Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate

Acyltransferases determine which extender units are incorporated into polyketide and fatty acid products. The ping-pong acyltransferase mechanism utilizes a serine in a conserved GHSxG motif. However, the role of the conserved histidine in this motif is poorly understood. We observed that a histidin...

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Main Authors:	Poust, Sean, Yoon, Isu, Adams, Paul D., Katz, Leonard, Petzold, Christopher J., Keasling, Jay D.
Format:	Online
Language:	English
Published:	Public Library of Science 2014
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4186864/

id	pubmed-4186864
recordtype	oai_dc
spelling	pubmed-41868642014-10-16 Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate Poust, Sean Yoon, Isu Adams, Paul D. Katz, Leonard Petzold, Christopher J. Keasling, Jay D. Research Article Acyltransferases determine which extender units are incorporated into polyketide and fatty acid products. The ping-pong acyltransferase mechanism utilizes a serine in a conserved GHSxG motif. However, the role of the conserved histidine in this motif is poorly understood. We observed that a histidine to alanine mutation (H640A) in the GHSxG motif of the malonyl-CoA specific yersiniabactin acyltransferase results in an approximately seven-fold higher hydrolysis rate over the wildtype enzyme, while retaining transacylation activity. We propose two possibilities for the reduction in hydrolysis rate: either H640 structurally stabilizes the protein by hydrogen bonding with a conserved asparagine in the ferredoxin-like subdomain of the protein, or a water-mediated hydrogen bond between H640 and the malonyl moiety stabilizes the malonyl-O-AT ester intermediate. Public Library of Science 2014-10-06 /pmc/articles/PMC4186864/ /pubmed/25286165 http://dx.doi.org/10.1371/journal.pone.0109421 Text en © 2014 Poust et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Poust, Sean Yoon, Isu Adams, Paul D. Katz, Leonard Petzold, Christopher J. Keasling, Jay D.
spellingShingle	Poust, Sean Yoon, Isu Adams, Paul D. Katz, Leonard Petzold, Christopher J. Keasling, Jay D. Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
author_facet	Poust, Sean Yoon, Isu Adams, Paul D. Katz, Leonard Petzold, Christopher J. Keasling, Jay D.
author_sort	Poust, Sean
title	Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
title_short	Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
title_full	Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
title_fullStr	Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
title_full_unstemmed	Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate
title_sort	understanding the role of histidine in the ghsxg acyltransferase active site motif: evidence for histidine stabilization of the malonyl-enzyme intermediate
description	Acyltransferases determine which extender units are incorporated into polyketide and fatty acid products. The ping-pong acyltransferase mechanism utilizes a serine in a conserved GHSxG motif. However, the role of the conserved histidine in this motif is poorly understood. We observed that a histidine to alanine mutation (H640A) in the GHSxG motif of the malonyl-CoA specific yersiniabactin acyltransferase results in an approximately seven-fold higher hydrolysis rate over the wildtype enzyme, while retaining transacylation activity. We propose two possibilities for the reduction in hydrolysis rate: either H640 structurally stabilizes the protein by hydrogen bonding with a conserved asparagine in the ferredoxin-like subdomain of the protein, or a water-mediated hydrogen bond between H640 and the malonyl moiety stabilizes the malonyl-O-AT ester intermediate.
publisher	Public Library of Science
publishDate	2014
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4186864/
_version_	1613141335051599872

Understanding the Role of Histidine in the GHSxG Acyltransferase Active Site Motif: Evidence for Histidine Stabilization of the Malonyl-Enzyme Intermediate

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