Date of Degree
PhD (Doctor of Philosophy)
David S. Weiss
First Committee Member
Craig D Ellermeier
Second Committee Member
Robert C Piper
Third Committee Member
Fourth Committee Member
Timothy L Yahr
Rare lipoprotein A (RlpA) has been studied previously only in Escherichia coli, where it localizes to the septal ring and scattered foci along the lateral wall, but mutants have no phenotypic change. In this thesis, we show rlpA mutants of Pseudomonas aeruginosa form chains of short, fat cells when grown in media of low osmotic strength. These morphological defects indicate RlpA is needed for efficient separation of daughter cells and maintenance of rod shape. Analysis of peptidoglycan sacculi from a ΔrlpA mutant revealed increased tetra and hexasaccharides that lack stem peptides (hereafter called "naked glycans"). Incubation of these sacculi with purified RlpA resulted in release of naked glycans containing 1,6-anhydro N-acetylmuramic acid ends. RlpA did not degrade sacculi from wild-type cells unless the sacculi were subjected to a limited digestion with an amidase to remove some of the stem peptides. Collectively, these findings indicate RlpA is a lytic transglycosylase with a strong preference for naked glycan strands. We propose that RlpA activity is regulated in vivo by substrate availability, and that amidases and RlpA work in tandem to degrade peptidoglycan in the division septum and lateral wall.
Our discovery that RlpA from P. aeruginosa is a lytic transglycosylase motivated us to reinvestigate RlpA from E. coli. We confirmed predictions that RlpA of E. coli is an outer membrane protein and determined its abundance to be about 600 molecules per cell. However, multiple efforts to demonstrate that E. coli RlpA is a lytic transglycosylase were unsuccessful and the function of this protein in E. coli remains obscure.
Double-psi beta barrel domain, Lytic transglycosylase, Outer membrane protein, Peptidoglycan, Septal ring, SPOR domain
xii, 155 pages
Includes bibliographical references (pages 140-155).
Copyright 2014 Matthew Allan Jorgenson
Jorgenson, Matthew Allan. "A tale of two RLPAs : studies of cell division in Escherichia coli and Pseudomonas aeruginosa." PhD (Doctor of Philosophy) thesis, University of Iowa, 2014.