Transition Path Sampling Study of the Feruloyl Esterase Mechanism

Author(s)

Silveira, Rodrigo L. ^{[1, 2, 3, 4]} ; Knott, Brandon C. ^[3] ; Pereira, Caroline S. ^{[1, 2, 3]} ; Crowley, Michael F. ^[3] ; Skaf, Munir S. ^{[1, 2]} ; Beckham, Gregg T. ^[3]

Affiliation

^[1] Univ Estadual Campinas, Inst Chem, BR-13084862 Campinas, SP - Brazil

^[2] Univ Estadual Campinas, Ctr Comp Engn & Sci, BR-13084862 Campinas, SP - Brazil

^[3] Natl Renewable Energy Lab, Renewable Resources & Enabling Sci Ctr, Golden, CO 80401 - USA

^[4] Univ Fed Rio de Janeiro, Inst Chem, BR-21941909 Rio De Janeiro, RJ - Brazil

• Document type: Journal article
• Source: Journal of Physical Chemistry B; v. 125, n. 8, p. 2018-2030, MAR 4 2021.
• Web of Science Citations: 0

Abstract

Serine hydrolases cleave peptide and ester bonds and are ubiquitous in nature, with applications in biotechnology, in materials, and as drug targets. The serine hydrolase two-step mechanism employs a serine-histidine-aspartate/glutamate catalytic triad, where the histidine residue acts as a base to activate poor nucleophiles (a serine residue or a water molecule) and as an acid to allow the dissociation of poor leaving groups. This mechanism has been the subject of debate regarding how histidine shuttles the proton from the nucleophile to the leaving group. To elucidate the reaction mechanism of serine hydrolases, we employ quantum mechanics/molecular mechanics-based transition path sampling to obtain the reaction coordinate using the Aspergillus niger feruloyl esterase A (AnFaeA) as a model enzyme. The optimal reaction coordinates include terms involving nucleophilic attack on the carbonyl carbon and proton transfer to, and dissociation of, the leaving group. During the reaction, the histidine residue undergoes a reorientation on the time scale of hundreds of femtoseconds that supports the ``moving histidine{''} mechanism, thus calling into question the ``ring flip{''} mechanism. We find a concerted mechanism, where the transition state coincides with the tetrahedral intermediate with the histidine residue pointed between the nucleophile and the leaving group. Moreover, motions of the catalytic aspartate toward the histidine occur concertedly with proton abstraction by the catalytic histidine and help stabilize the transition state, thus partially explaining how serine hydrolases enable poor nucleophiles to attack the substrate carbonyl carbon. Rate calculations indicate that the second step (deacylation) is rate-determining, with a calculated rate constant of 66 s(-1). Overall, these results reveal the pivotal role of active-site dynamics in the catalytic mechanism of AnFaeA, which is likely similar in other serine hydrolases. (AU)

• FAPESP's process: 15/25031-1 - Molecular Dynamics of Carbohydrate Modifying Enzymes for Lignocellulosic Biomass Deconstruction and Valorization
• Grantee: Caroline Simões Pereira
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 17/01151-3 - Transition path sampling QM/MM simulations of lytic transglycosylases
• Grantee: Caroline Simões Pereira
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• FAPESP's process: 16/22956-7 - Hybrid QM/MM simulations of feruloyl esterases: cleavage mechanism of lignin-carbohydrate complexes in plant cell walls
• Grantee: Rodrigo Leandro Silveira
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• FAPESP's process: 13/08293-7 - CCES - Center for Computational Engineering and Sciences
• Grantee: Munir Salomao Skaf
• Support Opportunities: Research Grants - Research, Innovation and Dissemination Centers - RIDC
• FAPESP's process: 14/10448-1 - Molecular aspects of plant cell wall architecture
• Grantee: Rodrigo Leandro Silveira
• Support Opportunities: Scholarships in Brazil - Post-Doctoral