The EccC enzyme of Mycobacterium tuberculosis ESX-1 secretion system is involved in EsxAB virulence factor secretion and offers an attractive target for antivirulence inhibitors development against M. tuberculosis. The EccCb1 polypeptide of the EccC enzyme contains two Ftsk/SpoIIIE type ATPase domains (D2 and D3) and binds to the EsxAB factor at the C-terminal region of the D3 domain. In the current study, we have determined a low-resolution structure of EccCb1, and its mechanism involved in ATPase activity and EsxAB factor binding. Small-angle X-ray scattering data yielded a double hexameric ring structure of EccCb1 in solution and was further confirmed by SEC-MALS and dynamic light scattering. ATPase activity of wild-type, D2, and D3 mutants showed that D2-K90A and D3-K382A mutations led to a complete loss of enzyme activity. The full-length EccCb1 showed ∼3.7-fold lower catalytic efficiency than D2 domain and ∼1.7 fold lower than D3 domain. The EsxAB factor binds EccCb1 with Kd ∼ 11.3 ± 0.6 nM and its affinity is enhanced ∼2 fold in presence of ATP + Mg2+. These data indicate the involvement of ATPase activity in EsxAB factor translocation. Molecular dynamics simulation on wild-type, ATP + Mg2+, and EsxAB + ATP + Mg2+ bound EccCb1 double-ring structure showed enhanced stability of enzyme upon ATP + Mg2+ and EsxAB binding. Overall, our study showed a low-resolution structure of EccCb1, and the mechanism involved in ATPase activity and EsxAB factor recognition, which can be targeted for the development of antivirulence drugs against M. tuberculosis.
Skip Nav Destination
Article navigation
July 2022
-
Cover Image
Cover Image
The cover image shows the double hexameric ring structures of Mycobacterium tuberculosis EccCb1 in apo and EsxAB+ATP+Mg2+ bound forms. The binding affinity EsxAB substrate to EccCb1 is enhanced ~2-fold in presence of ATP and Mg2+, which indicate the involvement of ATPase activity in EsxAB substrate translocation. Read more from Saxena and colleagues on pages 1559–1579
Research Article|
July 21 2022
Binding of the Mycobacterium tuberculosis EccCb1 ATPase double hexameric ring to the EsxAB virulence factor is enhanced by ATP
Arkita Bandyopadhyay;
Arkita Bandyopadhyay
1Rm-403/440, Structural Biology Lab, School of Life Sciences, Jawaharlal Nehru University, New Delhi 67, India
Search for other works by this author on:
Ramesh Kumar;
Ramesh Kumar
Investigation
1Rm-403/440, Structural Biology Lab, School of Life Sciences, Jawaharlal Nehru University, New Delhi 67, India
Search for other works by this author on:
Jyotsna Singh;
Jyotsna Singh
Investigation
2National Center for Cell Science, Savitribai Phule Pune University Campus, Ganeshkhind, Pune 411007, India
Search for other works by this author on:
Ajay K. Saxena
Conceptualization, Resources, Software, Supervision, Funding acquisition, Validation, Investigation, Methodology, Writing - original draft, Project administration, Writing - review & editing
1Rm-403/440, Structural Biology Lab, School of Life Sciences, Jawaharlal Nehru University, New Delhi 67, India
Correspondence: Ajay K. Saxena (ajaysaxena@mail.jnu.ac.in)
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
June 10 2021
Revision Received:
June 29 2022
Accepted:
June 30 2022
Accepted Manuscript online:
June 30 2022
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 2022 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society
2022
Biochem J (2022) 479 (14): 1559–1579.
Article history
Received:
June 10 2021
Revision Received:
June 29 2022
Accepted:
June 30 2022
Accepted Manuscript online:
June 30 2022
Citation
Arkita Bandyopadhyay, Ramesh Kumar, Jyotsna Singh, Ajay K. Saxena; Binding of the Mycobacterium tuberculosis EccCb1 ATPase double hexameric ring to the EsxAB virulence factor is enhanced by ATP. Biochem J 29 July 2022; 479 (14): 1559–1579. doi: https://doi.org/10.1042/BCJ20210430
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.
Biochemical Society Member Sign in
Sign InSign in via your Institution
Sign in via your InstitutionGet Access To This Article
399
Views