Protein-Protein Interactions in Prokaryotic Gene Expression

University Of Iowa, Iowa City IA

Investigators

Abstract

The Escherichia coli gcvTHP operon, coding the glycine cleavage enzyme system, provides one-carbon units for essential cellular methylation reactions. This operon is controlled by gcv -specific (GcvA and GcvR)and global-acting (Lrp, CRP and PurR)transcriptional regulatory proteins that are integrated into a regulatory network that allows the cell to respond to multiple environmental signals to either activate or repress transcription. The gcvB gene, encoding two small non-translated regulatory RNAs, is controlled solely by the GcvA and GcvR proteins. GcvA activates both operons in the presence of glycine, and represses the operons when glycine is limiting. Repression by GcvA requires the negative-acting GcvR protein. It is hypothesized that GcvR interacts directly with GcvA and not with DNA to repress the operons. Generally, repressors must bind to DNA to be functional. It is also hypothesized that a specific surface on GcvA interacts with different subunits of RNA polymerase to activate the gcvTHP operon and the gcvB gene. A combination of genetics, physiology and biochemistry will be used to address two major and interrelated questions to understand this regulatory network. First, what is the molecular mechanism used by the gcv -specific regulatory proteins GcvA and GcvR to control expression of the gcvTHP and gcvB operons? Second, how does a surface on the GcvA protein interact with different regions of RNA polymerase to activate transcription at the gcvTHP promoter versus the gcvB promoter? Since there are many functional and structural similarities between the transcriptional apparatus from both prokaryotic and eukaryotic organisms, understanding the mechanisms of bacterial transcription initiation will contribute to our general understanding of gene expression and its regulation.