Covalent Immobilisation of Transglutaminase: Stability and Applications in Protein PEGylation
Journal of drug targeting(2017)
摘要
Microbial transglutaminase enzyme (mTGase) is an extremely useful enzyme that is increasingly employed in the food and pharmaceutical industries and as a tool for protein modification and tagging. The current study describes how we immobilised mTGase (iTGase) on a solid support to improve its stability during the PEGylation process by which polyethylene glycol chains are attached to protein and peptide drugs. When the enzyme was immobilised at the N-terminal sequence on agarose beads, it retained more than 53% of its starting activity. Kinetic studies on the immobilised and free mTGase disclosed a 1.7 and 1.5 fold decrease of Km and Vmax, respectively. Protein PEGylation was carried out using α-lactalbumin (α-LA) and granulocyte colony stimulating factor (G-CSF). In the former case, the iTGase showed a selective conjugation towards only one Gln residue of α-LA, avoiding formation of a mono- and bi-conjugate mixture that is achieved using the free enzyme. In the latter case, the immobilised enzyme still remained selective towards only one Gln, but avoided the undesired formation of deamidated G-CSF that took place when free mTGase was used. Overall, the results of the current study highlight the suitability of iTGase in preparing site-selective protein-polymer conjugates.
更多查看译文
关键词
Transglutaminase,PEGylation,enzyme immobilisation,protein modification,polymer conjugation
AI 理解论文
溯源树
样例
生成溯源树,研究论文发展脉络
Chat Paper
正在生成论文摘要