Structure-function investigation of 3-methylaspartate ammonia lyase reveals substrate molecular determinants for the deamination reaction

Veronica Saez-Jimenez; Željka Sanader Maršić; Matteo Lambrughi; Jae Ho Shin; Robin van Havere; Elena Papaleo; Lisbeth Olsson; Valeria Mapelli

doi:10.1371/journal.pone.0233467

Structure-function investigation of 3-methylaspartate ammonia lyase reveals substrate molecular determinants for the deamination reaction

PLoS One. 2020 May 21;15(5):e0233467. doi: 10.1371/journal.pone.0233467. eCollection 2020.

Authors

Veronica Saez-Jimenez¹, Željka Sanader Maršić², Matteo Lambrughi², Jae Ho Shin¹, Robin van Havere¹, Elena Papaleo², Lisbeth Olsson¹, Valeria Mapelli¹

Affiliations

¹ Division of Industrial Biotechnology, Department of Biology and Biological Engineering, Chalmers University of Technology, Gothenburg, Sweden.
² Computational Biology Laboratory, Danish Cancer Society Research Center, Copenhagen, Denmark.

Abstract

The enzymatic reactions leading to the deamination of β-lysine, lysine, or 2-aminoadipic acid are of great interest for the metabolic conversion of lysine to adipic acid. Enzymes able to carry out these reactions are not known, however ammonia lyases (EC 4.3.1.-) perform deamination on a wide range of substrates. We have studied 3-methylaspartate ammonia lyase (MAL, EC 4.3.1.2) as a potential candidate for protein engineering to enable deamination towards β-lysine, that we have shown to be a competitive inhibitor of MAL. We have characterized MAL activity, binding and inhibition properties on six different compounds that would allow to define the molecular determinants necessary for MAL to deaminate our substrate of interest. Docking calculations showed that β-lysine as well as the other compounds investigated could fit spatially into MAL catalytic pocket, although they probably are weak or very transient binders and we identified molecular determinants involved in the binding of the substrate. The hydrophobic interactions formed by the methyl group of 3-methylaspartic acid, together with the presence of the amino group on carbon 2, play an essential role in the appropriate binding of the substrate. The results showed that β-lysine is able to fit and bind in MAL catalytic pocket and can be potentially converted from inhibitor to substrate of MAL upon enzyme engineering. The characterization of the binding and inhibition properties of the substrates tested here provide the foundation for future and more extensive studies on engineering MAL that could lead to a MAL variant able to catalyse this challenging deamination reaction.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Ammonia-Lyases / metabolism*
Binding Sites
Deamination
Models, Molecular*
Molecular Docking Simulation
Protein Conformation
Structure-Activity Relationship

Substances

Ammonia-Lyases
methylaspartate ammonia-lyase

Grants and funding

The study was funded by The Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS) (project number 942-2015-1628 awarded to LO and VM), Danmarks Grundforskningsfond (DNRF125) and the NovoNordisk Foundation under the program for Biotechnology-based Synthesis and Production Research (reference number NNF-17OC0027588, awarded to LO and EP). The calculations described in this paper were performed using the DeiC National Life Science Supercomputer Computerome at DTU (Denmark) and resources provided by the Swedish National Infrastructure for Computing (SNIC) at PDC Center for High Performance Computing under the project SNIC 2017/1-537. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.