A Molecular Biological and Biochemical Investigation on Mycobacterium tuberculosis MutT Protein

نویسندگان

  • Hsiu-Lin Huang
  • Ho-Ting Su
  • Chung-Hsiun Herbert Wu
  • Jyy-Jih Tsai-Wu
چکیده

BACKGROUND Mycobacterium tuberculosis is a vicious microbe co-existing with the infected host. This pathogen exploited opportunities to spread during periods of urbanization and social upheaval, and got retreated with improved hygiene. OBJECTIVES This investigation was designed to clone and characterize M. tuberculosis mutT gene, a homologue of a DNA repair protein in Escherichia coli. The aim was to depict the possible role of this homologue in the virulent microbe. MATERIALS AND METHODS A DNA fragment of the mutT gene was amplified with PCR from the genomic DNA of strain H37Rv M. tuberculosis. The expression vector was transformed into E. coli strains BL21 (DE3) and MK602 (DE3) (mutT-). The protein activity assay was performed by biochemical methods. RESULTS M. tuberculosis MutT shares 23% identity with the E. coli MutT protein. The mutT gene DNA fragment was subcloned into the expression vector pET28a(+) and the recombinant plasmid was overexpressed in E. coli. Purified and refolded M. tuberculosis MutT possesses a dGTPase activity, which is one of the most well-known preference nucleotidase activities of MutT in E. coli. This study also showed that the dGTPase activity of M. tuberculosis MutT was enhanced by magnesium and inhibited by Ni(2+) or EDTA. Endogenous MutT protein in M. tuberculosis lysate displayed a smear pattern in the Western blot, suggesting instability of this protein in the bacteria similar to the important proteins, such as P53 protein, tightly regulated by protein degradation. CONCLUSIONS The cloned M. tuberculosis mutT gene and MutT protein were characterized. M. tuberculosis MutT has a dGTPase activity, which is one of the most well-known preference nucleotidase activities of MutT in E. coli. These findings provide further understanding about the vicious bacterium.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Role of Molecular Interactions and Oligomerization in Chaperone Activity of Recombinant Acr from Mycobacterium tuberculosis

Background: The chaperone activity of Mycobacterium tuberculosis Acr is an important function that helps to prevent misfolding of protein substrates inside the host, especially in conditions of hypoxia. Objectives: The aim of this study was to establish the correlation of structure and function of recombinant Acr proteins both before and after ge...

متن کامل

Biochemical properties of MutT2 proteins from Mycobacterium tuberculosis and M. smegmatis and their contrasting antimutator roles in Escherichia coli.

Mycobacterium tuberculosis, the causative agent of tuberculosis, is at increased risk of accumulating damaged guanine nucleotides such as 8-oxo-dGTP and 8-oxo-GTP because of its residency in the oxidative environment of the host macrophages. By hydrolyzing the oxidized guanine nucleotides before their incorporation into nucleic acids, MutT proteins play a critical role in allowing organisms to ...

متن کامل

Biochemical characterization of PE_PGRS61 family protein of Mycobacterium tuberculosis H37Rv reveals the binding ability to fibronectin

Objective(s): The periodic binding of protein expressed by Mycobacterium tuberculosis H37Rv with the host cell receptor molecules i.e. fibronectin (Fn) is gaining significance because of its adhesive properties.  The genome sequencing of M. tuberculosis H37Rv revealed that the proline-glutamic (PE) proteins contain polymorphic GC-rich repetitive sequences (PGRS) which have clinical importance i...

متن کامل

PURIFICATION AND CHARACTERIZATION OF PROTEIN ANTIGENS ISOLATED FROM MYCOBACTERIUM TUBERCULOSIS (H37Rv STRA IN) AND THEIR EFFECTS ON CELL-MEDIATED IMMUNE RESPONSES IN GUINEA PIGS

Mycobacterium tuberculosis (H37Rv strain) was used in this study. The bacterial cells were disintegrated by sonication. The separation and characterization of the soluble molecules were attempted by various techniques including gel filtration, ion exchange chromatographies and polyacrylamide gel electrophoresis, using SDS and 2ME. Eight protein molecules with molecular weights ranging from...

متن کامل

Identification of Mycobacterium Tuberculosis Complex, Using Molecular Methods

Abstract Background and Objective: A high level of homogeneity observed within all bacteria in the Mycobacterium tuberculosis complex makes a property that seriously challenges traditional biochemical-based identification methods of these pathogens in the laboratory. The work presented here was conducted to characterize Mycobacterium tuberculosis complex isolates in Golestan, Northern Iran. ...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:

دوره 7  شماره 

صفحات  -

تاریخ انتشار 2014