Baptista R P, Chen L Y, Paixão A, Cabral J M S, Melo E P
Centro de Engenharia Biológica e Química, Instituto Superior Técnico, Av. Rovisco Pais, Lisboa, Portugal.
Biotechnol Bioeng. 2003 Jun 30;82(7):851-7. doi: 10.1002/bit.10641.
Cutinase in aqueous solution at pH 4.5 deactivates following a parallel pathway. At 53 degrees C, 88% of the cutinase molecules are in the unfolded conformation, which can aggregate with a reaction order of 3 if the protein concentration is high (>/=12 microM). The aggregates show a sixfold increase in size as determined by dynamic light scattering. This aggregation process is the first phase observed during a deactivation experiment; however, after significant cutinase depletion and maturation of the aggregates, a first-order step starts to dominate and a second phase independent of the protein concentration is observed. Kinetic partitioning between aggregation and first-order irreversible changes of the unfolded conformation can occur during enzyme deactivation when the equilibrium between the native and the unfolded conformation is shifted and kept toward the unfolded conformation.
在pH 4.5的水溶液中，角质酶通过平行途径失活。在53℃时，88%的角质酶分子处于未折叠构象，如果蛋白质浓度较高（≥12μM），它们会以三级反应顺序聚集。通过动态光散射测定，聚集体的尺寸增大了六倍。这种聚集过程是失活实验中观察到的第一阶段；然而，在角质酶大量消耗且聚集体成熟后，一级步骤开始占主导，观察到一个与蛋白质浓度无关的第二阶段。当天然构象和未折叠构象之间的平衡发生移动并保持向未折叠构象时，在酶失活过程中会发生聚集和未折叠构象的一级不可逆变化之间的动力学分配。
