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Rickettsial species have independently lost several genes owing to reductive evolution while retaining those predominantly implicated in virulence, survival, and biosynthetic pathways. In this study, we have identified a previously uncharacterized R. conorii gene RC0497 as an N-acetylmuramoyl-L-alanine amidase constitutively expressed during infection of cultured human microvascular endothelial cells at the levels of both mRNA transcript and encoded protein. Homology based search of rickettsial genomes reveals that RC0497 homologs, containing amidase_2 family and peptidoglycan binding domains, are highly conserved among spotted fever group rickettsiae. The recombinant RC0497 protein exhibits α-helix secondary structure, undergoes a conformational change in the presence of zinc, and exists as a dimer at higher concentrations. We have further ascertained the enzymatic activity of RC0497 via demonstration of its ability to hydrolyze Escherichia coli peptidoglycan. Confocal microscopy on E. coli expressing RC0497 and transmission immunoelectron microscopy of R. conorii revealed its localization predominantly to the cell wall, septal regions of replicating bacteria, and the membrane of vesicles pinching off the cell wall. In summary, we have identified and functionally characterized RC0497 as a peptidoglycan hydrolase unique to spotted fever rickettsiae, which may potentially serve as a novel moonlighting protein capable of performing multiple functions during host-pathogen interactions.
This article was published in the following journal.
Name: Biological chemistry
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An autolytic enzyme bound to the surface of bacterial cell walls. It catalyzes the hydrolysis of the link between N-acetylmuramoyl residues and L-amino acid residues in certain cell wall glycopeptides, particularly peptidoglycan. EC 220.127.116.11.
The interdisciplinary science that studies evolutionary biology, including the origin and evolution of the major elements required for life, their processing in the interstellar medium and in protostellar systems. This field also includes the study of chemical evolution and the subsequent interactions between evolving biota and planetary evolution as well as the field of biology that deals with the study of extraterrestrial life.
Evolution at the molecular level of DNA sequences and proteins. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)
A hexosyltransferase involved in the transfer of disaccharide molecules to the peptidoglycan structure of the CELL WALL SKELETON. It plays an important role in the genesis of the bacterial CELL WALL.
The techniques used to produce molecules exhibiting properties that conform to the demands of the experimenter. These techniques combine methods of generating structural changes with methods of selection. They are also used to examine proposed mechanisms of evolution under in vitro selection conditions.
Bioinformatics is the application of computer software and hardware to the management of biological data to create useful information. Computers are used to gather, store, analyze and integrate biological and genetic information which can then be applied...
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