Biomedical subjects

M Czjzek

Publications and source records attributed to M Czjzek.

25 records · Page 2Linked to original sources

PubMed1995

Crystal structure of the catalytic domain of a bacterial cellulase belonging to family 5.

BACKGROUND: Cellulases are glycosyl hydrolases--enzymes that hydrolyze glycosidic bonds. They have been widely studied using biochemical and microbiological techniques and have attracted industrial interest because of their potential in biomass conversion and in the paper and textile industries. Glycosyl hydrolases have lately been assigned to specific families on the basis of similarities in their amino acid sequences. The cellulase endoglucanase A produced by Clostridium cellulolyticum (CelCCA) belongs to family 5. RESULTS: We have determined the crystal structure of the catalytic domain of CelCCA at a resolution of 2.4 A and refined it to 1.6 A. The structure was solved by the multiple isomorphous replacement method. The overall structural fold, (alpha/beta)8, belongs to the TIM barrel motif superfamily. The catalytic centre is located at the C-terminal ends of the beta strands; the aromatic residues, forming the substrate-binding site, are arranged along a long cleft on the surface of the globular enzyme. CONCLUSIONS: Strictly conserved residues within family 5 are described with respect to their catalytic function. The proton donor, Glu170, and the nucleophile, Glu307, are localized on beta strands IV and VII, respectively, and are separated by 5.5 A, as expected for enzymes which retain the configuration of the substrate's anomeric carbon. Structure determination of the catalytic domain of CelCCA allows a comparison with related enzymes belonging to glycosyl hydrolase families 2, 10 and 17, which also display an (alpha/beta)8 fold.

Binding Sites↗

PubMed1995

Interfacial properties of the polyheme cytochrome c3 superfamily from Desulfovibrio.

In order to compare the interfacial behavior of the polyheme cytochromes c which belong to the cytochrome c3 superfamily, the monomolecular film technique was used to determine whether and how these metalloproteins interact with (phospho)lipids). Measurements of the variations of surface pressure and surface potential versus time have shown differences in their penetration capacity into phosphatidylcholine, dicaprin, and phosphatidylglycerol films. The Desulfovibrio vulgaris Hildenborough cytochrome with 16 hemes (Hmc) and Desulfovibrio desulfuricans Norway tetra- and octaheme cytochromes c3, which have been assumed to be soluble periplasmic molecules, may be considered as extrinsic membrane proteins, unlike the D. vulgaris Hildenborough cytochrome c3 (Mr 13 000). The interfacial properties are discussed in terms of the available three-dimensional structural data, the electrostatic potential calculation, and the results obtained by hydrophobic cluster analysis of the cytochrome sequences. The very different behavior of the two cytochromes c3 (Mr 13 000) enlightens the role of a particular surface loop in the interaction with a model membrane. A functional interpretation is proposed assuming that the D. vulgaris Hildenborough Hmc and both cytochromes c3 (Mr 13 000) and (Mr 26 000) from the Norway strain might provide the link between periplasmic hydrogen oxidation and cytoplasmic sulfate reduction.

Amino Acid Sequence↗

PubMed1994

Crystal structure of cytochrome c3 from Desulfovibrio desulfuricans Norway at 1.7 A resolution.

The crystal structure of cytochrome c3 (M(r) 13,000) from Desulfovibrio desulfuricans (118 residues, four heme groups) has been crystallographically refined to 1.7 A resolution using a simulated annealing method, based on the structure-model at 2.5 A resolution, already published. The final R-factor for 10,549 reflections was 0.198 covering the range from 5.5 to 1.7 A resolution. The individual temperature factors were refined for a total of 1059 protein atoms, together with 126 bound solvent molecules. The structure has been analyzed with respect to its detailed conformational properties, secondary structure features, temperature factor behaviour, bound solvent sites and heme geometry and ligation. The characteristic secondary structures of the polypeptide chain of this molecule are one extended alpha-helix, a short beta-strand and 13 reverse turns. The four heme groups are located in different structural environments, all highly exposed to solvent. The particular structural features of the heme environments are compared to the four hemes of the cytochrome c3 from Desulfovibrio vulgaris Miyazaki.

Amino Acid Sequence↗

PubMed1994

Molecular and structural basis of electron transfer in tetra- and octa-heme cytochromes.

The first three-dimensional structure of a dimeric, octa-heme cytochrome c3 (M(r) 26000) from Desulfovibrio desulfuricans Norway, established at 2.2 A resolution, is briefly presented and compared to the known 3-D-structures of different C3-type tetraheme cytochromes, in order to contribute to a better understanding of the function of multiheme clusters and of the role of conserved amino acids implicated in possible electron transfer pathways. The dimeric protein crystallizes in the space group P3(1)21 with a = 73.01 A, c = 61.81 A and the asymmetric unit contains one monomer subunit, the dimer being generated by the crystallographic two-fold axis. The 3-D-structure was solved using the molecular replacement method with a model based on the structure of the tetraheme cytochrome c3 (M(r) 13000) from D desulfuricans Norway, presently refined at 1.7 A resolution. The monomeric subunit has the same overall fold as all cytochromes c3 (M(r) 13000). Moreover, the heme core of all examined cytochromes c3 is highly conserved, but differences appear concerning the heme environments and the histidines, axial ligands of the heme-iron atoms.

Amino Acid Sequence↗

PubMed1993

Crystallization and preliminary X-ray analysis of the catalytic domain of endoglucanase from Clostridium cellulolyticum.

The catalytic domain of an endoglucanase belonging to family A (CelCCA) from an anaerobic bacterium (Clostridium cellulolyticum) has been crystallized in a form suitable for X-ray diffraction analysis. The crystals have been grown in the presence of polyethylene glycol 4000 using the vapour diffusion technique. The crystals, which diffract to 2.0 A resolution, belong to the orthorhombic space group P2(1)2(1)2(1) and have the following cell constants: a = 52.4 A, b = 76.2 A and c = 113.5 A.

Amino Acid Sequence↗

PubMed1992

Crystallization and preliminary crystallographic study of an octa-heme cytochrome c3 from Desulfovibrio desulfuricans Norway.

An octa-heme cytochrome c3, isolated as a dimeric molecule of about 30 kDa from the anaerobic bacteria Desulfovibro desulfuricans Norway, has been crystallized in a form suitable for atomic resolution X-ray structural investigations. The crystals are trigonal, space group P3(1)21 (or its enantiomorph P3(2)21), with cell dimensions: a = b = 72.9 A c = 62.7 A. The asymmetric unit contains most probably one monomer and a solvent content of about 60%. Under this assumption, the crystallographic 2-fold axis relates the two subunits of the dimer. Diffraction extends to 2.0 A.

Crystallization↗

PubMed1992

Site-directed mutagenesis of tetraheme cytochrome c3. Modification of oxidoreduction potentials after heme axial ligand replacement.

The nature of the axial ligands of a heme group is an important factor in maintaining the oxidation-reduction potential of a c-type cytochrome. Cytochrome c3 from Desulfovibrio vulgaris Hildenborough contains four bis-histidinyl coordinated hemes with low oxidation-reduction potentials. Site-directed mutagenesis was used to generate a mutant in which histidine 70, the sixth axial ligand of heme 4, has been replaced by a methionine. The mutant protein was expressed in Desulfovibrio desulfuricans G200 at a level similar to the wild type cytochrome. A model for the three-dimensional structure of D. vulgaris Hildenborough cytochrome c3 was generated on the basis of the crystal structure of D. vulgaris Miyazaki cytochrome c3 in order to investigate the effects of the H70M mutation. The model, together with NMR data, suggested that methionine 70 has effectively replaced histidine 70 as the sixth axial ligand of heme 4 without significant alteration of the structure. A large increase of at least 200 mV of one of the four oxidation-reduction potentials was observed by electrochemistry and is interpreted in terms of structure/potential relationships.

Amino Acid Sequence↗