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Interaction analysis of TcrX/Y two component system from Mycobacterium tuberculosis
Author(s) -
Bhattacharya Monolekha,
Biswas Ashis,
Das Amit Kumar
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.709.1
Subject(s) - autophosphorylation , surface plasmon resonance , phosphorylation , in silico , crosstalk , chemistry , histidine kinase , signal transduction , two component regulatory system , histidine , mycobacterium tuberculosis , dissociation constant , biochemistry , biophysics , biology , mutant , enzyme , materials science , nanotechnology , protein kinase a , physics , receptor , medicine , tuberculosis , pathology , nanoparticle , optics , gene
TcrX/Y is one of the twelve two component system (TCS) present in Mycobacterium tuberculosis . The objective was to investigate the TcrX/Y interaction by in silico studies, pull down assay, radioactive phosphotransfer, surface plasmon resonance as well as crosstalk analysis of TcrY with TcrA ‐ a non‐cognate response regulator. Sequence alignment of TcrY with other histidine kinases revealed His256 as the residue responsible for autophosphorylation. Modeled structure of TcrX predicts that Asp59 may be involved in phosphotransfer with His256 of TcrY. Docking analysis pointed to the interaction of TcrY/His256 with TcrX/Asp54. TcrY/Y dimerisation was also observed by in silico and crosslinking studies. Autophosphorylation of TcrY has been observed followed by the phosphate transfer to TcrX. The phosphorylation process required divalent metal ions like Mg 2+ or Ca 2+ ions as evident from the radioactive phosphorylation studies. Interaction was not observed in TcrY/A suggesting the signal transduction process is specific in TcrX/Y system. TcrY hydrolyzes ATP and the K m value has been found to be 10 mM. TcrX/Y interaction has been determined by surface plasmon resonance and dissociation constant (K D ) was evaluated to be 3.6 μM. We conclude from our results that TcrX/Y are part of the same signal transduction pathway without their involvement in crosstalk with non‐cognate counterpart. The work was funded by Department of Biotechnology, Govt. of India.