Synthesis of a membrane protein with two transmembrane regions

A membrane protein with two transmembrane domains was synthesized by means of the thioester method. The F1F0 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)-SCH2CH2Co-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-dihydro3-hydro-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 50degreesC, and to 26% when a mixture of chloroform and methanol was used as a solvent. Copyright (C) 2002 European Peptide Society and John Wiley Sons, Ltd.