Human-Xenopus chimeras of Gsα reveal a new region important for its activation of adenylyl cyclase

AbstractG proteins are heterotrimeric GTPases that play a key role in signal transduction. The α subunit of Gs bound to GTP is capable of activating adenylyl cyclase. The amino acid sequences derived from two X. laevis cDNA clones that apparently code for Gsα subunits are 92% identical to those found in the short form of human Gsα. Despite this high homology, the X. laevis Gsα clones expressed in vitro, yielded a protein that are not able to activate the adenylyl cyclase present in S49 cyc− membranes in contrast with human Gsα 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 Gsα. 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 Gsα. These eight constructs were expressed in vitro and equivalent amounts of the resulting proteins were assayed in the activation of adenylyl cyclase with GTPγs and isoproterenol. The results obtained here clearly indicate that the Gα sequence that extends from amino acid 70 to 140, is important for the functionality of human Gsα in activating adenylyl cyclase