Ostermann N, Lavie A, Padiyar S, Brundiers R, Veit T, Reinstein J, Goody R S, Konrad M, Schlichting I
Department of Physical Biochemistry, Max Planck Institute for Molecular Physiology, Otto-Hahn-Str. 11 D-44227 Dortmund, Germany.
J Mol Biol. 2000 Nov 17;304(1):43-53. doi: 10.1006/jmbi.2000.4175.
The 60-fold reduced phosphorylation rate of azidothymidine (AZT) monophosphate (AZTMP), the partially activated AZT metabolite, by human thymidylate kinase (TMPK) severely limits the efficacy of this anti-HIV prodrug. Crystal structures of different TMPK nucleotide complexes indicate that steric hindrance by the azido group of AZTMP prevents formation of the catalytically active closed conformation of the P-loop of TMPK. The F105Y mutant and a chimeric mutant that contains sequences of the human and Escherichia coli enzyme phosphorylate AZTMP 20-fold faster than the wild-type enzyme. The structural basis of the increased activity is assigned to stabilization of the closed P-loop conformation.
叠氮胸苷（AZT）单磷酸（AZTMP）是部分活化的AZT代谢产物，其被人胸苷酸激酶（TMPK）磷酸化的速率降低了60倍，这严重限制了这种抗HIV前药的疗效。不同TMPK核苷酸复合物的晶体结构表明，AZTMP叠氮基团造成的空间位阻会阻止TMPK P环形成具有催化活性的闭合构象。F105Y突变体以及包含人和大肠杆菌酶序列的嵌合突变体对AZTMP的磷酸化速度比野生型酶快20倍。活性增加的结构基础归因于闭合P环构象的稳定。
