Intragenic Suppression of Gal3^sup C^ Interaction With Gal80 in the Saccharomyces cerevisiae GAL Gene Switch

Gal4-mediated activation of GAL gene transcription in Saccharomyces cerevisiae requires the interaction of Gal3 with Gal80, the Gal4 inhibitor protein. While it is known that galactose and ATP activates Gal3 interaction with Gal80, neither the mechanism of activation nor the surface that binds to Gal80 is known. We addressed this through intragenic suppression of GAL3^sup C^ alleles that cause galactose-independent Gal3-Gal80 interaction. We created a new allele, GAL^sup SOC^, and showed that it suppressed a new GAL3^sup C^ allele. We tested the effect of GAL^sup SOC^ on several newly isolated and existing GAL3^sup C^ alleles that map throughout the gene. All except one GAL3^sup C^ allele, D368V, were suppressible by GAL3^sup SOC^. GAL3^sup SOC^ and all GAL3^sup C^ alleles were localized on a Gal3 homology model that is based on the structure of the highly related Gal1 protein. These results provide evidence for allosterism in the galactose- and ATP-activation of Gal3 binding to Gal80. In addition, because D368V and residues corresponding to Gal80-nonbinder mutations colocalized to a domain that is absent in homologous proteins that do not bind to Gal80, we suggest that D368 is a part of the Gal80-binding surface. [PUBLICATION ABSTRACT]