Crystal Structures of Human HMG-CoA Synthase Isoforms Provide Insights into Inherited Ketogenesis Disorders and Inhibitor Design

Shafqat, N.; Turnbull, A.P.; Zschocke, Johannes; Oppermann, U.; Yue, Wyatt W.

doi:10.1016/j.jmb.2010.03.034

Cited by 60 publications

(84 citation statements)

References 37 publications

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“…The covalent enzyme-product complex is hydrolyzed to release HMG-CoA. HMGCS is well characterized, including crystal structures of complexes between the enzyme and its product, substrates, and inhibitors (9)(10)(11)(12)(13)(14).…”

Section: Significancementioning

confidence: 99%

Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate

Maloney

Gerwick

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Alkyl branching at the β position of a polyketide intermediate is an important variation on canonical polyketide natural product biosynthesis. The branching enzyme, 3-hydroxy-3-methylglutaryl synthase (HMGS), catalyzes the aldol addition of an acyl donor to a β-keto-polyketide intermediate acceptor. HMGS is highly selective for two specialized acyl carrier proteins (ACPs) that deliver the donor and acceptor substrates. The HMGS from the curacin A biosynthetic pathway (CurD) was examined to establish the basis for ACP selectivity. The donor ACP (CurB) had high affinity for the enzyme (K d = 0.5 μM) and could not be substituted by the acceptor ACP. Highresolution crystal structures of HMGS alone and in complex with its donor ACP reveal a tight interaction that depends on exquisite surface shape and charge complementarity between the proteins. Selectivity is explained by HMGS binding to an unusual surface cleft on the donor ACP, in a manner that would exclude the acceptor ACP. Within the active site, HMGS discriminates between pre-and postreaction states of the donor ACP. The free phosphopantetheine (Ppant) cofactor of ACP occupies a conserved pocket that excludes the acetyl-Ppant substrate. In comparison with HMG-CoA (CoA) synthase, the homologous enzyme from primary metabolism, HMGS has several differences at the active site entrance, including a flexible-loop insertion, which may account for the specificity of one enzyme for substrates delivered by ACP and the other by CoA.natural products | polyketide synthase | curacin | HMG synthase | acyl carrier protein P olyketides are a large and chemically diverse group of natural products that includes many pharmaceuticals with a broad range of biological activities and applications as antibiotics, antifungals, antiinflammatory drugs, and cancer chemotherapeutic agents (1). Polyketide synthase (PKS) biosynthetic pathways are subjects of efforts to engineer diversification of natural products in pursuit of pharmaceutical leads and compounds of industrial importance (2). They are rich sources for development of chemoenzymatic catalysts based on PKS enzymes with unusual catalytic activities.Modular type I PKS pathways, among the most versatile of nature's systems, are biosynthetic assembly lines composed of modules that act in a defined sequence to produce complex products with a variety of functional groups and chiral centers. Each module is a set of fused catalytic domains that extend and modify a polyketide intermediate. Biosynthesis proceeds from intermediates tethered to acyl carrier protein (ACP) domains via a thioester link to a phosphopantetheine (Ppant) cofactor. A ketosynthase (KS) domain catalyzes extension of the intermediate, and subsequent modification domains typically catalyze reduction and/or methylation of the β-keto (3-keto) extension product. Beyond the enzymes for these core reactions, many PKS pathways also include other catalytic functionality. Among the most interesting of these noncanonical capabilities is alkylation at the β position by a set of β...

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Section: Significancementioning

confidence: 99%

Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate

Maloney

Gerwick

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Human HMCS2 deficiency (OMIM 605991) was first described in 1997 (Thompson et al 1997), and a small number of patients have subsequently been described with mutations identified in the HMGCS2 gene that encodes HMCS2 (Aledo et al 2001(Aledo et al , 2006Bouchard et al 2001;Ramos et al 2013;Shafqat et al 2010;Wolf et al 2003). HMCS2 deficiency is characterized by episodes of severe hypoglycemia and metabolic acidosis that occur during fasting and can progress rapidly to life-threatening coma.…”

Section: Introductionmentioning

confidence: 99%

Mitochondrial 3‐hydroxy‐3‐methylglutaryl‐CoA synthase deficiency: urinary organic acid profiles and expanded spectrum of mutations

Pitt

Peters

Boneh

et al. 2014

J of Inher Metab Disea

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Mitochondrial 3-hydroxy-3-methylglutaryl CoA synthase (HMCS2) deficiency results in episodes of hypoglycemia and increases in fatty acid metabolites. Metabolite abnormalities described to date in HMCS2 deficiency are nonspecific and overlap with other inborn errors of metabolism, making the biochemical diagnosis of HMCS2 deficiency difficult. Urinary organic acid profiles from periods of metabolic decompensation were studied in detail in HMCS2-deficient patients from four families. An additional six unrelated patients were identified from clinical presentation and/or qualitative identification of abnormal organic acids. The diagnosis was confirmed by sequencing and deletion/duplication analysis of the HMGCS2 gene. Seven related novel organic acids were identified in urine profiles. Five of them (3,5-dihydroxyhexanoic 1,5 lactone; trans-5-hydroxyhex-2-enoate; 4-hydroxy-6-methyl-2-pyrone; 5-hydroxy-3-ketohexanoate; 3,5-dihydroxyhexanoate) were identified by comparison with synthesized or commercial authentic compounds. We provisionally identified trans-3-hydroxyhex-4-enoate and 3-hydroxy-5-ketohexanoate by their mass spectral characteristics. These metabolites were found in samples taken during periods of decompensation and normalized when patients recovered. When cutoffs of adipic >200 and 4-hydroxy-6-methyl-2-pyrone >20 μmol/mmol creatinine were applied, all eight samples taken from five HMCS2-deficient patients during episodes of decompensation were flagged with a positive predictive value of 80% (95% confidence interval 35-100%). Some ketotic patients had increased 4-hydroxy-6-methyl-2-pyrone. Molecular studies identified a total of 12 novel mutations, including a large deletion of HMGCS2 exon 1 in two families, highlighting the need to perform quantitative gene analyses. There are now 26 known HMGCS2 mutations, which are reviewed in the text. 4-Hydroxy-6-methyl-2-pyrone and related metabolites are markers for HMCS2 deficiency. Detection of these metabolites will streamline the biochemical diagnosis of this disorder.

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“…Human HMGCS is compartmentalised into the cytosolic isoform HMGCS1 which participates in the MVA pathway, and the mitochondrial isoform HMGCS2 where mutation results in an inherited enzyme deficiency (OMIM ID 605911). We have recently determined the structures of HMGCS1 and HMGCS2 with bound reaction products in the active site (Shafqat et al 2010), revealing subtle amino acid differences surrounding the active site when compared with previously reported structures from the bacterial and plant HMGCS proteins. The amino acid side-chains of these non-conserved residues generate different surface features among the different orthologues and give rise to isoform-specific or organism-specific 'sub-pockets' around their active sites (e.g.…”

Section: Structure Determination Of Metabolic Enzymesmentioning

confidence: 94%

High‐throughput structural biology of metabolic enzymes and its impact on human diseases

Yue

Oppermann

2011

J of Inher Metab Disea

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The Structural Genomics Consortium (SGC) is a public-private partnership that aims to determine the three-dimensional structures of human proteins of medical relevance and place them into the public domain without restriction. To date, the Oxford Metabolic Enzyme Group at SGC has deposited the structures of more than 140 human metabolic enzymes from diverse protein families such as oxidoreductases, hydrolases, oxygenases and fatty acid transferases. A subset of our target proteins are involved in the inherited disorders of carbohydrate, fatty acid, amino acid and vitamin metabolism. This article will provide an overview of the structural data gathered from our high-throughput efforts and the lessons learnt in the structure-function relationship of these enzymes, small molecule development and the molecular basis of disease mutations.

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Crystal Structures of Human HMG-CoA Synthase Isoforms Provide Insights into Inherited Ketogenesis Disorders and Inhibitor Design

Cited by 60 publications

References 37 publications

Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate

Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate

Mitochondrial 3‐hydroxy‐3‐methylglutaryl‐CoA synthase deficiency: urinary organic acid profiles and expanded spectrum of mutations

High‐throughput structural biology of metabolic enzymes and its impact on human diseases

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