CiteULike: neils's Feng

Structural Basis of Substrate Recognition in Thiopurine S-Methyltransferase

Yi Peng — 2008-05-19T01:44:46-00:00

Biochemistry (17 May 2008)

Abstract: Thiopurine S-methyltransferase (TPMT) modulates the cytotoxic effects of thiopurine prodrugs such as 6-mercaptopurine by methylating them in a reaction using S-adenosyl-L-methionine as the donor. Patients with TPMT variant allozymes exhibit diminished levels of protein and/or enzyme activity and are at risk for thiopurine drug-induced toxicity. We have determined two crystal structures of murine TPMT, as a binary complex with the product S-adenosyl-L-homocysteine and as a ternary complex with S-adenosyl-L-homocysteine and the substrate 6-mercaptopurine, to 1.8 and 2.0 Å resolution, respectively. Comparison of the structures reveals that an active site loop becomes ordered upon 6-mercaptopurine binding. The positions of the two ligands are consistent with the expected SN2 reaction mechanism. Arg147 and Arg221, the only polar amino acids near 6-mercaptopurine, are highlighted as possible participants in substrate deprotonation. To probe whether these residues are important for catalysis, point mutants were prepared in the human enzyme. Substitution of Arg152 (Arg147 in murine TPMT) with glutamic acid decreases Vmax and increases Km for 6-mercaptopurine but not Km for S-adenosyl-L-methionine. Substitution at this position with alanine or histidine and similar substitutions of Arg226 (Arg221 in murine TPMT) result in no effect on enzyme activity. The double mutant Arg152Ala/Arg226Ala exhibits a decreased Vmax and increased Km for 6-mercaptopurine. These observations suggest that either Arg152 or Arg226 may participate in some fashion in the TPMT reaction, with one residue compensating when the other is altered, and that Arg152 may interact with substrate more directly than Arg226, consistent with observations in the murine TPMT crystal structure.

The draft genome of the transgenic tropical fruit tree papaya (Carica papaya Linnaeus)

Ray Ming — 2008-04-23T19:37:29-00:00

Nature, Vol. 452, No. 7190. (24 April 2008), pp. 991-996.

PPSP: prediction of PK-specific phosphorylation site with Bayesian decision theory.

Yu Xue — 2007-12-04T03:22:11-00:00

BMC Bioinformatics, Vol. 7 (2006)

BACKGROUND: As a reversible and dynamic post-translational modification (PTM) of proteins, phosphorylation plays essential regulatory roles in a broad spectrum of the biological processes. Although many studies have been contributed on the molecular mechanism of phosphorylation dynamics, the intrinsic feature of substrates specificity is still elusive and remains to be delineated. RESULTS: In this work, we present a novel, versatile and comprehensive program, PPSP (Prediction of PK-specific Phosphorylation site), deployed with approach of Bayesian decision theory (BDT). PPSP could predict the potential phosphorylation sites accurately for approximately 70 PK (Protein Kinase) groups. Compared with four existing tools Scansite, NetPhosK, KinasePhos and GPS, PPSP is more accurate and powerful than these tools. Moreover, PPSP also provides the prediction for many novel PKs, say, TRK, mTOR, SyK and MET/RON, etc. The accuracy of these novel PKs are also satisfying. CONCLUSION: Taken together, we propose that PPSP could be a potentially powerful tool for the experimentalists who are focusing on phosphorylation substrates with their PK-specific sites identification. Moreover, the BDT strategy could also be a ubiquitous approach for PTMs, such as sumoylation and ubiquitination, etc.