Date of Award

2017

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Molecular Biology

Abstract

Post-translational modifications, such as N(epsilon)-lysine acetylation, are known to alter the behavior of transcriptional regulators in eukaryotes, but very little is known about the consequences of acetylation on transcriptional regulation in bacteria. Here, I provide evidence that a global transcriptional regulator of carbon metabolism, cAMP Receptor Protein (CRP), promotes both enzymatic and non-enzymatic lysine acetylation in E. coli. Non-enzymatic lysine acetylation occurs when cells ferment acetate, such as during growth on high concentrations of glucose. Intriguingly, CRP can be non-enzymatically acetylated on several lysines, including lysine 100 (K100). I provide evidence that neutralization of the K100 positive charge, as would occur upon K100 acetylation, has a dual effect on CRP activity. First, K100 neutralization decreases CRP activity at some Class II promoters. This decreased activity likely results from disruption of the interaction between Activating Region 2 (AR2) of CRP and the RNA polymerase alpha subunit N-terminal domain. Second, K100 neutralization increases the CRP half-life, leading to increased CRP steady state levels. Due to increased steady state levels, CRP activity is increased at some Class I promoters, in which CRP does not require AR2. Taken together, I propose that CRP promotes global acetylation, including CRP K100 acetylation, when cells are grown on glucose by positively regulating non-enzymatic acetylation. A consequence of K100 acetylation may be inverse regulation of Class II and Class I promoters under these conditions. This mechanism may help regulate carbon flux though central metabolism.

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