GroEL2 of Mycobacterium tuberculosis Reveals the Importance of Structural Pliability in Chaperonin Function

Abstract

Intracellular protein folding is mediated by molecular chaperones, the best studied among which are the chaperonins GroEL and GroES. Conformational changes and allosteric transitions between different metastable states are hallmarks of the chaperonin mechanism. These conformational transitions between three structural domains of GroEL are anchored at two hinges. Although hinges are known to be critical for mediating the communication between different domains of GroEL, the relative importance of hinges on GroEL oligomeric assembly, ATPase activity, conformational changes, and functional activity is not fully characterized. We have exploited the inability of Mycobacterium tuberculosis GroEL2 to functionally complement an Escherichia coli groEL mutant to address the importance of hinge residues in the GroEL mechanism. Various chimeras of M. tuberculosis GroEL2 and E. coli GroEL allowed us to understand the role of hinges and dissect the consequences of oligomerization and substrate binding capability on conformational transitions. The present study explains the concomitant conformational changes observed with GroEL hinge variants and is best supported by the normal mode analysis. IMPORTANCEConformational changes and allosteric transitions are hallmarks of the chaperonin mechanism. We have exploited the inability of M. tuberculosis GroEL2 to functionally complement a strain of E. coli in which groEL expression is repressed to address the importance of hinges. The significance of conservation at the hinge regions stands out as a prominent feature of the GroEL mechanism in binding to GroES and substrate polypeptides. The hinge residues play a significant role in the chaperonin activity in vivo and in vitro. M olecular chaperones are helper proteins, which play essential roles in folding, assembly, and transport of several cellular proteins (1, 2). Chaperonins, a subclass of the molecular chaperones, are homo-or hetero-oligomeric proteins, which carry out the substrate protein folding in a sequestered cavity. One of the best-characterized chaperonins is the 60-kDa chaperonin of Escherichia coli, GroEL (3). Chaperonins are highly conserved proteins and are known to interact with nonnative substrate proteins in an ATP-dependent manner. The E. coli genome possesses a single copy of groEL, arranged in an operonic arrangement with groES and expressed under all growth conditions (4). GroEL forms a cylindrical assembly with two heptameric rings and functions in coordination with the heptameric GroES (5-7). Each subunit of GroEL comprises three domains: (i) an equatorial domain (residues 2 to 133 and 409 to 548), which exhibits ATPase activity and participates in intersubunit and interring interactions; (ii) an apical domain (residues 191 to 374), which binds to substrate polypeptide and cochaperonin GroES; and (iii) an intermediate domain (residues 134 to 190 and 375 to 408), which links apical and equatorial domains in sequence and structure and mediates allosteric communication between the two (8, 9). The two rings...

A novel function of Mycobacterium tuberculosis chaperonin paralog GroEL1 in copper homeostasis

“…GroEL1, which belongs to the chaperonin family of proteins is unable to form higher oligomer. However, the conformational change seen in the apical domain corroborate the earlier study that apical domain requires greater flexibility for canonical chaperonin function [53]. It is also imperative to identify the residues involved in metal binding.…”

Mycobacterium tuberculosis GroEL2 Modulates Dendritic Cell Responses

“…Furthermore, data from biochemical and structural studies indicate that GroEL2 may be involved in promoting antigen presentation. GroEL2 possesses specific protein domains that have the potential to bind peptide substrates, a process that likely facilitates their subsequent association with MHC molecules (23,24). Together, our results highlight GroEL2 cleavage as a mechanism employed by M. tuberculosis to modulate DCmediated immunity during infection.…”

Functional Differences between E. coli and ESKAPE Pathogen GroES/GroEL

“…Although complete replacement of E. coli GroES/GroEL with the chaperonins from other organisms is possible ( 33 , 34 ), experiments where exogenous chaperonins were used to rescue GroES/GroEL-deficient E. coli , LG6 ( 18 ), have produced unexpected results ( 35 – 38 ). The most intriguing example came from the Mande group, where they studied the ability of Mycobacterium tuberculosis GroEL2 and E. coli / M. tuberculosis GroEL chimeras to rescue GroES/GroEL-deficient E. coli ( 39 ). Although M. tuberculosis GroEL2 is essential for M. tuberculosis survival, this chaperonin could not rescue GroES/GroEL-deficient E. coli despite significant amino acid identity with that of E. coli GroEL.…”