Investigation of the physiological significance of protein acetylation in Bacillus subtilis

ABSTRACT
Until recently, N-lysine acetylation was thought to be rare in bacteria, but is now appreciated to affect hundreds of
bacterial proteins with diverse cellular functions. Acetylation was initially discovered as a post-translational modification
(PTM) on the unstructured, highly basic N-terminal tails of eukaryotic histones. Histone acetylation constitutes part of the
“histone code,” and regulates chromosome compaction and various DNA processes, such as gene expression, replication,
repair and recombination. In eukaryotes, acetylation regulates many other proteins in addition to histones, involved in a
wide array of important biological processes. This observation is also true in bacteria, as evidenced by the characterization
of the acetylomes of more than 30 different bacterial species. However, the physiological significance of the vast majority
of these modifications remains unknown. In addition, the mechanisms of acetylation and deacetylation, and the
bacterial enzymes involved are not completely understood. To address these gaps in knowledge, we have focused on
studying the acetylation of the essential, histone-like protein HBsu in Bacillus subtilis. In bacteria, the nucleoid is
compacted and organized by the action of nucleoid-associated proteins (NAPs). HBsu is a member of the most widely
conserved NAP family, and is considered a functional equivalent of eukaryotic histones. We found that HBsu contains
seven novel acetylation sites, and this raised the exciting possibility that these modifications represent a “histone-like”
code in bacteria. So far, we discovered that acetylation of HBsu at key lysine residues is required to maintain normal
chromosome compaction. Additionally, we identified the second protein acetyltransferase in B. subtilis. The overall goal
of our research program is to decipher this code. Our recent progress supports the hypotheses that acetylation of HBsu
regulates cell division and sporulation, and that there are additional enzymes involved in regulating acetylation. The short-
term goals of this work are to define the enzymatic mechanism of regulation of HBsu acetylation and determine the
significance of HBsu acetylation in the regulation of DNA transactions, stationary phase development and drug tolerance.
Additionally, we will develop new biochemical and mass-spectrometry based proteomics techniques for the study of
acetylation in bacteria. Our long-term goals are to characterize additional HBsu PTMs, identify and characterize novel
enzymes of acetylation, and perform a detailed structural and biochemical analysis with acetylated HBsu and novel
enzymes. Ultimately, we will design novel inhibitors of bacterial acetylation enzymes or acetylated HBsu and assess their
efficacy as potential novel antimicrobial therapies. Together, these studies may demonstrate the existence of a histone-
like code in bacteria, an unexpected and exciting new field of biology. Furthermore, these studies will provide the
foundation for designing novel antimicrobial drugs that target protein acetylation, either the enzymes or key acetylated
targets.

Grant Number: 5R35GM138303-05
NIH Institute/Center: NIH
Principal Investigator: Valerie Carabetta