Kendrick B S, Cleland J L, Lam X, Nguyen T, Randolph T W, Manning M C, Carpenter J F
Department of Pharmaceutical Sciences, University of Colorado Health Sciences Center, Campus Box C238, Denver, Colorado 80262, USA.
J Pharm Sci. 1998 Sep;87(9):1069-76. doi: 10.1021/js9801384.
Protein aggregation is a complex phenomenon that can occur in vitro and in vivo, usually resulting in the loss of the protein's biological activity. While many aggregation studies focus on a mechanism due to a specific stress, this study focuses on the general nature of aggregation. Recombinant human interferon-gamma (rhIFN-gamma) provides an ideal model for studying protein aggregation, as it has a tendency to aggregate under mild denaturing stresses (low denaturant concentration, temperature below the Tm, and below pH 5). All of the aggregates induced by these stresses have a similar structure (high in intermolecular beta-sheet content and a large loss of alpha-helix) as determined by infrared and circular dichroism spectroscopy. Thermally induced and denaturant-induced aggregation processes follow first-order kinetics under the conditions of this study. Spectroscopic and kinetic data suggest that rhIFN-gamma aggregates through an intermediate form possessing a large amount of residual secondary structure. In contrast to the aggregates formed under denaturing stresses, the salted-out protein has a remarkably nativelike secondary structure.
蛋白质聚集是一种复杂的现象，可在体外和体内发生，通常会导致蛋白质生物活性丧失。虽然许多聚集研究聚焦于特定应激导致的机制，但本研究关注的是聚集的一般性质。重组人干扰素-γ（rhIFN-γ）为研究蛋白质聚集提供了一个理想模型，因为它在温和变性应激（低变性剂浓度、低于熔点温度以及pH值低于5）下有聚集倾向。通过红外光谱和圆二色光谱测定，这些应激诱导产生的所有聚集体都具有相似的结构（分子间β-折叠含量高且α-螺旋大量丧失）。在本研究条件下，热诱导和变性剂诱导的聚集过程遵循一级动力学。光谱和动力学数据表明，rhIFN-γ通过具有大量残余二级结构的中间形式聚集。与在变性应激下形成的聚集体不同，盐析蛋白具有显著的类似天然的二级结构。
