高压处理对木瓜蛋白酶活性的影响及分子动力学模拟

为研究超高压处理对木瓜蛋白酶活性的影响及分子机制，将木瓜蛋白酶溶液经0.1、200和600 MPa处理，测定其活性变化，并对木瓜蛋白酶在这3种压力下进行150 ns的分子模拟，分析RMSD（均方根误差）、RMSF（均方根涨落）、回旋半径、氢键、溶剂可及表面积、体积、二级结构和分子表面结构等的变化。结果显示，200 MPa对木瓜蛋白酶起到激活作用，而600 MPa条件下酶活性降低26.2%。分子模拟表明高压处理能减小酶蛋白结构的波动；减少蛋白体积，使蛋白结构更加致密；蛋白溶剂可及表面积降低，特别是疏水表面积；高压能破坏酶蛋白间的氢键，使蛋白中β-折叠增加，而α-螺旋则是在600 MPa明显减少。200 MPa下酶的激活可能与该压力下酶活性中心的结合口袋变大因而更易与目标蛋白质结合有关；而600 MPa下酶活性的部分抑制则与该压力下酶结构遭受较大破坏有关，如α-螺旋和氢键的破坏、疏水表面积和蛋白体积的减小。从分子机制上明确了高压下木瓜蛋白酶活性变化的机理。

Effect of High Pressure Treatment on Papain Activity and Molecular Dynamics Simulation

To study the effect of ultrahigh pressure treatment on papain activity and the molecular mechanism, papain solutions were treated with 0.1, 200 and 600 MPa to determine the changes in activity. And molecular simulation of papain at these three pressures were performed for 150 ns, the changes of root mean square deviation (RMSD), root mean square fluctuation (RMSF), radius of gyration (Rg), hydrogen bonds, solvent accessible surface area, volume, secondary structure and molecular surface structure was evaluated. The results showed that 200 MPa had an activating effect on papain, while the enzyme activity was reduced by 26.2% at 600 MPa. Molecular simulation showed that high pressure treatment could reduce the fluctuation of protein structure, protein volume, and protein solvent accessible surface area, especially hydrophobic surface area, and make the protein structure more compact. High pressure could destroy the hydrogen bond between enzyme proteins and increase the content of β-sheet, while decrease the α-helix obviously at 600 MPa. The activation of the enzyme at 200 MPa may be related to the larger binding pocket in the active center of the enzyme at this pressure, which made it easier to bind to the target protein. The partial inhibition of enzyme activity under 600 MPa was related to the destruction of enzyme structure under this pressure, for example, the destruction of α-helix and hydrogen, the reduction of hydrophobic surface area and protein volume. The molecular mechanism of papain activity change under high pressure was clarified.