TY - JOUR
T1 - Thioredoxin regulates human mercaptopyruvate sulfurtransferase at physiologically-relevant concentrations
AU - Yadav, Pramod Kumar
AU - Vitvitsky, Victor
AU - Carballal, Sebastián
AU - Seravalli, Javier
AU - Banerjee, Ruma
N1 - Publisher Copyright:
© 2020 Yadav et al. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc.
PY - 2020/5/8
Y1 - 2020/5/8
N2 - 3-Mercaptopyruvate sulfur transferase (MPST) catalyzes the desulfuration of 3-mercaptopyruvate (3-MP) and transfers sulfane sulfur from an enzyme-bound persulfide intermediate to thiophilic acceptors such as thioredoxin and cysteine. Hydrogen sulfide (H2S), a signaling molecule implicated in many physiological processes, can be released from the persulfide product of the MPST reaction. Two splice variants of MPST, differing by 20 amino acids at the N terminus, give rise to the cytosolic MPST1 and mitochondrial MPST2 isoforms. Here, we characterized the poorly-studied MPST1 variant and demonstrated that substitutions in its Ser–His–Asp triad, proposed to serve a general acid–base role, minimally affect catalytic activity. We estimated the 3-MP concentration in murine liver, kidney, and brain tissues, finding that it ranges from 0.4 μmol kg-1 in brain to 1.4 μmol kg-1 in kidney. We also show that N-acetylcysteine, a widely-used antioxidant, is a poor substrate for MPST and is unlikely to function as a thiophilic acceptor. Thioredoxin exhibits substrate inhibition, increasing the KM for 3-MP ~15-fold compared with other sulfur acceptors. Kinetic simulations at physiologically-relevant substrate concentrations predicted that the proportion of sulfur transfer to thioredoxin increases ~3.5-fold as its concentration decreases from 10 to 1 μM, whereas the total MPST reaction rate increases ~7-fold. The simulations also predicted that cysteine is a quantitatively-significant sulfane sulfur acceptor, revealing MPST’s potential to generate low-molecular-weight persulfides. We conclude that the MPST1 and MPST2 isoforms are kinetically indistinguishable and that thioredoxin modulates the MPST-catalyzed reaction in a physiologically-relevant concentration range.
AB - 3-Mercaptopyruvate sulfur transferase (MPST) catalyzes the desulfuration of 3-mercaptopyruvate (3-MP) and transfers sulfane sulfur from an enzyme-bound persulfide intermediate to thiophilic acceptors such as thioredoxin and cysteine. Hydrogen sulfide (H2S), a signaling molecule implicated in many physiological processes, can be released from the persulfide product of the MPST reaction. Two splice variants of MPST, differing by 20 amino acids at the N terminus, give rise to the cytosolic MPST1 and mitochondrial MPST2 isoforms. Here, we characterized the poorly-studied MPST1 variant and demonstrated that substitutions in its Ser–His–Asp triad, proposed to serve a general acid–base role, minimally affect catalytic activity. We estimated the 3-MP concentration in murine liver, kidney, and brain tissues, finding that it ranges from 0.4 μmol kg-1 in brain to 1.4 μmol kg-1 in kidney. We also show that N-acetylcysteine, a widely-used antioxidant, is a poor substrate for MPST and is unlikely to function as a thiophilic acceptor. Thioredoxin exhibits substrate inhibition, increasing the KM for 3-MP ~15-fold compared with other sulfur acceptors. Kinetic simulations at physiologically-relevant substrate concentrations predicted that the proportion of sulfur transfer to thioredoxin increases ~3.5-fold as its concentration decreases from 10 to 1 μM, whereas the total MPST reaction rate increases ~7-fold. The simulations also predicted that cysteine is a quantitatively-significant sulfane sulfur acceptor, revealing MPST’s potential to generate low-molecular-weight persulfides. We conclude that the MPST1 and MPST2 isoforms are kinetically indistinguishable and that thioredoxin modulates the MPST-catalyzed reaction in a physiologically-relevant concentration range.
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U2 - 10.1074/jbc.RA120.012616
DO - 10.1074/jbc.RA120.012616
M3 - Article
C2 - 32179647
AN - SCOPUS:85084721251
SN - 0021-9258
VL - 295
SP - 6299
EP - 6310
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 19
ER -