Methionine sulfoxide reductase contributes to meeting dietary methionine requirements
Highlights
► We demonstrate a role in mammalian nutrition for the methionine sulfoxide reductases. ► Weanling mice on methionine restricted diets are growth retarded. ► Knocking out methionine sulfoxide reductase A and B1 increases the growth retardation. ► The reductases likely act by preventing the loss of methionine as its sulfoxide.
Introduction
Virtually all organisms from bacteria to mammals have several methionine sulfoxide reductases (Msr)1 that catalyze the reduction of methionine sulfoxide back to methionine. The oxidation of methionine to its sulfoxide can be effected by reactive oxygen and nitrogen species. Approximately 98% of methionine in organisms is in proteins, and the steady state content of methionine sulfoxide in mouse tissues is 4–10% of the total methionine [1], [2]. Recovery of this oxidized methionine could therefore contribute to meeting the nutritional requirement for methionine (Fig. 1).
The oxidation of methionine produces two stereospecific forms of methionine sulfoxide. The R-form is reduced specifically by MsrB and the S-form by MsrA. In mammals, there is only one form of MsrA while there are three isoforms of MsrB: MsrB1, MsrB2 and MsrB3, each encoded by a different gene. MsrB1 is the most abundant of the B isozymes and is found in the cytosol and the nucleus. There is considerable evidence that MsrA and MsrB provide an antioxidant defense by scavenging reactive oxygen species through cyclic oxidation and reduction of methionine and methionine sulfoxide [3], [4], [5], [6], [7], [8], [9], [10], [11], [12]. The reductase reaction also conserves methionine for metabolic processes by preventing its loss as methionine sulfoxide. Methionine is an essential amino acid in mammals [13], required for protein initiation, incorporation into proteins, and one carbon metabolism. We hypothesized that the Msr contribute to normal nutrition, especially in animals living in the wild with limited food sources. We tested this hypothesis with a classical biochemical nutrition experiment, namely following the growth of weanling mice on diets with decreasing methionine content. We studied the growth of wild-type mice and of mice genetically modified to lack MsrA, MsrB1, or both and also of mice overexpressing MsrA. On a low methionine diet, we expected mice lacking the reductase to exhibit blunted growth and that this growth retardation might be prevented by overexpression of the reductase.
Section snippets
Generation of Msr overexpressing and knockout mice
All mice described in this work were generated on C57BL/6 background. Mice were treated in accordance with the Guide for the Care and Use of Laboratory Animals (NIH publication 85-23, 1996), and the study was approved by the Animal Care and Use Committee of the National Heart, Lung, and Blood Institute. MsrA transgenic mice were generated as described [14]. MsrA residues 21–233 were included in the construct TgCyto_Myr for the cytosolic targeted transgenic mice, and the overexpressed MsrA was
Results
We recently characterized the MsrA transgenic mice used in this study [19], and for this study also generated a KO_B1 (Fig. 2C). Crossing the two knockouts generated the double knockout, KO_AB1 (Figs. 2C and 3). The protein levels of MsrA in liver, kidney, and brain were determined by quantitative immunoblotting (Fig. 3). In the single knockout animals, there was no compensatory change in level of the remaining Msr. As found previously for the MsrA_KO, the MsrB1_KO and the MsrAB1_KO mice were
Discussion
A goal of our laboratory is to define the in vivo functions of the methionine sulfoxide reductases. One approach we adopted was the study of mice lacking MsrA, MsrB1, or both along with transgenic mice overexpressing MsrA in the cytosol or the mitochondria [19]. For the cytosolic MsrA overexpression, we created strains in which the enzyme was myristoylated and non-myristoylated to facilitate study of the effect of myristoylation. All of these genetically altered mice developed normally and
Acknowledgments
We thank the NIH Building 50 animal facility staff for their important support in this study. We also thank the Biochemistry Core facility of the National Heart, Lung, and Blood Institute for access to key instruments. The authors’ responsibilities were as follows: HZ, GK, and RLL designed the research; HZ conducted the research; HZ and RLL analyzed the data; HZ and RLL wrote the manuscript; GK provided essential materials; RLL had primary responsibility for the final content of the manuscript.
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2019, Free Radical Biology and MedicineCitation Excerpt :The 4 methionine sulfoxide reductases are located on 4 different chromosomes in the mouse, facilitating generation of the quadruple knockout. We had previously generated a double knockout, MsrA−/−/MsrB1−/− [40], so that only MsrB2 and MsrB3 knockouts were required to eventually generate the quadruple knockout. CRISPR/Cas9 was used to target the first coding exon of MsrB2−/− and MsrB3−/−.
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2014, Biochimica et Biophysica Acta - Proteins and ProteomicsCitation Excerpt :While mice showed no phenotype under normal dietary conditions, the reduced Met diet caused growth retardation. This was especially pronounced in MsrA−/− MsrB−/− mice, whereas MsrA−/− mice and MsrB1−/− mice grew like wild-type mice [58]. This suggests that Msrs contribute to satisfy the nutritional requirements for Met by reducing Met-SO back to Met, which is supported by the fact that the steady state content of Met-SO in mouse tissues is 4–10% of the total Met [101].