title: FK506-binding protein of the hyperthermophilic archaeum, Thermococcus sp. KS-1, a cold-shock-inducible peptidyl-prolyl cis-trans isomerase with activities to trap and refold denatured proteins.
abstract: The FK506 (tacrolimus)-binding protein (FKBP) type peptidyl-prolyl cis-trans isomerase (PPIase) in the hyperthermophilic archaeum Thermococcus sp. KS-1 was shown to be induced by temperature downshift to growth temperatures lower than the optimum. This PPIase (TcFKBP18) showed chaperone-like protein refolding activity in addition to PPIase activity in vitro. It refolded unfolded citrate synthase (CS) and increased the yield of the refolded protein. At a molar ratio of 15:1 ([TcFKBP18] to [CS]) in the refolding mixture, the recovered yield of folded CS was maximal at 62%, whereas that of spontaneous refolding was 11%. Increasing FKBP above a 15:1 ratio decreased the final yield, whereas the aggregation of unfolded CS was suppressed. A cross-linking analysis showed the formation of a complex between TcFKBP18 and unfolded CS (1:1 complex) at molar ratios of 3:1 to 15:1. However, molar ratios of 15:1 or 60:1 induced the binding of multiple FKBP molecules to an unfolded CS molecule (multimeric complex). Disrupting hydrophobic interaction by adding ethylene glycol at a molar ratio of 60:1 ([TcFKBP18] to [CS]) suppressed the formation of this multimeric complex, simultaneously enhancing CS refolding. FK506 also suppressed the formation of the multimeric complex while increasing the chaperone-like activity. These results suggest that the hydrophobic region of TcFKBP18, probably the FK506-binding pocket, was important for the interaction with unfolded proteins. No cross-linked product was detected between TcFKBP18 and native dimeric CS. TcFKBP18 probably traps the unfolded protein, then refolds and releases it in a native form. This FKBP might be important at growth temperatures lower than the optimum in Thermococcus sp. KS-1 cells.
authors: Marine Biotechnology Institute Co. Ltd., 3-75-1 Heita, Kamaishi, Iwate 026-0001, Japan. akira.ideno@kamaishi.mbio.co.jp
authors: A
authors: A
authors: Ideno
authors: T
authors: T
authors: Yoshida
authors: T
authors: T
authors: Iida
authors: M
authors: M
authors: Furutani
authors: T
authors: T
authors: Maruyama
mesh_terms: D002950:Citrate (si)-Synthase / Q000737:chemistry / Q000378:metabolism; D003080:Cold Temperature; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic; D007700:Kinetics; D011489:Protein Denaturation; D017510:Protein Folding; D022021:Tacrolimus Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D019712:Thermococcus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology
chemical_list: D002950:Citrate (si)-Synthase; D022021:Tacrolimus Binding Proteins