STUDY OF THE ROLE OF SEPARATE AMINO-ACID- RESIDUES IN CGMP PHOSPHODIESTERASE-GAMMA SUBUNIT FROM BOVINE ROD OUTER SEGMENTS FUNCTIONING BY MEANS OF SITE-DIRECTED MUTAGENESIS - 2-STEP MECHANISM OF THE GAMMA-SUBUNIT INTERACTION WITH PHOSPHODIESTERASE CATALYTIC COMPLEX

The recombinant and 21 mutant forms of bovine rod phosphodiesterase ga mma subunit genes have been expressed by sequential transcription and translation in vitro. Inhibitory properties of these mutants and their interaction with phosphodiesterase catalytic alphabeta subunits and t ransducin alpha subunit have been studied. Two-step mechanism of the g amma subunit interaction with catalytic ones has been proposed: primar y binding and immediate inhibition. The central region of the gamma su bunit, enriched with basic amino acid residues, takes part In the prim ary binding, while the C-terminus plays a key role in inhibition. The spatial orientation of the gamma subunit C-terminal region is of great importance for inhibition. C-terminal part also demonstrates some inf luence on the gamma subunit affinity to phosphodiesterase catalytic co mplex.