里氏木霉内切-1,4-β-木聚糖酶II及其三个二硫键突变体的不可逆热变性：差示扫描量热法表征

Irreversible thermal denaturation of Trichoderma reesei endo-1,4-beta-xylanase II and its three disulfide mutants characterized by differential scanning calorimetry.

作者信息

Jänis Janne, Rouvinen Juha, Vainiotalo Pirjo, Turunen Ossi, Shnyrov Valery L

机构信息

University of Joensuu, Department of Chemistry, PO Box 111, FI-80101 Joensuu, Finland.

出版信息

Int J Biol Macromol. 2008 Jan 1;42(1):75-80. doi: 10.1016/j.ijbiomac.2007.09.012. Epub 2007 Sep 29.

DOI:10.1016/j.ijbiomac.2007.09.012

PMID:17988729

Abstract

Kinetic as well as energetic aspects of the thermal denaturation of Trichoderma reesei endo-1,4-beta-xylanase II (TRX II) and its three thermostable disulfide mutants were characterized by means of differential scanning calorimetry (DSC) in different solution conditions. The calorimetric transitions were strongly scan-rate dependent, characteristic for an irreversible, kinetically controlled protein denaturation. The DSC-determined T*-values (the temperature at which the denaturation rate constant equals 1min(-1)), and the activation free energies for the transitions are consistent with the apparent transition temperatures of TRX II determined earlier by mass spectrometry. Protein aggregation, connected with the irreversibility of the transitions, was present in all cases but was less pronounced with the mutants as well as highly dependent on experimental conditions.

摘要

在不同溶液条件下，通过差示扫描量热法（DSC）对里氏木霉内切 - 1,4 - β - 木聚糖酶II（TRX II）及其三个热稳定二硫键突变体的热变性动力学和能量学方面进行了表征。量热转变强烈依赖扫描速率，这是不可逆的、动力学控制的蛋白质变性的特征。DSC测定的T*值（变性速率常数等于1min⁻¹时的温度）以及转变的活化自由能与早期通过质谱法测定的TRX II的表观转变温度一致。与转变的不可逆性相关的蛋白质聚集在所有情况下均存在，但在突变体中不太明显，并且高度依赖于实验条件。