce novel chemical structures.