Novel site-specific immobilization of a functional protein using a preferred substrate sequence for transglutaminase 2
- 作者：Yoshiaki Sugimura ; Hiroshi Ueda ; Masatoshi Maki ; Kiyotaka Hitomi
- 关键词：Transglutaminase ; Glutathione-S-transferase ; Single chain fragment antibody ; Cross-linking ; TRANSIM
- 刊名：Journal of Biotechnology
- 出版年：2007
- 期刊代码：53_01681656
- 类别：imm
- 出版时间：31 August 2007
- 卷：131
- 期：2
- 页码：121-127
- 文件大小：479 K
摘要
Transglutaminase (TGase) catalyzes the formation of a covalent cross-link between a peptide-bound glutamine residue and a lysine residue or primary amine. We have recently identified specific preferred sequences as glutamine-donor substrates in TGase 2 and Factor XIII reactions. By taking advantage of preference of the 12-amino acid sequence for the enzymatic reaction, an efficient immobilization method was established using two different model proteins, glutathione S-transferase (GST) and single-chain fragment antibody (scFv). Both proteins were genetically attached with the preferred substrate sequence to produce a fusion protein. Attachment of the sequence enables the recombinant proteins to act as prominent TGase-substrates and enables them to be immobilized onto chemically amine-terminated gels. Investigation of the biological activities of the two proteins demonstrated their effective immobilization in comparison with that by using a chemically immobilizing method. This established system, which we designated as Transglutaminase-mediated site-specific immobilization method (TRANSIM), would provide site-specific and biologically active conjugation between proteins and several non-protein materials.