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Phosphorylation of methylated-DNA-protein-cysteine S-methyltransferase at serine-204 significantly increases its resistance to proteolytic digestion.

Lim, IK; Park, TJ; Paik, WK
The Biochemical journal, 352(Pt3):801-808, 2000
Journal Title
The Biochemical journal
In a previous paper [Lim, Park, Jee, Lee and Paik (1999) J. Cancer Res. Clin. Oncol. 125, 493-499], we showed two major forms of active DNA-6-O-methylguanine:protein-L-cysteine S-methyltransferase (MGMT; EC in the liver with N-nitrosodiethylamine (DEN)-induced carcinogenesis: these were 26 and 24 kDa species. Here we show that a 2 kDa C-terminal fragment was cleaved from the 26 kDa species in vitro by thrombin or microsomal fractions isolated from DEN-treated rat livers. When Ser(204) of the 26 kDa protein was replaced with Ala by site-directed mutagenesis, phosphorylation of the protein was completely abolished, indicating Ser(204) to be the site of phosphorylation. We also show that the phosphorylation was performed by Ca(2+)-independent protein kinase isoenzymes, and that the phosphorylated rat MGMT protein was resistant to digestion by protease(s) whose activity was increased during DEN-induced hepatocarcinogenesis and also by digestion with endopeptidase Glu-C (V8 protease).
MeSH terms
Amino Acid SequenceAmino Acid Substitution/geneticsAnimalsBlotting, WesternCalcium/physiologyCarcinogens/pharmacologyDiethylnitrosamine/pharmacologyEndopeptidases/*metabolismEnzyme Induction/drug effectsHumansIsoenzymes/metabolismLiver Neoplasms/chemically induced/enzymology/metabolismMaleMicrosomes, Liver/drug effects/enzymology/metabolismMolecular Sequence DataMolecular WeightO(6)-Methylguanine-DNA Methyltransferase/*chemistry/genetics/*metabolismPhosphorylation/drug effectsPhosphoserine/*metabolismProtein Kinase C/antagonists & inhibitors/metabolismRatsRats, Sprague-DawleySequence AlignmentSerine Endopeptidases/metabolismThrombin/metabolism
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Journal Papers > School of Medicine / Graduate School of Medicine > Biochemistry & Molecular Biology
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임, 인경박, 태준백, 운기
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