IDENTIFICATION OF PALMITOYLATION SITES WITHIN THE L-TYPE CALCIUM-CHANNEL BETA(2A) SUBUNIT AND EFFECTS ON CHANNEL FUNCTION

The hydrophilic beta(2a) subunit of the L-type calcium channel was rec ently shown to be a membrane-localized, post-translationally modified protein (Chien, A. J., Zhao, X. L., Shirokov, R. E., Purl, T. S., Chan g, C. F., Sun, D. D., Rios, E., and Hosey, M. M. (1995) J. Biol, Chem, 270, 30036-30044), In this study, we demonstrate that the rat beta(2a ) subunit was palmitoylated through a hydroxylamine sensitive thioeste r linkage, Palmitoylation required a pair of cysteines in the N termin us, Cys(3) and Cys(4); mutation of these residues to serines resulted in mutant beta(2a) subunits that were unable to incorporate palmitic a cid, Interestingly, a palmitoylation-deficient beta(2a) mutant still l ocalized to membrane particulate fractions and was still able to targe t functional channel complexes to the plasma membrane similar to wild- type beta(2a). However, channels formed with a palmitoylation-deficien t beta(2a) subunit exhibited a dramatic decrease in ionic current per channel, indicating that although mutations eliminating palmitoylation did not affect channel targeting by the beta(2a) subunit, they were i mportant determinants of channel modulation by the beta(2a) subunit, T hree other known beta subunits that were analyzed were not palmitoylat ed, suggesting that palmitoylation could provide a basis for the regul ation of L-type channels through modification of a specific beta isofo rm.