Intragenic suppression of Gal3C 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 GAL3C alleles that cause galactose-independent Gal3-Gal80 interaction. We created a new allele, GAL3SOC, and showed that it suppressed a new GAL3C allele. We tested the effect of GAL3SOC on several newly isolated and existing GAL3C alleles that map throughout the gene. All except one GAL3C allele, D368V, were suppressible by GAL3SOC. GAL3SOC and all GAL3C 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.
Diep
Cuong Q
CQ
Department of Biochemistry and Molecular Biology, Pennsylvania State University, College of Medicine, Hershey, Pennsylvania 17033, USA.
Peng
Gang
G
Bewley
Maria
M
Pilauri
Vepkhia
V
Ropson
Ira
I
Hopper
James E
JE
D000255:Adenosine Triphosphate / Q000378:metabolism; D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D005690:Galactose / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D011485:Protein Binding; D011487:Protein Conformation; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013489:Suppression, Genetic; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D000519:alpha-Galactosidase / Q000378:metabolism
C055002:GAL80 protein, S cerevisiae; C099289:Gal3 protein, S cerevisiae; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000255:Adenosine Triphosphate; D000519:alpha-Galactosidase; D005690:Galactose