Cloning and Sequencing of a Trimubin cDNA from Trimeresurus mucrosquamatus

• Main Authors: Wu, Hui-Hua, 吳惠華
• Other Authors: Chao, Chung-Fays ; Guo, Yaw-Wen
• Format: Others
• Language: zh-TW
• Published: 1995
• Online Access: http://ndltd.ncl.edu.tw/handle/11721294897029834395

碩士 === 國防醫學院 === 解剖學研究所 === 83 === Trimeresurus mucrosquamatus, a common venomous snake in Taiwan, belongs to the family Viperidae. In previous study we have obtained a partial cDNA clone by immunoscreening from venom gland cDNA library of T. mucrosquamatus. Partial cDNA sequence analysis indicate a strong homology with that of snake venom protein batroxobin. Batroxobin is a thrombin-like enzyme isolated from Bothrops atrox which are also member of the family Viperidae. Batroxobin has a defibrinogenerating effect and is clinically used for the treatment of thrombotic diseases. In this study, a full-length cDNA isolated from T. mucrosquamatus and designated as trimubin. The cDNA sequence and the structure and functional relationships of trimubin cDNA is characterized. The 5''-rapid amplification of cDNA end (RACE) are used in the isolation and construction of the full-length cDNA. Both RACE amplified cDNA and cloned partial cDNA detects a message with a similar size of 1.5 kb. Northern blot analysis by this result suggests that the RACE amplifi ed cDNA and cloned partial cDNA is in the same gene but on the opposite end. The full-length trimubin cDNA consists of 1576 bp with a 5''-noncoding region of 188 bp and a 3''-noncoding segment of 614 bp. Based on the sequence homology comparison and Kozak sequence analysis, the protein translation begins at 189 base and stops at 962 base. The encoding region in between is 771 bases and could code for 257 amino acids with a calculated molecular weight of 28.1 kDa. A typical poly (A) signal, AATAAA is located 15 bases in the upstream of the poly (A) tail. Amino acid sequence comparison shows strong homology with venombin A, ancrod and batroxobin. trimubin is a serine proteinase.