Feng Jianwen, Stieglitz Kimberly, Roberts Mary F
Merkert Chemistry Center, Boston College, Chestnut Hill, Massachusetts 02467, USA.
J Am Chem Soc. 2004 Feb 4;126(4):1008-9. doi: 10.1021/ja038529u.
Two mutations, R69D and K115E, converted a bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) to a phosphatase with much higher specific activity toward glucose-6-phosphate than inositol-1-phosphate. PI-PLC single mutations R69D and K115E can cleave PI but lack any demonstrable phosphatase activity. The bacterial PI-PLC has no sequence homology with known glucose-6-phosphatase enzymes, which need His, Arg, and negatively charged residues (Asp or Glu) at the active site. The change in chemical reaction and substrate specificity can be rationalized by energy minimization of the mutant with I-1-P or G-6-P bound.
R69D和K115E这两个突变将一种细菌磷脂酰肌醇特异性磷脂酶C（PI-PLC）转变为一种磷酸酶，该磷酸酶对葡萄糖-6-磷酸的比活性远高于对肌醇-1-磷酸的比活性。PI-PLC的单突变R69D和K115E能够切割PI，但缺乏任何可证明的磷酸酶活性。这种细菌PI-PLC与已知的葡萄糖-6-磷酸酶没有序列同源性，后者在活性位点需要组氨酸、精氨酸和带负电荷的残基（天冬氨酸或谷氨酸）。通过结合I-1-P或G-6-P的突变体的能量最小化，可以解释化学反应和底物特异性的变化。
