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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 (Hsp16.3-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. • Lysine acetylation in Hsp16.3 perturbs its secondary, tertiary and quaternary structure. • This post-translational modification enhances the surface hydrophobicity of Hsp16.3. • The chaperone function of Hsp16.3 is enhanced upon lysine acetylation. • The improved chaperone function of Hsp16.3 influences the growth of H37Rv strain. [ABSTRACT FROM AUTHOR]

• Two biosurfactants (Taurocholic acid and Taurodeoxycholic acid) efficiently inhibit the primary fibrillation of insulin. • TDCA is also able to suppress the secondary fibrillation of insulin. • Both these bile acids attenuate insulin fibril-induced cellular toxicity. • They interact to the C-ter of insulin B-chain via hydrophobic & H-bond interactions. • Such interactions stabilize insulin secondary structure and reduce the aggregate size. Insulin-induced amyloidosis, a typical example of protein fibrillation, reduces the effectiveness of insulin treatment in type-II diabetic patients, which demands intense investigations to mitigate insulin fibrillation. Herein, the inhibitory effect of two taurine conjugated bile acids [taurocholic acid (TCA) and taurodeoxycholic acid (TDCA)] towards insulin fibrillation has been elucidated using various biophysical, imaging and computational tools. TCA delays lag time of fibrillation while TDCA targets fibril elongation step and thus impeding insulin primary fibrillation. TDCA also inhibits the secondary fibrillation of insulin efficiently. The greater inhibition of both fibrillation processes is evidenced once insulin is kept with TDCA under fibrillogenic conditions. In silico studies reveal that these two bile acids interact with several aggregation-prone residues (e.g. Tyr16, Phe24, Tyr26) present at the C-terminal portion of insulin B-chain. Such interactions stabilize the native insulin structure, reduce the surface hydrophobicity and aggregate size as well as modulate the fibril morphology. Both these molecules are non-toxic and attenuate insulin fibril-induced hemolysis of erythrocytes and cytotoxicity in HepG2 cells. Overall, our findings provide novel insights to the study of amyloidal protein fibrillation and lay foundation towards the utilization of these two taurine conjugated bile acids as inhibitors of insulin fibrillation and insulin-induced amyloidosis. [ABSTRACT FROM AUTHOR]