Probing membrane protein unfolding ... | Article | H1 Connect

This article describes a simple but effective method to determine the stability of membrane proteins that has a number of advantages over existing methods. The method is based on simple pulse proteolysis and gel electrophoresis experiments. It extends a previous work by Park's group, which has used this method to determine the stability and ligand binding of soluble proteins to study membrane proteins. A well-characterised membrane protein, bacteriorhodopsin, is used as a test case, and the stability of the protein towards denaturation by sodium dodecyl sulfate (SDS) using the pulse-proteolysis technique compares favourably with previous studies using optical spectroscopy. The technique has several advantages over conventional methods -- first, no optical signal is needed, making it widely applicable to all membrane proteins; second, only small quantities of protein are needed thus making it possible to study endogenous or transiently expressed membrane proteins; and third, through use of Western blots, the technique can be used on unpurified proteins. Not only will this technique potentially be of much use to those groups interested in the study of the folding of membrane proteins but it can also be used to investigate the effects of disease-related mutations on membrane protein stability, and may be used to screen potential stabilising mutations in order to aid structural studies on membrane proteins. In addition, this technique may also be used to study ligand binding.