TY - JOUR
T1 - Redox regulation of cytosolic glycerol-3-phosphate dehydrogenase
T2 - Cys102 is the target of the redox control and essential for the catalytic activity
AU - Kim, Ji Young
AU - Park, Hee Sae
AU - Kang, Soo Im
AU - Choi, Eui Ju
AU - Kim, Ick Young
N1 - Funding Information:
This work was supported by Grant No. KOSEF 2000-2-20900-008-5 from the Korea Science and Engineering Foundation to I.Y.K.
PY - 2002/1/15
Y1 - 2002/1/15
N2 - Cytosolic glycerol-3-phosphate dehydrogenase (cG3PDH) occupies the branch point between the glycolytic pathway and triglyceride biosynthesis. However, the regulatory mechanism of the cG3PDH activity has remained obscure. Here we report that cG3PDH is efficiently inhibited by modification of the thiol group through a redox mechanism. In this study, we found that sodium selenite and nitric oxide (NO) donors such as S-nitroso-N-acetylpenicillamine and 3-morpholinosydnonimine inhibited cG3PDH activity, and that similar effects could be achieved with selenium metabolites such as selenocysteine and selenomethionine. Furthermore, we found that reducing agents, such as dithiothreitol and β-mercaptoethanol, restored the cG3PDH activity suppressed by selenite and NO both in vitro and in cultured cells. Buthionine sulfoximine depleted levels of both reduced glutathione and the oxidized form but had no effect on the suppression of cG3PDH activity by selenite in cultured cells. Moreover, thiol-reactive agents, such as N-ethylmaleimide and o-iodosobenzoic acid, blocked the enzyme activity of cG3PDH through the modification of redox-sensitive cysteine residues in cG3PDH. The inhibitor of NO synthase, L-NG-nitro-arginine, restored the cG3PDH activity inhibited by NO in cultured cells, whereas the inhibitor of guanylyl cyclase, 1H-[1,2,4] oxadiazole[4,3-α] quinoxalin-1-one (ODQ), has no effect. NO directly inhibits cG3PDH activity not via a cGMP-dependent mechanism. Finally, using site-directed mutagenesis, we found that Cys102 of cG3PDH was sensitive to both selenite and NO. From the results, we suggest that cG3PDH is a target of cellular redox regulation.
AB - Cytosolic glycerol-3-phosphate dehydrogenase (cG3PDH) occupies the branch point between the glycolytic pathway and triglyceride biosynthesis. However, the regulatory mechanism of the cG3PDH activity has remained obscure. Here we report that cG3PDH is efficiently inhibited by modification of the thiol group through a redox mechanism. In this study, we found that sodium selenite and nitric oxide (NO) donors such as S-nitroso-N-acetylpenicillamine and 3-morpholinosydnonimine inhibited cG3PDH activity, and that similar effects could be achieved with selenium metabolites such as selenocysteine and selenomethionine. Furthermore, we found that reducing agents, such as dithiothreitol and β-mercaptoethanol, restored the cG3PDH activity suppressed by selenite and NO both in vitro and in cultured cells. Buthionine sulfoximine depleted levels of both reduced glutathione and the oxidized form but had no effect on the suppression of cG3PDH activity by selenite in cultured cells. Moreover, thiol-reactive agents, such as N-ethylmaleimide and o-iodosobenzoic acid, blocked the enzyme activity of cG3PDH through the modification of redox-sensitive cysteine residues in cG3PDH. The inhibitor of NO synthase, L-NG-nitro-arginine, restored the cG3PDH activity inhibited by NO in cultured cells, whereas the inhibitor of guanylyl cyclase, 1H-[1,2,4] oxadiazole[4,3-α] quinoxalin-1-one (ODQ), has no effect. NO directly inhibits cG3PDH activity not via a cGMP-dependent mechanism. Finally, using site-directed mutagenesis, we found that Cys102 of cG3PDH was sensitive to both selenite and NO. From the results, we suggest that cG3PDH is a target of cellular redox regulation.
KW - Cytosolic glycerol-3-phosphate dehydrogenase
KW - Redox mechanism
KW - Reducing agent
KW - S-Nitroso-N-acetylpenicillamine
KW - Sodium selenite
KW - Thiol
UR - http://www.scopus.com/inward/record.url?scp=0037081758&partnerID=8YFLogxK
U2 - 10.1016/S0304-4165(01)00236-7
DO - 10.1016/S0304-4165(01)00236-7
M3 - Article
C2 - 11853959
AN - SCOPUS:0037081758
SN - 0304-4165
VL - 1569
SP - 67
EP - 74
JO - Biochimica et Biophysica Acta - General Subjects
JF - Biochimica et Biophysica Acta - General Subjects
IS - 1-3
ER -