Functional Expression and Purification of the Recombinant Pineapple Stem Lectin with GST Fusion from E. coli

• Main Authors: Ciou-Wun Liao, 廖秋雯
• Other Authors: Sorgan S. K. Tai
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/e2t7f6

碩士 === 國立虎尾科技大學 === 生物科技研究所 === 101 === Lectins are carbohydrate-binding proteins that can agglutinate cells and precipitate glycoconjugates. Plants contain the lectins that bind preferentially to mannose or glucose occurred in many monocot or dicot species. The lectins are important tool for the research of carbohydrates and to identify their function in glycobiology. One of the mainly protein in pineapple stem has been identified as a lectin. The lectin denoted as ACL (Ananas Comosus Lectin) has been isolated, purified and cloned from the pineapple stem. The coding region of ACL clone was introduction into expression vector, pGEX-4T-1 with GST-tag fusion, named pGEX-lec and transformed to Escherichia coli BL21 (DE3). Overexpression of the recombinant ACL (rACL) in the lower culture temperature the water-soluble protein of rACL was found. The proteins subjected to a glutathione affinity column to purify the GST-fusion rACL. Western analysis of rACL confirmed that rACL is proper expressed. In this study, rACL exhibited a similar hemagglutation activity to purified native ACL (nACL) that could agglutinate erythrocytes. Functional expression of rACL and establishing the produce system is an important step forward for future study of the ACL.