title: Domain structure and denaturation of a dimeric Mip-like peptidyl-prolyl cis-trans isomerase from Escherichia coli.
abstract: FKBP22, a protein expressed by Escherichia coli, possesses PPIase (peptidyl-prolyl cis-trans isomerase) activity, binds FK506 (an immunosuppressive drug), and shares homology with Legionella Mip (a virulence factor) and its related proteins. To understand the domain structure and the folding-unfolding mechanism of Mip-like proteins, we investigated a recombinant E. coli FKBP22 (His-FKBP22) as a model protein. Limited proteolysis indicated that His-FKBP22 harbors an N-terminal domain (NTD), a C-terminal domain (CTD), and a long flexible region linking the two domains. His-FKBP22, NTD(+) (NTD with the entire flexible region), and CTD(+) (CTD with a truncated flexible region) were unfolded by a two-state mechanism in the presence of urea. Urea induced the swelling of dimeric His-FKBP22 molecules at the pretransition state but dissociated it at the early transition state. In contrast, guanidine hydrochloride (GdnCl)-induced equilibrium unfolding of His-FKBP22 or NTD(+) and CTD(+) seemed to follow three-step and two-step mechanisms, respectively. Interestingly, the intermediate formed during the unfolding of His-FKBP22 with GdnCl was not a molten globule but was thought to be composed of the partially unfolded dimeric as well as various multimeric His-FKBP22 molecules. Dimeric His-FKBP22 did not dissociate gradually with increasing concentrations of GdnCl. Very low GdnCl concentrations also had little effect on the molecular dimensions of His-FKBP22. Unfolding with either denaturant was found to be reversible, as refolding of the unfolded His-FKBP22 completely, or nearly completely, restored the structure and function of the protein. Additionally, denaturation of His-FKBP22 appeared to begin at the CTD(+).
authors.affiliation: Department of Biochemistry, Bose Institute, P-1/12, CIT Scheme VII M, Kolkata 700054, West Bengal, India.
authors.forename: Biswanath
authors.initials: B
authors.lastname: Jana
authors.forename: Amitava
authors.initials: A
authors.lastname: Bandhu
authors.forename: Rajkrishna
authors.initials: R
authors.lastname: Mondal
authors.forename: Anindya
authors.initials: A
authors.lastname: Biswas
authors.forename: Keya
authors.initials: K
authors.lastname: Sau
authors.forename: Subrata
authors.initials: S
authors.lastname: Sau
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D011489:Protein Denaturation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D058767:Protein Unfolding; D017386:Sequence Homology, Amino Acid; D016559:Tacrolimus / Q000737:chemistry; D022021:Tacrolimus Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D037521:Virulence Factors / Q000737:chemistry / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D037521:Virulence Factors; C077482:Mip-like protein, Chlamydia trachomatis; C403422:FKBP22; D022021:Tacrolimus Binding Proteins; D016559:Tacrolimus