Understanding the role of histidine in the GHSxG acyltransferase active site motif: Evidence for histidine stabilization of the malonyl-enzyme intermediate
Abstract
Acyltransferases determine which extender units are incorporated into polyketide and fatty acid products. Thus, 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.
- Authors:
-
- Univ. of California, Berkeley, CA (United States)
- Joint BioEnergy Institute, Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
- Synthetic Biology Engineering Research Center, Emeryville, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States)
- Univ. of California, Berkeley, CA (United States); Synthetic Biology Engineering Research Center, Emeryville, CA (United States); Joint BioEnergy Institute, Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
- Publication Date:
- Research Org.:
- Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)
- Sponsoring Org.:
- USDOE Advanced Research Projects Agency - Energy (ARPA-E); USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1211510
- Grant/Contract Number:
- 0000206-1577; AC02-05CH11231
- Resource Type:
- Accepted Manuscript
- Journal Name:
- PLoS ONE
- Additional Journal Information:
- Journal Volume: 9; Journal Issue: 10; Journal ID: ISSN 1932-6203
- Publisher:
- Public Library of Science
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; hydrolysis; histidine; sequence motif analysis; hydrogen bonding; serine; alanine; esters; fatty acids
Citation Formats
Poust, Sean, Yoon, Isu, Adams, Paul D., Katz, Leonard, Petzold, Christopher J., and Keasling, Jay D. Understanding the role of histidine in the GHSxG acyltransferase active site motif: Evidence for histidine stabilization of the malonyl-enzyme intermediate. United States: N. p., 2014.
Web. doi:10.1371/journal.pone.0109421.
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. United States. https://doi.org/10.1371/journal.pone.0109421
Poust, Sean, Yoon, Isu, Adams, Paul D., Katz, Leonard, Petzold, Christopher J., and Keasling, Jay D. Mon .
"Understanding the role of histidine in the GHSxG acyltransferase active site motif: Evidence for histidine stabilization of the malonyl-enzyme intermediate". United States. https://doi.org/10.1371/journal.pone.0109421. https://www.osti.gov/servlets/purl/1211510.
@article{osti_1211510,
title = {Understanding the role of histidine in the GHSxG acyltransferase active site motif: Evidence for histidine stabilization of the malonyl-enzyme intermediate},
author = {Poust, Sean and Yoon, Isu and Adams, Paul D. and Katz, Leonard and Petzold, Christopher J. and Keasling, Jay D.},
abstractNote = {Acyltransferases determine which extender units are incorporated into polyketide and fatty acid products. Thus, 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.},
doi = {10.1371/journal.pone.0109421},
journal = {PLoS ONE},
number = 10,
volume = 9,
place = {United States},
year = {Mon Oct 06 00:00:00 EDT 2014},
month = {Mon Oct 06 00:00:00 EDT 2014}
}
Web of Science
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Works referencing / citing this record:
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Structural and Biochemical Insight into the Recruitment of Acyl Carrier Protein‐Linked Extender Units in Ansamitocin Biosynthesis
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Control Mechanism for cis Double-Bond Formation by Polyunsaturated Fatty-Acid Synthases
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