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Title: Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography

Abstract

Glycoside hydrolases (GHs) commonly use the retaining or inverting mechanisms to hydrolyze carbohydrates, and the rates of catalysis are usually pH dependent. Deeper understanding of these pH-dependent reaction mechanisms is of great importance for protein engineering and drug design. We used high-resolution X-ray crystallography to analyze the sugar ring configurations of an oligosaccharide ligand during hydrolysis for the family 11 GH, and the results support the 1S3 → 4H3 → 4C1 conformational itinerary. These results indicate that sugar ring flexibility may help to distort and break the glycosidic bond. Constant pH molecular dynamics simulations and neutron crystallography demonstrate that the catalytic glutamate residue (E177) has alternate conformational changes to transfer a proton to cleave the glycosidic bond. Furthermore, a neutron crystallography analysis shows that the H-bond length between E177 and its nearby tyrosine residue (Y88) is shortened when the pH increases, preventing E177 from rotating downward and obtaining a proton from the solvent for catalysis. This result indicates that the H-bond length variation may play a key role in the pH-dependent reaction mechanism. Here, our results demonstrate that both sugar ring flexibility and protein dynamics are important in the pH-dependent reaction mechanism and may help to engineer GHs with differentmore » pH optima.« less

Authors:
 [1];  [1];  [1];  [1];  [1];  [1];  [1];  [1]; ORCiD logo [2]; ORCiD logo [3]; ORCiD logo [3];  [4]; ORCiD logo [1]
+ Show Author Affiliations
  1. Nanjing Agricultural Univ., Nanjing (People’s Republic of China)
  2. Forschungszentrum Jülich GmbH, Garching (Germany)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  4. Jimei Univ., Xiamen (People’s Republic of China)
Publication Date:
Research Org.:
Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1509581
Grant/Contract Number:  
AC05-00OR22725
Resource Type:
Accepted Manuscript
Journal Name:
ACS Catalysis
Additional Journal Information:
Journal Volume: 8; Journal Issue: 9; Journal ID: ISSN 2155-5435
Publisher:
American Chemical Society (ACS)
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; constant pH molecular dynamics; glycoside hydrolase; H-bond length; neutron crystallography; pH-dependent reaction mechanism

Citation Formats

Li, Zhihong, Zhang, Xiaoshuai, Wang, Qingqing, Li, Chunran, Zhang, Nianying, Zhang, Xinkai, Xu, Birui, Ma, Baoliang, Schrader, Tobias E., Coates, Leighton, Kovalevsky, Andrey, Huang, Yandong, and Wan, Qun. Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography. United States: N. p., 2018. Web. doi:10.1021/acscatal.8b01472.
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Li, Zhihong, Zhang, Xiaoshuai, Wang, Qingqing, Li, Chunran, Zhang, Nianying, Zhang, Xinkai, Xu, Birui, Ma, Baoliang, Schrader, Tobias E., Coates, Leighton, Kovalevsky, Andrey, Huang, Yandong, & Wan, Qun. Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography. United States. https://doi.org/10.1021/acscatal.8b01472
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Li, Zhihong, Zhang, Xiaoshuai, Wang, Qingqing, Li, Chunran, Zhang, Nianying, Zhang, Xinkai, Xu, Birui, Ma, Baoliang, Schrader, Tobias E., Coates, Leighton, Kovalevsky, Andrey, Huang, Yandong, and Wan, Qun. Wed . "Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography". United States. https://doi.org/10.1021/acscatal.8b01472. https://www.osti.gov/servlets/purl/1509581.
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@article{osti_1509581,
title = {Understanding the pH-Dependent Reaction Mechanism of a Glycoside Hydrolase Using High-Resolution X-ray and Neutron Crystallography},
author = {Li, Zhihong and Zhang, Xiaoshuai and Wang, Qingqing and Li, Chunran and Zhang, Nianying and Zhang, Xinkai and Xu, Birui and Ma, Baoliang and Schrader, Tobias E. and Coates, Leighton and Kovalevsky, Andrey and Huang, Yandong and Wan, Qun},
abstractNote = {Glycoside hydrolases (GHs) commonly use the retaining or inverting mechanisms to hydrolyze carbohydrates, and the rates of catalysis are usually pH dependent. Deeper understanding of these pH-dependent reaction mechanisms is of great importance for protein engineering and drug design. We used high-resolution X-ray crystallography to analyze the sugar ring configurations of an oligosaccharide ligand during hydrolysis for the family 11 GH, and the results support the 1S3 → 4H3 → 4C1 conformational itinerary. These results indicate that sugar ring flexibility may help to distort and break the glycosidic bond. Constant pH molecular dynamics simulations and neutron crystallography demonstrate that the catalytic glutamate residue (E177) has alternate conformational changes to transfer a proton to cleave the glycosidic bond. Furthermore, a neutron crystallography analysis shows that the H-bond length between E177 and its nearby tyrosine residue (Y88) is shortened when the pH increases, preventing E177 from rotating downward and obtaining a proton from the solvent for catalysis. This result indicates that the H-bond length variation may play a key role in the pH-dependent reaction mechanism. Here, our results demonstrate that both sugar ring flexibility and protein dynamics are important in the pH-dependent reaction mechanism and may help to engineer GHs with different pH optima.},
doi = {10.1021/acscatal.8b01472},
journal = {ACS Catalysis},
number = 9,
volume = 8,
place = {United States},
year = {Wed Jul 18 00:00:00 EDT 2018},
month = {Wed Jul 18 00:00:00 EDT 2018}
}
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