HUMAN-XENOPUS CHIMERAS OF G(S)ALPHA REVEAL A NEW REGION IMPORTANT FOR ITS ACTIVATION OF ADENYLYL-CYCLASE

G proteins are heterotrimeric GTPases that play a key role in signal transduction. The alpha subunit of G(s) bound to GTP is capable of activating adenylyl cyclase. The amino acid sequences derived from two X. laevis cDNA clones that apparently code for G(s) alpha subunits are 92% identical to those found in the short form of human G(s) alpha. Despite this high homology, the X. laevis G(s) alpha clones expressed in vitro, yielded a protein that are not able to activate the adenylyl cyclase present in S49 cy $$($) over bar c membranes in contrast with human G(s) alpha similarly expressed. This finding suggested that the few amino acid substitutions found in the amphibian subunit are important in defining the functionality of the human G(s) alpha. The construction of chimeras composed of different fractions of the cDNAs of the two species was adopted as an approach in determining the regions of the molecule important in its functionality in this assay. Four pairs of chimeras were constructed using reciprocal combinations of the cDNAs coding for human and Xenopus G(s) alpha. These eight constructs were expressed in vitro and equivalent amounts of the resulting proteins were assayed in the activation of adenylyl cyclase with GTP gamma s and isoproterenol. The results obtained here clearly indicate that the G alpha sequence that extends from amino acid 70 to 140, is important for the functionality of human G(s) alpha in activating adenylyl cyclase.