Structure and Electron-Transfer Pathway of the Human Methionine Sulfoxide Reductase MsrB3.

Gabriel Javitt,Ronen Gabizon,Efrat Resnick,Neil J Bulleid,Deborah Fass,Zhenbo Cao

doi:10.1089/ars.2020.8037

Gabriel Javitt, Ronen Gabizon + Show 4 more

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https://doi.org/10.1089/ars.2020.8037

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Abstract

Aims: The post-translational oxidation of methionine to methionine sulfoxide (MetSO) is a reversible process, enabling the repair of oxidative damage to proteins and the use of sulfoxidation as a regulatory switch. MetSO reductases catalyze the stereospecific reduction of MetSO. One of the mammalian MetSO reductases, MsrB3, has a signal sequence for entry into the endoplasmic reticulum (ER). In the ER, MsrB3 is expected to encounter a distinct redox environment compared with its paralogs in the cytosol, nucleus, and mitochondria. We sought to determine the location and arrangement of MsrB3 redox-active cysteines, which may couple MsrB3 activity to other redox events in the ER.Results: We determined the human MsrB3 structure by using X-ray crystallography. The structure revealed that a disulfide bond near the protein amino terminus is distant in space from the active site. Nevertheless, biochemical assays showed that these amino-terminal cysteines are oxidized by the MsrB3 active site after its reaction with MetSO.Innovation: This study reveals a mechanism to shuttle oxidizing equivalents from the primary MsrB3 active site toward the enzyme surface, where they would be available for further dithiol-disulfide exchange reactions.Conclusion: Conformational changes must occur during the MsrB3 catalytic cycle to transfer oxidizing equivalents from the active site to the amino-terminal redox-active disulfide. The accessibility of this exposed disulfide may help couple MsrB3 activity to other dithiol-disulfide redox events in the secretory pathway.

Highlights

The reversible modification of the sulfur-containing amino acid side chains cysteine and methionine is a common mechanism for regulation of protein function
Innovation: This study reveals a mechanism to shuttle oxidizing equivalents from the primary MsrB3 active site toward the enzyme surface, where they would be available for further dithiol-disulfide exchange reactions
The major finding from the human MsrB3 crystal structure is that the resolving cysteines, Cys3 and Cys9, form a disulfide bond and are distant from the active site in the observed conformation of the enzyme

Summary

Introduction

The reversible modification of the sulfur-containing amino acid side chains cysteine and methionine is a common mechanism for regulation of protein function. Our knowledge of how cysteine side chains are oxidized and reduced is well advanced, but we are only beginning to appreciate how methionine side chains are reversibly modified [23]. Methionine in proteins can be oxidized to methionine sulfoxide (MetSO) either nonspecifically by reactive oxygen species or as a specific, enzyme-catalyzed post-translational modification. Two main classes of methionine sulfoxide reductase (Msr) enzymes catalyze MetSO reduction to restore the unmodified methionine: MsrA enzymes reduce the S epimer, and MsrB enzymes reduce the R epimer. As an example of regulation using this chemistry, enzymes of the Mical family stereoselectively oxidize methionine in actin, resulting in polymer disassembly [19], and cytosolic MsrB activity counteracts this process by reducing the actin MetSOs [28].

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