Lysine acetylation of Hsp16.3: Effect on its structure, chaperone function and influence towards the growth of Mycobacterium tuberculosis

Subhashree Barik; Alok Kumar Panda; Viplov Kumar Biswas; Sheetal Das; Ayon Chakraborty; Shibangini Beura; Rahul Modak; Sunil Kumar Raghav; Rajiv K Kar; Ashis Biswas

doi:10.1016/j.ijbiomac.2024.131763

Lysine acetylation of Hsp16.3: Effect on its structure, chaperone function and influence towards the growth of Mycobacterium tuberculosis

Int J Biol Macromol. 2024 Apr 23;268(Pt 2):131763. doi: 10.1016/j.ijbiomac.2024.131763. Online ahead of print.

Authors

Affiliations

¹ School of Basic Sciences, Indian Institute of Technology Bhubaneswar, Bhubaneswar, India.
² Environmental Science Laboratory, School of Applied Sciences, Kalinga Institute of Industrial Technology, Deemed to be University, Bhubaneswar, Odisha 751024, India.
³ Immunogenomics and Systems Biology Laboratory, Institute of Life Sciences (ILS), Bhubaneswar, Odisha 751023, India; School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Bhubaneswar, Odisha, India.
⁴ Jyoti and Bhupat Mehta School of Health Sciences and Technology, Center for Nanotechnology, Indian Institute of Technology Guwahati, Assam, India.
⁵ University Institute of Biotechnology, University Centre for Research & Development, Chandigarh University, Mohali, India.
⁶ Infection and Epigenetics Laboratory, School of Biotechnology, Kalinga Institute of Industrial Technology (KIIT), Bhubaneswar, Odisha, India.
⁷ Immunogenomics and Systems Biology Laboratory, Institute of Life Sciences (ILS), Bhubaneswar, Odisha 751023, India.
⁸ School of Basic Sciences, Indian Institute of Technology Bhubaneswar, Bhubaneswar, India. Electronic address: abiswas@iitbbs.ac.in.

PMID: 38657928
DOI: 10.1016/j.ijbiomac.2024.131763

Abstract

Hsp16.3 plays a vital role in the slow growth of Mycobacterium tuberculosis via its chaperone function. Many secretory proteins, including Hsp16.3 undergo acetylation in vivo. Seven lysine (K) residues (K64, K78, K85, K114, K119, K132 and K136) in Hsp16.3 are acetylated inside pathogen. However, how lysine acetylation affects its structure, chaperone function and pathogen's growth is still elusive. We examined these aspects by executing in vitro chemical acetylation (acetic anhydride modification) and by utilizing a lysine acetylation mimic mutant (K64Q/K78Q/K85Q/K114Q/K119Q/K132Q/K136Q). Far- and near-UV CD measurements revealed that the chemically acetylated proteins(s) and acetylation mimic mutant has altered secondary and tertiary structure than unacetylated/wild-type protein. The chemical modification and acetylation mimic mutation also disrupted the oligomeric assembly, increased surface hydrophobicity and reduced stability of Hsp16.3, as revealed by GF-HPLC, 4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid binding and urea denaturation experiments, respectively. These structural changes collectively led to an enhancement in chaperone function (aggregation and thermal inactivation prevention ability) of Hsp16.3. Moreover, when the H37Rv strain expressed the acetylation mimic mutant protein, its growth was slower in comparison to the strain expressing the wild-type/unacetylated Hsp16.3. Altogether, these findings indicated that lysine acetylation improves the chaperone function of Hsp16.3 which may influence pathogen's growth in host environment.

Keywords: Lysine acetylation; Molecular chaperone; Mycobacterium tuberculosis Hsp16.3; Small heat shock proteins (sHsps); Tuberculosis.