Structure, Biochemistry, and Substrate Selectivity of the Hydroxymethylglutaryl Synthase of Polyketide-Branching

Natural Products;Curacin;Polyketide Synthase;Structural Biology;HMG Synthase;Acyl Carrier Protein;Biological Chemistry;Science;Chemical Biology

Maloney, FinnTesmer, John J G ;

University of Michigan

关键词: Natural Products;    Curacin;    Polyketide Synthase;    Structural Biology;    HMG Synthase;    Acyl Carrier Protein;    Biological Chemistry;    Science;    Chemical Biology;   

Others  :  https://deepblue.lib.umich.edu/bitstream/handle/2027.42/140829/fpmalone_1.pdf?sequence=1&isAllowed=y

瑞士|英语

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

【 摘 要 】

Modular polyketide synthase (PKS) pathways generate a diverse array of pharmaceutically significant small molecule natural products, and synthetic PKS biology may facilitate pharmaceutical development, production of industrially important compounds, and discovery of chemoenzymatic reagents. Of interest to these efforts are biosynthetic schemes that install unusual functional groups, including ;;beta-branching” enzymes that generate alkyl substituents found in some polyketides. PKS enzymes act on substrates linked to acyl carrier proteins (ACP) via a phosphopantetheine arm (Ppant), but beta-branching enzymes are further selective for an intermediate linked to a specialized, branch-acceptor ACP (ACPA). ACP-enzyme interactions are a poorly understood facet of PKS biology and selectivity of β-branching enzymes for ACPA is essential for fidelity of the biosynthetic pathway.A hydroxymethylglutaryl synthase (HMGS) initiates beta-branching using an acetyl nucleophile that is delivered to HMGS by a distinct, branch-donor ACP (ACPD). This thesis summarizes research into the structural basis for the distinct selectivity of HMGS for its acetyl-ACPD and polyketide-ACPA substrates. We solved crystal structures of HMGS and determined features that both distinguish it from its primary metabolism homolog and are involved in ACP interaction. Structures of the ACPD/HMGS complex revealed that ACPD recognition is dependent on electrostatic interactions and on unique structural features of ACPD. In these first structures of a natively bound PKS ACP/enzyme complex, we discovered that ACPD Ppant positioning is substrate dependent and identified distinct pre- and post-reaction positions of the Ppant. Furthermore, differences in the ACPA and ACPD interactions with HMGS apparently result in different Ppant positions that we selectively disrupted with active site substitutions. Finally, we demonstrated that HMGS is reactive with a non-natural donor-substrate, which has promising implications for the use of HMGS in synthetic biology and chemoenzymatic applications.