Department of Pharmaceutical Chemistry, University of California, San Francisco, 1700 Fourth Street, San Francisco, California 94143-2550, United States.
J Med Chem. 2021 Apr 8;64(7):4109-4116. doi: 10.1021/acs.jmedchem.0c02253. Epub 2021 Mar 24.
Small molecule colloidal aggregates adsorb and partially denature proteins, inhibiting them artifactually. Oddly, this inhibition is typically time-dependent. Two mechanisms might explain this: low concentrations of the colloid and enzyme might mean low encounter rates, or colloid-based protein denaturation might impose a kinetic barrier. These two mechanisms should have different concentration dependencies. Perplexingly, when enzyme concentration was increased, incubation times actually lengthened, inconsistent with both models and with classical chemical kinetics of solution species. We therefore considered molecular crowding, where colloids with lower protein surface density demand a shorter incubation time than more crowded colloids. To test this, we grew and shrank colloid surface area. As the surface area shrank, the incubation time lengthened, while as it increased, the converse was true. These observations support a crowding effect on protein binding to colloidal aggregates. Implications for drug delivery and for detecting aggregation-based inhibition will be discussed.
小分子胶态聚集物吸附并部分变性蛋白质，从而人为地抑制它们。奇怪的是，这种抑制通常是时间依赖性的。两种机制可能解释这一点：胶体和酶的低浓度可能意味着低的相遇率，或者基于胶体的蛋白质变性可能施加一个动力学障碍。这两种机制应该有不同的浓度依赖性。令人费解的是，当酶浓度增加时，孵育时间实际上延长了，这与两种模型以及溶液物种的经典化学动力学都不一致。因此，我们考虑了分子拥挤，其中蛋白质表面密度较低的胶体比更拥挤的胶体需要更短的孵育时间。为了验证这一点，我们生长和收缩了胶体的表面积。随着表面积的缩小，孵育时间延长，而当表面积增加时，情况则相反。这些观察结果支持了蛋白质与胶态聚集物结合的拥挤效应。这些结果对药物传递和检测基于聚集的抑制具有重要意义。
