Faculty of Science, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
Appl Biochem Biotechnol. 2012 Jun;167(3):612-20. doi: 10.1007/s12010-012-9728-2. Epub 2012 May 12.
In silico and experimental investigations were conducted to explore the effects of substituting hydrophobic residues, Val, Met, Leu, Ile, Trp, and Phe into Gln 114 of T1 lipase. The in silico investigations accurately predicted the enzymatic characteristics of the mutants in the experimental studies and provided rationalization for some of the experimental observations. Substitution with Leu successfully improved the conformational stability and enzymatic characteristics of T1 lipase. However, replacement of Gln114 with Trp negatively affected T1 lipase and resulted in the largest disruption of protein stability, diminished lipase activity and inferior enzymatic characteristics. These results suggested that the substitution of a larger residue in a densely packed area of the protein core can have considerable effects on the structure and function of an enzyme. This is especially true when the residue is next to the catalytic serine as demonstrated with the Phe and Trp mutation.
进行了计算机模拟和实验研究，以探讨将疏水残基 Val、Met、Leu、Ile、Trp 和 Phe 替代 T1 脂肪酶中的 Gln114 的效果。计算机模拟准确预测了实验研究中突变体的酶学特性，并为一些实验观察提供了合理化解释。用 Leu 取代成功地提高了 T1 脂肪酶的构象稳定性和酶学特性。然而，用 Trp 取代 Gln114 对 T1 脂肪酶产生了负面影响，导致蛋白质稳定性的最大破坏、脂肪酶活性的降低和较差的酶学特性。这些结果表明，在蛋白质核心密集堆积区域中替换较大的残基会对酶的结构和功能产生相当大的影响。当残基靠近催化丝氨酸时，这种情况尤其如此，如 Phe 和 Trp 突变所证明的那样。
