Functional similarities between the small heat shock proteins Mycobacterium tuberculosis HSP 16.3 and human αB‐crystallin

Abstract

Mycobacterium tuberculosis heat shock protein 16.3 (MTB HSP 16.3) accumulates as the dominant protein in the latent stationary phase of tuberculosis infection. MTB HSP 16.3 displays several characteristics of small heat shock proteins (sHsps): its expression is increased in response to stress, it protects against protein aggregation in vitro, and it contains the core Ôa-crystallinÕ domain found in all sHsps. In this study we characterized the chaperone activity of recombinant MTB HSP 16.3 in several different assays and compared the results to those obtained with recombinant human aB-crystallin, a well characterized member of the sHsp family. Recombinant MTB HSP 16.3 was expressed in Escherichia coli and purified to apparent homogeneity. Similar to aB-crystallin, MTB HSP16.3 suppressed citrate synthase aggregation and in the presence of 3.5 mM ATP the chaperone activity was enhanced by twofold. ATP stabilized MTB HSP 16.3 against proteolysis by chymotrypsin, and no effect was observed with ATPcS, a nonhydrolyzable analog of ATP. Increased expression of MTB HSP 16.3 resulted in protection against thermal killing in E. coli at 48°C. While the sequence similarity between human aB-crystallin and MTB HSP 16.3 is only 18%, these results suggest that the functional similarities between these proteins containing the core Ôa-crystallinÕ domain are much closer.Keywords: ATP; human aB-crystallin; molecular chaperone; Mycobacterium tuberculosis HSP 16.3; small heat shock proteins.One-third of the world's population is infected with latent inactive tuberculosis and active tuberculosis is the leading cause of death due to an infectious disease [1]. Each year, new infections occur in 54 million people; 6.8 million people develop clinical disease, and 2.4 million cases result in death [2]. There is still limited knowledge of the molecular pathogenesis of the latent stage of this organism [3]. Individuals who have been infected with Mycobacterium tuberculosis can harbor stable dormant bacilli for decades before developing an active infection later in life [4]. Recent reports indicate an important role for M. tuberculosis (MTB) heat shock protein (HSP) 16.3 in the survival of MTB during prolonged periods of infection [5][6][7]. It was shown that MTB HSP 16.3, initially described as the immunodominant 14-or 16-kDa antigen [8][9][10][11], was a major component in tuberculosis infection in humans and played an important role in enhancing protein stability and survival [5]. Eighty-five percent of patients with active tuberculosis showed a positive reaction to this antigen, suggesting that this protein expressed in vivo had a key role in MTB infection [11,12]. The 14K antigen was later renamed MTB HSP 16.3 [13]. MTB HSP 16.3 accumulates to become the dominant protein in the latent stationary phase of M. tuberculosis infection [7]. Over-expression of HSP 16.3 in log phase growth of M. tuberculosis slowed the growth rate and protected against stationary phase autolysis in vitro [7]. MTB HSP 16.3 has been characterized as a membrane asso...

Is mycobacterial heat shock protein 16kDa, a marker of the dormant stage of Mycobacterium tuberculosis, a sarcoid antigen?

“…Mtb-hsp16), a bystander activation, and an anergy altering an expression of costimulatory molecules, regulatory T-cells, phagocytosis, apoptosis, or oxidative stress [3,[8][9][10][11][19][20][21][22][23][24][25][26][27][28][29][30]36]. It is worthwhile to note that cytotoxicity is often directly attributed to NO, especially under conditions of induced oxidative stress resulting from overproduction of superoxide at the sites of inflammation.…”

“…Mtb-hsp16, with following persistent antigenemia, immunocomplexemia and increased local proliferation of B-and T-cells with granuloma formation (Fig. 1) [1,3,9,[39][40][41][42][43][44]. The ability of CIs to induce a severe local inflammatory response by synthesis and release of prostaglandins, and hydrolases from phagocytes, may also be an important pathogenic mechanism in SA [43].…”

“…In contrast to other authors [46,47], who did not find any significant results in a measurement of NO, Ziora et al [48] and Moodley et al [49] detected an increased concentration of exhaled nitric oxide in patients with SA in comparison to healthy individuals. It is possible that Mtb-hsps, especially Mtb-hsp16, depending on the expression and/or level of iNOS, NOx, and antioxidants (our study is in process), may develop active or latent TB, or sarcoidosis in individuals with different genetic predisposition [3,7,9,10,13,[14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32]35].…”

“…Due to similar clinical, radiological, and histopathological pictures of SA and tuberculosis (TB), mycobacterial antigens, e.g., the 6 kDa early secretory antigenic target of Mycobacterium tuberculosis (ESAT6), catalase-peroxidase (mKatG), superoxide dismutase A (SODA), and heat shock proteins (Mtb-hsp) have been considered as infectious factors in the etiopathogenesis of SA [1,2,[4][5][6][7]. Mycobacterial hsp70, hsp65, and hsp16 may provide a link between the latent infection and autoimmunity [3,[8][9][10][11][12][13][14].…”

“…An active tuberculosis develops only in 5-10% of individuals infected with M. tuberculosis. Therefore we hypothesized that in genetically different predisposed hosts, Mtb-hsp16 may induce LTBI or active TB or autoimmune response in SA [3,[7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26]. To our knowledge, there are no studies on the expression of Mtb-hsp in precipitated CIs, or on the serum levels of nitric oxide stable metabolites in patients with sarcoidosis and tuberculosis in the context of an induction of the persistent state of mycobacteria in sarcoidosis.…”