Synthesis of the C-terminal region of opioid receptor like 1 in an SDS micelle by the native chemical ligation:: effect of thiol additive and SDS concentration on ligation efficiency

In the process of developing a method for the synthesis of membrane proteins, the conditions for native chemical ligation, namely, detergent concentration and the chemical characteristics of the thiol additive were investigated in detail. The C-terrninal region of the opioid receptor like 1, ORLI(288-370), which contains C-terminal intracellular and transmembrane domains, was chosen as a model. The building blocks, ORL1(329-370) and ORL1(288-328)-SR-GIy-Arg5-Lcu (-SR- : -SCH2CH2CO-) were most effectively condensed slightly below the critical micelle concentration of SIDS and in the presence of mercaptoethanesulfonic acid as a thiol additive. The results showed that the concentration of SDS and the charge on the thiol additive are crucial factors for the effective synthesis of a membrane protein by native chemical ligation. Copyright (c) 2004 European Peptide Society and John Wiley & Sons, Ltd.