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BMC Microbiology

, 14:174

Clinical microbiology and vaccines

Abstract

BackgroundMany bacteria modulate and evade the immune defenses of their hosts through peptidoglycan PG deacetylation.
The PG deacetylases from Streptococcus pneumonia, Listeria monocytogenes and Lactococcus lactis have been characterized.
However, thus far, the PG deacetylase of Mycobacterium tuberculosis has not been identified.

ResultsIn this study, we cloned the Rv1096 gene from the M.
tuberculosis H37Rv strain and expressed Rv1096 protein in both Escherichia coli and M.
smegmatis.
The results showed that the purified Rv1096 protein possessed metallo-dependent PG deacetylase activity, which increased in the presence of Co.
The kinetic parameters of the PG deacetylase towards M.
smegmatis PG as a substrate were as follows: Km, 0.910 ± 0.007 mM; Vmax, 0.514 ± 0.038 μMmin; and Kcat = 0.099 ± 0.007 S.
Additionally, the viability of M.
smegmatis in the presence of over-expressed Rv1096 protein was 10-fold higher than that of wild-type M.
smegmatis after lysozyme treatment.
Additionally, light microscopy and scanning electron microscopy showed that in the presence of over-expressed Rv1096 protein, M.
smegmatis kept its regular shape, with an undamaged cell wall and smooth surface.
These results indicate that Rv1096 caused deacetylation of cell wall PG, leading to lysozyme resistance in M.
smegmatis.

ConclusionWe have determined that M.
tuberculosis Rv1096 is a PG deacetylase.
The PG deacetylase activity of Rv1096 contributed to lysozyme resistance in M.
smegmatis.
Our findings suggest that deacetylation of cell wall PG may be involved in evasion of host immune defenses by M.
tuberculosis.

KeywordsMycobacterium tuberculosis Cell wall Rv1096 Peptidoglycan deacetylase Lysozyme AbbreviationsPGPeptidoglycan

mAGPMycolyl-arabinogalactan-peptidoglycan

NodNucleotide-binding oligomerization domain

IPTGIsopropyl-D-thiogalactopyranoside

PMSFPhenylmethyl-sulphonyl fluoride

BCIP-NBT5-Bromo-4-chloro-3-indolyl phosphate-Nitro blue tetrazolium

SDSSodium dodecyl sulfate

CFUColony forming unit

OsO4Omium tetroxide

SEMScanning electronic microscopy

CE-4Carbohydrate esterase 4.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2180-14-174 contains supplementary material, which is available to authorized users.

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Autor: Shufeng Yang - Fei Zhang - Jian Kang - Wenli Zhang - Guoying Deng - Yi Xin - Yufang Ma

Fuente: https://link.springer.com/



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