Goodson R J, Katre N V
Cetus Corporation, Emeryville, CA 94608.
Biotechnology (N Y). 1990 Apr;8(4):343-6. doi: 10.1038/nbt0490-343.
We have modified recombinant interleukin-2 (rIL-2) to facilitate site-directed covalent attachment of monomethoxy polyethylene glycol (PEG). The site chosen for modification and subsequent covalent attachment with PEG (PEGylation) was the single glycosylation position found in the native interleukin-2 (IL-2). The mutant protein was expressed in E. coli, purified, and PEGylated with a PEG-maleimide reagent to obtain PEG-cys3-rIL-2. The PEG-cys3-rIL-2 had full bioactivity relative to the unmodified molecule and had an increase in hydrodynamic size sufficient to increase its systemic exposure by approximately 4 fold. This method has general applicability for modifying any therapeutic protein at a specific site and thereby alter its potency. In particular, it can be used to attach PEG to prokaryotically expressed recombinant proteins at their glycosylation sites.
我们对重组白细胞介素-2（rIL-2）进行了改造，以促进单甲氧基聚乙二醇（PEG）的定点共价连接。选择进行改造并随后与PEG共价连接（PEG化）的位点是天然白细胞介素-2（IL-2）中发现的单一糖基化位点。突变蛋白在大肠杆菌中表达、纯化，并用PEG-马来酰亚胺试剂进行PEG化，以获得PEG-cys3-rIL-2。与未修饰的分子相比，PEG-cys3-rIL-2具有完全的生物活性，并且其流体力学尺寸增加，足以使其全身暴露增加约4倍。该方法具有普遍适用性，可在特定位点修饰任何治疗性蛋白质，从而改变其效力。特别是，它可用于在原核表达的重组蛋白的糖基化位点连接PEG。
