Gabriel Javitt1, Zhenbo Cao2, Efrat Resnick3, Ronen Gabizon4, Neil Bulleid5, Deborah Fass6. 1. Weizmann Institute of Science, Structural Biology, Rehovot, Israel; gabriel.javitt@weizmann.ac.il. 2. University of Glasgow, 3526, Institute of Molecular, Cellular and Systems Biology, Glasgow, Glasgow, United Kingdom of Great Britain and Northern Ireland; Zhenbo.cao@glasgow.ac.uk. 3. Weizmann Institute of Science, Organic Chemistry, Rehovot, Israel; efrat.resnick@weizmann.ac.il. 4. Weizmann Institute of Science, Organic Chemistry, Rehovot, Israel; ronen.gabizon@weizmann.ac.il. 5. University of Glasgow, 3526, Institute of Molecular, Cellular and Systems Biology, Glasgow, Glasgow, United Kingdom of Great Britain and Northern Ireland; neil.bulleid@glasgow.ac.uk. 6. Weizmann Institute of Science, Structural Biology, Department of Structural Biology, Weizmann Institute of Science, Rehovot, Israel, 76100; deborah.fass@weizmann.ac.il.
Abstract
INTRODUCTION: The post-translational oxidation of methionine to methionine sulfoxide is a reversible process, enabling repair of oxidative damage to proteins and the use of sulfoxidation as a regulatory switch. Methionine sulfoxide reductases catalyze the stereospecific reduction of methionine sulfoxide. One of the mammalian methionine sulfoxide 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 to its paralogs in the cytosol, nucleus, and mitochondria. AIMS: 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 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 methionine sulfoxide. 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.
INTRODUCTION: The post-translational oxidation of methionine to methionine sulfoxide is a reversible process, enabling repair of oxidative damage to proteins and the use of sulfoxidation as a regulatory switch. Methionine sulfoxide reductases catalyze the stereospecific reduction of methionine sulfoxide. One of the mammalian methionine sulfoxide 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 to its paralogs in the cytosol, nucleus, and mitochondria. AIMS: 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 humanMsrB3 structure 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 methionine sulfoxide. 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.
Authors: Tomasz Pędzinski; Katarzyna Grzyb; Konrad Skotnicki; Piotr Filipiak; Krzysztof Bobrowski; Chryssostomos Chatgilialoglu; Bronislaw Marciniak Journal: Int J Mol Sci Date: 2021-04-30 Impact factor: 5.923