<i>Rhodobacter</i> <i>sphaeroides</i> methionine sulfoxide reductase P reduces <i>R</i> - and <i>S</i> -diastereomers of methionine sulfoxide from a broad-spectrum of protein substrates

Abstract

Methionine (Met) is prone to oxidation and can be converted to Met sulfoxide (MetO), which exists as <i>R</i>- and <i>S</i>-diastereomers. MetO can be reduced back to Met by the ubiquitous methionine sulfoxide reductase (Msr) enzymes. Canonical MsrA and MsrB were shown to be absolutely stereospecific for the reduction of <i>S</i>-diastereomer and <i>R-</i>diastereomer, respectively. Recently, a new enzymatic system, MsrQ/MsrP which is conserved in all gram-negative bacteria, was identified as a key actor for the reduction of oxidized periplasmic proteins. The haem-binding membrane protein MsrQ transmits reducing power from the electron transport chains to the molybdoenzyme MsrP, which acts as a protein-MetO reductase. The MsrQ/MsrP function was well established genetically, but the identity and biochemical properties of MsrP substrates remain unknown. In this work, using the purified MsrP enzyme from the photosynthetic bacteria <i>Rhodobacter sphaeroides</i> as a model, we show that it can reduce a broad spectrum of protein substrates. The most efficiently reduced MetO is found in clusters, in amino acid sequences devoid of threonine and proline on the C-terminal side. Moreover, <i>R. sphaeroides</i> MsrP lacks stereospecificity as it can reduce both <i>R</i>- and <i>S</i>-diastereomers of MetO, similarly to its <i>Escherichia coli</i> homolog, and preferentially acts on unfolded oxidized proteins. Overall, these results provide important insights into the function of a bacterial envelop protecting system, which should help understand how bacteria cope in harmful environments.