Synthesis of a membrane protein with two transmembrane regions

✍ Scribed by Takeshi Sato; Toru Kawakami; Kenichi Akaji; Hiroki Konishi; Koji Mochizuki; Toshimichi Fujiwara; Hideo Akutsu; Saburo Aimoto

• Book ID: 105360192
• Publisher: John Wiley and Sons
• Year: 2002
• Tongue: English
• Weight: 135 KB
• Volume: 8
• Category: Article
• ISSN: 1075-2617
• DOI: 10.1002/psc.381

✦ Synopsis

Abstract

A membrane protein with two transmembrane domains was synthesized by means of the thioester method. The F~1~F~0~ ATP synthase subunit c (Sub.c), which consists of 79 amino acid residues (MW 8257), was chosen as a target. For synthetic purposes, two building blocks, Boc‐[Lys^34^(Boc)]‐Sub.c(1‐38)‐SCH~2~CH~2~CO‐Ala and Sub.c(39‐79), were synthesized via solid‐phase methods using Boc chemistry. RP‐HPLC purification conditions for the transmembrane peptide were examined. As a result, a combination of a mixture of formic acid, 1‐propanol and water with a phenyl column was found to be useful for separating the transmembrane peptide. The purified building blocks were condensed in DMSO in the presence of silver chloride, 3,4‐dihydro‐3‐hydroxy‐4‐oxo‐1,2,3‐benzotriazine (HOOBt), N,N‐diisopropylethylamine to give the product, Sub.c, after removal of Boc groups (yield 16%). The yield of the condensation reaction could be improved to 23% by raising the reaction temperature to 50 °C, and to 26% when a mixture of chloroform and methanol was used as a solvent. Copyright © 2002 European Peptide Society and John Wiley & Sons, Ltd.