Characterization of an extracellular protein, Rv1076 from M. tuberculosis with a potential role in humoral response.
Many mycobacterial proteins involved in lipid metabolism are reported to be essential for survival and pathogenesis of M. tuberculosis. Rv1076 of M. tuberculosis has been annotated as a putative esterase/lipase based on the consensus sequence 'GXSXG'. It is conserved in all the mycobacterial species. Therefore, in the present study we have characterized Rv1076 gene product in detail. The gene rv1076 was expressed in E. coli and purified from inclusion bodies with approx. 40% yield. The protein showed high specific activity with pNP- butyrate as preferred substrate. The enzyme was stable upto 50°C and in pH range of 6-8 i.e. under acidic conditions. Ser-140, Glu-239 and His-269 were confirmed as active site residues using site directed mutagenesis. The specific activity, Kmand Vmaxof enzyme was determined to be 177Umg-1protein, 334μM and 262μmolml-1min-1, respectively. Western blot analysis established Rv1076 to be an extracellular protein. Several putative immunodominant epitopes were predicted in Rv1076. Rv1076 elicited strong humoral response in both extrapulmonary and relapsed cases of TB patients. Therefore, we conclude that Rv1076 is a novel secretory esterase of M. tuberculosis which could be a potential immunodominant antigen of M. tuberculosis.
Department of Biotechnology, BMS Block-I, South Campus, Panjab University, Sector 25, Chandigarh, 160014, India. Electronic address: sangha.gurkamal@gmail.com.
Gurkamaljit
G
Kaur
Department of Pulmonary Medicine, Government Medical College and Hospital, Sec-32, Chandigarh, India. Electronic address: saini_vks@yahoo.com.
Varinder
V
Saini
Department of Biotechnology, BMS Block-I, South Campus, Panjab University, Sector 25, Chandigarh, 160014, India. Electronic address: vandanadhiman21@gmail.com.
Bandana
B
Kumari
Department of Biochemistry, Government Medical College and Hospital, Sec-32, Chandigarh, India. Electronic address: jasbinderkaur@yahoo.co.in.
Jasbinder
J
Kaur
Department of Biotechnology, BMS Block-I, South Campus, Panjab University, Sector 25, Chandigarh, 160014, India. Electronic address: jagsekhon@yahoo.com.
Jagdeep
J
Kaur
D000907:Antibodies, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D003001:Cloning, Molecular; D017124:Conserved Sequence; D005110:Extracellular Space / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D056724:Immunity, Humoral; D007700:Kinetics; D008958:Models, Molecular; D009169:Mycobacterium tuberculosis / Q000166:cytology* / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D011487:Protein Conformation; D021381:Protein Transport; D013696:Temperature; D014397:Tuberculosis, Pulmonary / Q000097:blood / Q000276:immunology
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins
Esterase; Extracellular; Humoral response; Lip gene family; M. tuberculosis; Rv1076