Abstract
The ltsA gene of Corynebacterium glutamicum encodes a purF-type glutamine-dependent amidotransferase, and mutations in this gene result in increased susceptibility to lysozyme. Recently, it was shown that the LtsA protein catalyzes the amidation of diaminopimelate residues in the lipid intermediates of peptidoglycan biosynthesis. In this study, intracellular localization of wild-type and mutant LtsA proteins fused with green fluorescent protein (GFP) was investigated. The GFP-fused wild-type LtsA protein showed a peripheral localization pattern characteristic of membrane-associated proteins. The GFP-fusions with a mutation in the N-terminal domain of LtsA, which is necessary for the glutamine amido transfer reaction, exhibited a similar localization to the wild type, whereas those with a mutation or a truncation in the C-terminal domain, which is not conserved among the purF-type glutamine-dependent amidotransferases, did not. These results suggest that the C-terminal domain is required for peripheral localization. Differential staining of cell wall structures with fluorescent dyes revealed that formation of the mycolic acid-containing layer at the cell division planes was affected in the ltsA mutant cells. This was also confirmed by observation that bulge formation was induced at the cell division planes in the ltsA mutant cells upon lysozyme treatment. These results suggest that the LtsA protein function is required for the formation of a mycolic acid-containing layer at the cell division planes and that this impairment results in increased susceptibility to lysozyme.
Highlights
A non-pathogenic species of coryneform bacteria, Corynebacterium glutamicum, has been known as a glutamate-overproducing microorganism [1,2] and utilized for the production of various amino acids, such as lysine [3,4], valine [5] and threonine [6]
In order to investigate the roles of LtsA protein in cell surface formation of C. glutamicum, we examined the intracellular localization of fusion proteins featuring wild-type and mutant LtsA proteins with a green fluorescent protein (GFP) in C. glutamicum cells
S1), indicating that the LtsA-GFP fusion protein is functional in lysozyme sensitivity and temperature-sensitive growth of the ltsA mutant strain KY9714
Summary
A non-pathogenic species of coryneform bacteria, Corynebacterium glutamicum, has been known as a glutamate-overproducing microorganism [1,2] and utilized for the production of various amino acids, such as lysine [3,4], valine [5] and threonine [6]. Glutamate production by C. glutamicum is triggered by the limitation of biotin, which is essential for its growth, and addition of Tween 40 or penicillin [9,10,11]. Biotin is a cofactor for acetyl-CoA carboxylase, which is responsible for fatty acid biosynthesis. Penicillin inhibits the biosynthesis of peptidoglycan, which is a component of the bacterial cell wall. These inducing treatments for glutamate production affect cell surface formation in C. glutamicum. The mechanosensitive channel, NCgl1221, is responsible for the secretion of glutamate produced in C. glutamicum [12]. During glutamate production by C. glutamicum, it is thought that the NCgl1221 protein senses the change in tension of the cell surface and the resultant conformational
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
