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The TK2203 protein from the hyperthermophilic archaeon Thermococcus kodakarensis KOD1 (262 residues, 29 kDa) is a putative extradiol dioxygenase catalyzing the cleavage of C-C bonds in catechol derivatives. It contains three metal-binding residues, but has no significant sequence similarity to proteins for which structures have been determined. Here, the first crystal structure of the TK2203 protein was determined at 1.41 Å resolution to investigate its functional role. Structure analysis reveals that this protein shares the same fold and catalytic residues as other extradiol dioxygenases, strongly suggesting the same enzymatic activity. Furthermore, the important region contributing to substrate selectivity is discussed. [ABSTRACT FROM AUTHOR]

Coenzyme A (CoA) plays pivotal roles in a variety of metabolic pathways in all organisms. The biosynthetic pathway of CoA is strictly regulated by feedback inhibition. In the hyperthermophilic archaeon Thermococcus kodakarensis, ketopantoate reductase (KPR), which catalyzes the NAD(P)H-dependent reduction of 2-oxopantoate, is a target of feedback inhibition by CoA. The crystal structure of KPR from T. kodakarensis (Tk-KPR) complexed with CoA and 2-oxopantoate has previously been reported. The structure provided an explanation for the competitive inhibition mechanism. Here, further biochemical analyses of Tk-KPR and the crystal structure of Tk-KPR in complex with NADP+ are reported. A mutational analysis implies that the residues in the binding pocket cooperatively contribute to the recognition of CoA. The structure reveals the same dimer architecture as the Tk-KPR-CoA-2-oxopantoate complex. Moreover, the positions of the residues involved in the dimer interaction are not changed by the binding of CoA and 2-oxopantoate, suggesting individual conformational changes of Tk-KPR monomers. [ABSTRACT FROM AUTHOR]

Coenzyme A (CoA) plays pivotal roles in a variety of metabolic pathways in all organisms. The biosynthetic pathway of CoA is strictly regulated by feedback inhibition. In the hyperthermophilic archaeon Thermococcus kodakarensis, ketopantoate reductase (KPR), which catalyzes the NAD(P)H-dependent reduction of 2-oxopantoate, is a target of feedback inhibition by CoA. The crystal structure of KPR from T. kodakarensis (Tk-KPR) complexed with CoA and 2-oxopantoate has previously been reported. The structure provided an explanation for the competitive inhibition mechanism. Here, further biochemical analyses of Tk-KPR and the crystal structure of Tk-KPR in complex with NADP+ are reported. A mutational analysis implies that the residues in the binding pocket cooperatively contribute to the recognition of CoA. The structure reveals the same dimer architecture as the Tk-KPR-CoA-2-oxopantoate complex. Moreover, the positions of the residues involved in the dimer interaction are not changed by the binding of CoA and 2-oxopantoate, suggesting individual conformational changes of Tk-KPR monomers. [ABSTRACT FROM AUTHOR]

Proper protein folding is an essential process for all organisms. Prefoldin (PFD) is a molecular chaperone that assists protein folding by delivering non-native proteins to group II chaperonin. A heterohexamer of eukaryotic PFD has been shown to specifically recognize and deliver non-native actin and tubulin to chaperonin-containing TCP-1 (CCT), but the mechanism of specific recognition is still unclear. To determine its crystal structure, recombinant human PFD was reconstituted, purified and crystallized. X-ray diffraction data were collected to 4.7 Å resolution. The crystals belonged to space group P21212, with unit-cell parameters a = 123.2, b = 152.4, c = 105.9 Å. [ABSTRACT FROM AUTHOR]