Clostridium active center

Figure 8.1 The active center of Clostridium pasteurianum hydrogenase.

The core-enzymes, prepared in our laboratory, and containing the active centers, were successfully crystallized (Dr. Jones, Uppsala, communicated) and tertiary structures will be described in the near future. Chemical modification studies on these enzymes are currently being undertaken in our laboratory identification of important catalytic residues and location of the active centers will lead to more functional information on these enzymes. Other cellulases such as some endoglucanases from Clostridium thermocel-lum (EG A, EG B, EG D) (10) and EngA and Exg from Cellulomonas fimi (19) also contain sequences of conserved, terminally located and sometimes reiterated, amino acids. Some of these sequences are preceded by proline-serine rich domains. Thus, a bistructural-bifunctional organization seems to be a rather common feature among cellulases, at least for EngA and Exg from C. fimi and the enzymes from Trichoderma reesei. 

Crystal structures of feruloyl esterases from Clostridium thermocellum XYNY and XYNZ have been published (47). The enzymes display the ajp-hydrolase fold and possess a classical Ser-His-Asp catalytic triad. The active center reveals the binding site for ferulic acid and related compounds. Ferulate... 

Many microbes contain a hydrogenase that lacks nickel. The best studied of these proteins is the hydrogenase from C. pasteurianum (Adams, 1990). The active site of this protein has been called the H cluster. Two structures of this protein recently appeared, a 1.6 structure of the het-erodimeric Desulfovibrio desuljuricans enzyme (Nicolet et al., 1999) and the 1.8 structure of the Cpl enzyme from Clostridium pasteurianum (Peters et al., 1998). The H cluster at the active site contains 6Fe as proposed earlier (Adams et al., 1989). There are two subcomponents a typical [4Fe64S] cubane bridged by a cysteine residue to an active site binuclear Fe center (Figure 4b). The unusual nature of the iron site in the NiFe hydrogenase is... 

Menon, S., and Ragsdale, S. W., 1998, Role of the [4Ee64S] cluster in reductive activation of the cobalt center of the corrinoid iron-sulfur protein from Clostridium thermoaceticum during acetyl-CoA synthesis Biochem. 37(16) 5689n5698. 

This enzyme, as well as nicotinic acid hydroxylase was recently reported by Andreesan to be a selenoenzyme. The discovery of both these enzymes was based on the clever assumption that selenium might well be a component of multisubunit enzymes containing redox centers such as iron-sulfur, flavin, molybdenum, etc. When Clostridium acidiurici was cultured in media with supplemental selenium, an elevated activity of xanthine dehydrogenase was observed. The clostridial enzyme is comparable to mammalian xanthine oxidases in that it contains flavin adeninedinucleotide (FAD), molybdenum and nonheme iron. This enzyme functions in vivo under anaerobic conditions and appears to catalyze the reduction of uric acid to xanthine. Again it will be interesting to learn the form of selenium in this enzyme. 

Ferredoxins, Fat low-molecular- mass iron-sulfur proteins which transfer electrons from one enzyme system to another, without possessing any enzyme activity themselves. The name was coined by Mortenson for iron-containing proteins from Clostridium pasteu-ranum. The 8-Fe-Fd take part in many election transport processes in organisms like Clostridia and photosynthetic bacteria (Table). The primary structures of many of these proteins have been determined. They consist of about SS amino adds, including 8 cysteines, which occupy the same positions in each Fd. The molecule contains two identical 4Fe-4S clusters, each one forming a cube and covalently bonded to 4 cysteine residues in the peptide chain. Each 4Fe-4S center can transfer one electron. 

The distal iron center is the putative site of H2 activation. The addition of CO gas to preparations of the Hox(g=2.10) form of the enzyme derived from Desul-fovibrio desuljuricans Hildenborough (DdH) or Clostridium pasteurianum I (CpI) leads to an inhibited form of the enzyme that is incapable of catalyzing reduction or H2 oxidation and the appearance of an additional v(C-O) band. The molecular structure derived from single-crystal X-ray diffraction studies of the CO-inhibited form of the [FeFe]H2ase enzyme shows that the distal iron center is coordinated by an additional CO ligand. 

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