Insights into Function, Catalytic Mechanism, and Fold Evolution of Selenoprotein Methionine Sulfoxide Reductase B1 through Structural Analysis*

Finn L Aachmann,Lena S Sal,Hwa-Young Kim,Stefano M Marino,Vadim N Gladyshev,Alexander Dikiy

doi:10.1074/jbc.m110.132308

Abstract

Methionine sulfoxide reductases protect cells by repairing oxidatively damaged methionine residues in proteins. Here, we report the first three-dimensional structure of the mammalian selenoprotein methionine sulfoxide reductase B1 (MsrB1), determined by high resolution NMR spectroscopy. Heteronuclear multidimensional spectra yielded NMR spectral assignments for the reduced form of MsrB1 in which catalytic selenocysteine (Sec) was replaced with cysteine (Cys). MsrB1 consists of a central structured core of two β-sheets and a highly flexible, disordered N-terminal region. Analysis of pH dependence of NMR signals of catalytically relevant residues, comparison with the data for bacterial MsrBs, and NMR-based structural analysis of methionine sulfoxide (substrate) and methionine sulfone (inhibitor) binding to MsrB1 at the atomic level reveal a mechanism involving catalytic Sec(95) and resolving Cys(4) residues in catalysis. The MsrB1 structure differs from the structures of Cys-containing MsrBs in the use of distal selenenylsulfide, residues needed for catalysis, and the mode in which the active form of the enzyme is regenerated. In addition, this is the first structure of a eukaryotic zinc-containing MsrB, which highlights the structural role of this metal ion bound to four conserved Cys. We integrated this information into a structural model of evolution of MsrB superfamily.

Highlights

Dative stress, but can be enzymatically reduced back to methionine by Methionine sulfoxide (MetSO) reductases (Msrs)
Structure Description—methionine sulfoxide reductase B1 (MsrB1) solution structure was calculated based on the NOE-derived geometrical constraints, dihedral angles obtained from TALOS [20], and distance constraints for the tetrahedral Zn2ϩ ion derived from x-ray structures of proteins containing zinc coordinated by Cys residues [21,22,23]
The catalytic site of MsrB1 is composed of Sec/Cys95, Cys4 and Trp43, His80, Phe82, Asp83, Arg93, and Phe97, which assist in the reduction of the oxidized methionine

Summary

Introduction

Dative stress, but can be enzymatically reduced back to methionine by MetSO reductases (Msrs). This observation is remaining Cys, Cys4, is situated in the mobile N-terminal consistent with the 15N-{H} NOE and 15N backbone relaxation region, which is flexible enough to flip in and form the proposed data, even if the latter are not very well defined for some protein selenenylsulfide bond with the catalytically active Sec95 in the regions (supplemental Fig. S1).

Results

Conclusion