The Uncommon Enzymology of Cis-Acyltransferase Assembly Lines

Keatinge‐Clay, Adrian T.

doi:10.1021/acs.chemrev.6b00683

Cited by 90 publications

(109 citation statements)

References 257 publications

(586 reference statements)

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“…However, a conformational change would be necessary since the side chain of H” is observed on the surface of the DH domain. Another hypothesis for double dehydration is that the dehydration of the δ-hydroxyl group occurs before reduction of the β-keto group, similar to catalysis by the final DH in the ajudazol assembly line (Buntin et al, 2010; Keatinge-Clay, 2017) (Route 2 in Figure 3b). If this mechanism were operative, the ketone oxygen could insert into the same oxyanion hole as the thioester oxygen during the canonical dehydration step.…”

Section: Resultsmentioning

confidence: 98%

“…These PKSs have been distinguished as cis -acyltransferase and trans -acyltransferase ( cis -AT and trans -AT) PKSs based on the reliance of the trans -AT PKSs upon separately-encoded ATs for selecting α-carboxyacyl-CoA polyketide building blocks. While cis -AT PKSs, such as the erythromycin PKS, have been fairly well characterized both structurally and functionally, the trans -AT PKSs are largely uncharacterized due to their more recent discovery (Keatinge-Clay, 2012; Keatinge-Clay, 2017, Piel, 2010; Helfrich and Piel 2016). Beyond the eponymous distinction between these synthase types, trans -AT PKSs apparently deviate from the colinearity rules much more often than cis -AT PKSs, such that the polyketides produced by trans -AT PKSs cannot be confidently predicted by inspecting which enzymes are present within its modules [e.g., ketosynthase (KS), methyltransferase (MT), ketoreductase (KR), dehydratase (DH), enoylreductase (ER)] (Nguyen et al 2008).…”

Section: Introductionmentioning

confidence: 99%

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Structural and Functional Trends in Dehydrating Bimodules from trans-Acyltransferase Polyketide Synthases

et al. 2017

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SUMMARY In an effort to uncover the structural motifs and biosynthetic logic of the relatively uncharacterized trans-acyltransferase polyketide synthases, we have begun the dissection of the enigmatic dehydrating bimodules common in these enzymatic assembly lines. We report the 1.98 Å-resolution structure of a ketoreductase (KR) from the first half of a type A dehydrating bimodule and the 2.22 Å-resolution structure of a dehydratase (DH) from the second half of a type B dehydrating bimodule. The KR, from the third module of the bacillaene synthase, and the DH, from the tenth module of the difficidin synthase, possess features not observed in structurally-characterized homologs. The DH architecture provides clues for how it catalyzes a unique double dehydration. Correlations between the chemistries proposed for dehydrating bimodules and bioinformatic analysis indicate that type A dehydrating bimodules generally produce an α/β-cis alkene moiety while type B dehydrating bimodules generally produce an α/β-trans, γ/δ-cis diene moiety.

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Section: Resultsmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

Structural and Functional Trends in Dehydrating Bimodules from trans-Acyltransferase Polyketide Synthases

et al. 2017

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“…Several hybrid assembly lines composed of both trans ‐AT and cis ‐AT modules are now known. The chivosazole, enacyloxin, kirromycin, and nosperin assembly lines are primarily comprised of trans ‐AT modules, the ambruticin, DKxanthene, and pyoluteorin assembly lines are primarily comprised of cis ‐AT modules, and the neocarzilin assembly line contains an equal mix of trans ‐AT and cis ‐AT modules …”

Section: Resultsmentioning

confidence: 99%

The modules of trans‐acyltransferase assembly lines redefined with a central acyl carrier protein

Wood

Keatinge‐Clay

2018

Proteins

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Here, the term "module" is redefined for trans-acyltransferase (trans-AT) assembly lines to agree with how its domains cooperate and evolutionarily co-migrate. The key domain in both the polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) modules of assembly lines is the acyl carrier protein (ACP). ACPs not only relay growing acyl chains through the assembly line but also collaborate with enzymes in modules, both in cis and in trans, to add a specific chemical moiety. A ketosynthase (KS) downstream of ACP often plays the role of gatekeeper, ensuring that only a single intermediate generated by the enzymes of a module is passed downstream. Bioinformatic analysis of 526 ACPs from 33 characterized trans-AT assembly lines reveals ACPs from the same module type generally clade together, reflective of the co-evolution of these domains with their cognate enzymes. While KSs downstream of ACPs from the same module type generally also clade together, KSs upstream of ACPs do not-in disagreement with the traditional definition of a module. Beyond nomenclature, the presented analysis impacts our understanding of module function, the evolution of assembly lines, pathway prediction, and assembly line engineering.

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“…Several mechanisms of pathway initiation have been described; frequently, a loading acyltransferase (AT L ) primes the ACP with an acyl group such as acetyl-, malonyl-, or methylmalonyl-CoA. An α-carboxylated acyl group can be decarboxylated by a non-extending ketosynthase domain 5 . In other cases, the loading modules select chemically diverse starter units derived from amino acids, cyclohexanecarboxylic acids, fatty acids, or branched chain CoAs 5, 6 .…”

Section: Introductionmentioning

confidence: 99%

A Mononuclear Iron-Dependent Methyltransferase Catalyzes Initial Steps in Assembly of the Apratoxin A Polyketide Starter Unit

et al. 2017

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Natural product biosynthetic pathways contain a plethora of enzymatic tools to carry out difficult biosynthetic transformations. Here we discover an unusual mononuclear iron-dependent methyltransferase that acts in the initiation steps of apratoxin A biosynthesis (AprA MT1). Fe3+-replete AprA MT1 catalyzes one or two methyl transfer reactions on the substrate malonyl-ACP (acyl carrier protein), whereas Co2+, Fe2+, Mn2+ and Ni2+ support only a single methyl transfer. MT1 homologs exist within the “GNAT” (GCN5-related N-acetyltransferase) loading modules of several modular biosynthetic pathways with propionyl, isobutyryl or pivaloyl starter units. GNAT domains are thought to catalyze decarboxylation of malonyl-CoA and acetyl transfer to a carrier protein. In AprA the GNAT domain lacks both decarboxylation and acyl transfer activity. A crystal structure of the AprA MT1-GNAT didomain with bound Mn2+, malonate and the methyl donor S-adenosylmethionine (SAM) reveals that the malonyl substrate is a bidentate metal ligand, indicating that the metal acts as a Lewis acid to promote methylation of the malonyl α-carbon. The GNAT domain is truncated relative to functional homologs. These results afford an expanded understanding of MT1-GNAT structure and activity, and permit the functional annotation of homologous GNAT loading modules both with and without methyltransferases, additionally revealing their rapid evolutionary adaptation in different biosynthetic contexts.

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The Uncommon Enzymology of Cis-Acyltransferase Assembly Lines

Cited by 90 publications

References 257 publications

Structural and Functional Trends in Dehydrating Bimodules from trans-Acyltransferase Polyketide Synthases

Structural and Functional Trends in Dehydrating Bimodules from trans-Acyltransferase Polyketide Synthases

The modules of trans‐acyltransferase assembly lines redefined with a central acyl carrier protein

A Mononuclear Iron-Dependent Methyltransferase Catalyzes Initial Steps in Assembly of the Apratoxin A Polyketide Starter Unit

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