Liu Hongcheng, Gaza-Bulseco Georgeen, Sun Joanne
Protein Analytics, Process Sciences Department, 100 research Drive, Abbott Bioresearch Center, Worcester, MA 01605, USA.
J Chromatogr B Analyt Technol Biomed Life Sci. 2006 Jun 6;837(1-2):35-43. doi: 10.1016/j.jchromb.2006.03.053. Epub 2006 Apr 27.
The susceptible degradation sites of therapeutic proteins are routinely assessed under accelerated conditions such as exposure to chemicals or incubation at elevated temperature or a combination of both. A fully human monoclonal IgG(1) antibody was characterized after incubation at 40 degrees C for 6 months by employing mass spectrometry and chromatography analyses. It was found that deamidation, fragmentation and N-terminal glutamate cyclization to form pyroglutamate are the major degradation pathways. Three major deamidation sites were identified and one site in a small tryptic peptide accounted for more than 80% of the total. Peptide cleavage was observed at several positions between different pairs of amino acids. Most of the cleavage sites were located in the hinge or other flexible regions of the IgG molecule.
治疗性蛋白质的敏感降解位点通常在加速条件下进行评估，如暴露于化学物质、在高温下孵育或两者结合。通过质谱和色谱分析，对一种全人源单克隆IgG(1)抗体在40℃孵育6个月后的情况进行了表征。结果发现，脱酰胺、片段化和N端谷氨酸环化形成焦谷氨酸是主要的降解途径。确定了三个主要的脱酰胺位点，一个位于小胰蛋白酶肽段中的位点占总量的80%以上。在不同氨基酸对之间的几个位置观察到了肽段裂解。大多数裂解位点位于IgG分子的铰链区或其他柔性区域。
