Structural Characterization of the Reaction and Substrate Specificity Mechanisms of Pathogenic Fungal Acetyl-CoA Synthetases

Jezewski, Andrew J.; Alden, Katy M.; Esan, Taiwo E.; DeBouver, Nicholas D.; Abendroth, Jan; Bullen, Jameson C.; Calhoun, Brandy M.; Potts, Kristy T.; Murante, Daniel M.; Hagen, Timothy J.; Fox, David; Krysan, Damian J.

doi:10.1021/acschembio.1c00484

Structural Characterization of the Reaction and Substrate Specificity Mechanisms of Pathogenic Fungal Acetyl-CoA Synthetases

Journal Article · Mon Aug 09 00:00:00 EDT 2021 · ACS Chemical Biology

DOI:https://doi.org/10.1021/acschembio.1c00484· OSTI ID:1821734

Jezewski, Andrew J. ^[1]; Alden, Katy M. ^[1]; Esan, Taiwo E. ^[2]; DeBouver, Nicholas D. ^[3]; Abendroth, Jan ^[3]; Bullen, Jameson C. ^[3]; Calhoun, Brandy M. ^[3]; Potts, Kristy T. ^[3]; Murante, Daniel M. ^[1]; ^[2]; Fox, David ^[4]; ^[1]

Univ. of Iowa, Iowa City, IA (United States)
Northern Illinois Univ., DeKalb, IL (United States)
UCB Pharma, Bainbridge Island, WA (United States); Seattle Structural Genomics Center for Infectious Disease (SSGCID), WA (United States)
UCB Pharma, Bainbridge Island, WA (United States); Beryllium Discovery Corp., Bainbridge Island, WA (United States); Seattle Structural Genomics Center for Infectious Disease (SSGCID), WA (United States)

Acetyl CoA synthetases (ACSs) are Acyl-CoA/NRPS/Luciferase (ANL) superfamily enzymes that couple acetate with CoA to generate acetyl CoA, a key component of central carbon metabolism in eukaryotes and prokaryotes. Normal mammalian cells are not dependent on ACSs, while tumor cells, fungi, and parasites rely on acetate as a precursor for acetyl CoA. Consequently, ACSs have emerged as a potential drug target. As part of a program to develop antifungal ACS inhibitors, we characterized fungal ACSs from five diverse human fungal pathogens using biochemical and structural studies. ACSs catalyze a two-step reaction involving adenylation of acetate followed by thioesterification with CoA. Our structural studies captured each step of these two half-reactions including the acetyl-adenylate intermediate of the first half-reaction in both the adenylation conformation and the thioesterification conformation and thus provide a detailed picture of the reaction mechanism. We also used a systematic series of increasingly larger alkyl adenosine esters as chemical probes to characterize the structural basis of the exquisite ACS specificity for acetate over larger carboxylic acid substrates. Consistent with previous biochemical and genetic data for other enzymes, structures of fungal ACSs with these probes bound show that a key tryptophan residue limits the size of the alkyl binding site and forces larger alkyl chains to adopt high energy conformers, disfavoring their efficient binding. Together, our analysis provides highly detailed structural models for both the reaction mechanism and substrate specificity that should be useful in designing selective inhibitors of eukaryotic ACSs as potential anticancer, antifungal, and antiparasitic drugs.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: National Institute of Allergy and Infectious Diseases (NIAID); National Institutes of Health (NIH); USDOE Office of Science (SC)

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 1821734

Alternate ID(s):: OSTI ID: 1823157

Journal Information:: ACS Chemical Biology, Journal Name: ACS Chemical Biology Journal Issue: 8 Vol. 16; ISSN 1554-8929

Publisher:: American Chemical Society (ACS)Copyright Statement

Country of Publication:: United States

Language:: ENGLISH

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Structural Characterization of the Reaction and Substrate Specificity Mechanisms of Pathogenic Fungal Acetyl-CoA Synthetases

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