G-Quadruplex-Forming DNA Aptamers Inhibit the DNA-Binding Function of HupB and Mycobacterium tuberculosis Entry into Host Cells

Priya Kalra; Subodh Kumar Mishra; Surinder Kaur; Amit Kumar; Hanumanthappa Krishna Prasad; Tarun Kumar Sharma; Jaya Sivaswami Tyagi

doi:10.1016/j.omtn.2018.08.011

G-Quadruplex-Forming DNA Aptamers Inhibit the DNA-Binding Function of HupB and Mycobacterium tuberculosis Entry into Host Cells

Mol Ther Nucleic Acids. 2018 Dec 7:13:99-109. doi: 10.1016/j.omtn.2018.08.011. Epub 2018 Aug 22.

Authors

Priya Kalra¹, Subodh Kumar Mishra², Surinder Kaur¹, Amit Kumar², Hanumanthappa Krishna Prasad¹, Tarun Kumar Sharma³, Jaya Sivaswami Tyagi⁴

Affiliations

¹ Department of Biotechnology, All India Institute of Medical Sciences, New Delhi, Delhi 110029, India.
² Discipline of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Simrol, Madhya Pradesh 453552, India.
³ Centre for Biodesign and Diagnostics, Translational Health Science and Technology Institute, Faridabad, Haryana 121001, India. Electronic address: tarun@thsti.res.in.
⁴ Department of Biotechnology, All India Institute of Medical Sciences, New Delhi, Delhi 110029, India; Centre for Biodesign and Diagnostics, Translational Health Science and Technology Institute, Faridabad, Haryana 121001, India. Electronic address: jayatyagi.aiims@gmail.com.

Abstract

The entry and survival of Mycobacterium tuberculosis (Mtb) within host cells is orchestrated partly by an essential histone-like protein HupB (Rv2986c). Despite being an essential drug target, the lack of structural information has impeded the development of inhibitors targeting the indispensable and multifunctional C-terminal domain (CTD) of HupB. To bypass the requirement for structural information in the classical drug discovery route, we generated a panel of DNA aptamers against HupB protein through systemic evolution of ligands by exponential (SELEX) enrichment. Two G-quadruplex-forming high-affinity aptamers (HupB-4T and HupB-13T) were identified, each of which bound two distinct sites on full-length HupB, with an estimated K_D of ∼1.72 μM and ∼0.17 μM, respectively, for the high-affinity sites. While HupB-4T robustly inhibited DNA-binding activity of HupB in vitro, both the aptamers recognized surface-located HupB and significantly blocked Mtb entry into THP-1 monocytic cells (p < 0.0001). In summary, DNA aptamers generated in this study block DNA-binding activity of HupB, inhibit virulent Mtb infection in host cells, and demonstrate aptamers to be inhibitors of HupB functions. This study also illustrates the utility of SELEX in developing inhibitors against essential targets for whom structural information is not available.

Keywords: G-quadruplex; HupB; Mtb; Mycobacterium tuberculosis; SELEX; aptamers.