Knockout of the alanine racemase gene in Aeromonas hydrophila HBNUAh01 results in cell wall damage and enhanced membrane permeability

This study focused on the alanine racemase gene (alr-2), which is involved in the synthesis of d-alanine that forms the backbone of the cell wall. A stable alr-2 knockout mutant of Aeromonas hydrophila HBNUAh01 was constructed. When the mutant was supplemented with d-alanine, growth was unaffected;...

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Bibliographic Details
Published in:FEMS microbiology letters 2015-07, Vol.362 (13), p.fnv089-fnv089
Main Authors: Liu, Dong, Zhang, Lu, Xue, Wen, Wang, Yaping, Ju, Jiansong, Zhao, Baohua
Format: Article
Language:English
Subjects:
Aeromonas hydrophila
Aeromonas hydrophila - enzymology
Aeromonas hydrophila - genetics
Aeromonas hydrophila - physiology
Aeromonas hydrophila - ultrastructure
Alanine
Alanine - metabolism
Alanine racemase
Alanine Racemase - genetics
Alanine Racemase - metabolism
Cell culture
Cell Membrane Permeability - genetics
Cell Wall - genetics
Cell Wall - metabolism
Cell Wall - ultrastructure
Cell walls
D-Alanine
Deactivation
Deprivation
Gene Knockout Techniques
Inactivation
Lysis
Membrane permeability
Membranes
Microbial Viability - genetics
Microbiology
Mutants
Mutation
Perforation
Permeability
Transmission electron microscopy
Viability
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Summary:This study focused on the alanine racemase gene (alr-2), which is involved in the synthesis of d-alanine that forms the backbone of the cell wall. A stable alr-2 knockout mutant of Aeromonas hydrophila HBNUAh01 was constructed. When the mutant was supplemented with d-alanine, growth was unaffected; deprivation of d-alanine caused the growth arrest of the starved mutant cells, but not cell lysis. No alanine racemase activity was detected in the culture of the mutant. Additionally, a membrane permeability assay showed increasing damage to the cell wall during d-alanine starvation. No such damage was observed in the wild type during culture. Scanning and transmission electron microscopy analyses revealed deficiencies of the cell envelope and perforation of the cell wall. Leakage of UV-absorbing substances from the mutants was also observed. Thus, the partial viability of the mutants and their independence of d-alanine for growth indicated that inactivation of alr-2 does not impose an auxotrophic requirement for d-alanine. Knockout of the alanine racemase gene in Aeromonas hydrophila HBNUAh01 results in cell wall damage and enhanced membrane permeability.
ISSN:1574-6968
0378-1097
1574-6968
DOI:10.1093/femsle/fnv089