The membrane-bound hydrogenase of Alcaligenes eutrophus: II. Localization and immunological comparison with other hydrogenase systems

1980 ◽  
Vol 46 (1) ◽  
pp. 1-14 ◽  
Author(s):  
B. Schink ◽  
H. G. Schlegel
Keyword(s):  
Alcaligenes Eutrophus ◽  
Immunological Comparison ◽  
Membrane Bound

scholarly journals Functional and Structural Role of the Cytochrome b Subunit of the Membrane-Bound Hydrogenase Complex of Alcaligenes Eutrophus H16

1997 ◽  
Vol 248 (1) ◽  
pp. 179-186 ◽  
Author(s):  
Michael Bernhard ◽  
Bruna Benelli ◽  
Alejandro Hochkoeppler ◽  
Davide Zannoni ◽  
Barbel Friedrich
Keyword(s):  
Cytochrome B ◽  
Alcaligenes Eutrophus ◽  
Structural Role ◽  
B Subunit ◽  
Alcaligenes Eutrophus H16 ◽  
Membrane Bound

scholarly journals Antigenic determinants of the membrane-bound hydrogenase in Alcaligenes eutrophus are exposed toward the periplasm.

1995 ◽  
Vol 177 (21) ◽  
pp. 6309-6312 ◽  
Author(s):  
K Eismann ◽  
K Mlejnek ◽  
D Zipprich ◽  
M Hoppert ◽  
H Gerberding ◽  
...  
Keyword(s):  
Alcaligenes Eutrophus ◽  
Antigenic Determinants ◽  
Membrane Bound

scholarly journals Purification of hydrogenases by affinity chromatography on Procion Red-agarose

1983 ◽  
Vol 213 (2) ◽  
pp. 391-398 ◽  
Author(s):  
K Schneider ◽  
M Pinkwart ◽  
K Jochim
Keyword(s):  
Enzyme Preparation ◽  
Methyl Viologen ◽  
Specific Activity ◽  
Specific Interaction ◽  
Potassium Phosphate ◽  
Kinetic Studies ◽  
Fold Increase ◽  
Alcaligenes Eutrophus ◽  
Membrane Bound ◽  
Triazine Dye

The agarose-coupled triazine dye Procion Red HE-3B has been demonstrated to be applicable as an affinity gel for the purification of five diverse hydrogenases, namely the soluble, NAD-specific and the membrane-bound hydrogenase of Alcaligenes eutrophus, the membrane-bound hydrogenase of the N2-fixing Alcaligenes latus, the reversible H2-evolving and the unidirectional H2-oxidizing hydrogenase of Clostridium pasteurianum. In the case of the soluble hydrogenase of A. eutrophus, chromatography on Procion Red-agarose even permitted the separation of inactive from active enzyme, thus yielding a 2-3-fold increase in specific activity. For the homogeneous enzyme preparation obtained after two column steps (Procion Red-agarose, DEAE-Sephacel), a specific activity of 121 mumol of H2 oxidized/min per mg of protein was determined. Kinetic studies with free Procion Red provided evidence that the diverse hydrogenases are competitively inhibited by the dye, each with respect to the electron carrier (NAD, Methylene Blue, Methyl Viologen), indicating a specific interaction between Procion Red and the catalytic centres of the enzymes. For the highly purified preparations of the soluble and the membrane-bound hydrogenase of A. eutrophus, in 50 mM-potassium phosphate, pH 7.0, Ki values for Procion Red of 103 and 19 microM have been determined.

scholarly journals The properties of adenosine triphosphatase from exponential and synchronous cultures of Alcaligenes eutrophus H16

1978 ◽  
Vol 172 (2) ◽  
pp. 253-260 ◽  
Author(s):  
Clive Edwards ◽  
Juliet A. Spode ◽  
Colin W. Jones
Keyword(s):  
Cell Cycle ◽  
Adenosine Triphosphatase ◽  
Alcaligenes Eutrophus ◽  
Chain Structure ◽  
Whole Cells ◽  
Synchronous Cultures ◽  
Maximum Inhibition ◽  
A Cell ◽  
Membrane Bound ◽  
Endogenous Substrates

The properties of Alcaligenes eutrophus ATPase (adenosine triphosphatase) were investigated by using subcellular fractions prepared from cells growing in exponential and synchronous cultures. Both the soluble and membrane-bound forms of the ATPase were inhibited non-competitively (Ki 142μm) by Nbf-Cl (4-chloro-7-nitrobenzofurazan), whereas only the membrane-bound enzyme was inhibited (non-competitive; Ki 750μm) by NN′-dicyclohexylcarbodi-imide. Neither the activity of the ATPase nor its sensitivity to these two inhibitors varied during exponential growth. However, marked variations in ATPase activity were observed during synchronous growth, which were characterized by maxima at approx. 0.4 and 0.9 of a cell cycle and minima at approx. 0.1 and 0.6 of a cycle. Sensitivity to Nbf-Cl and NN′-dicyclohexylcarbodi-imide also varied during the cell cycle; maximum inhibition by the former occurred at approx. 0.4 and 0.9 of a cell cycle, whereas maximum inhibition by the latter was located at approx. 0.1 and 0.6 of a cell cycle. Proton conductance by whole cells was also periodic during the cell cycle, the lowest rates occurring at approx. 0.15 and 0.55 of a cycle and the highest rates at approx. 0.4 and 0.9 of a cycle, but →H+/O quotients for the oxidation of endogenous substrates remained relatively constant and indicated the presence of four proton-translocating respiratory segments throughout the cell cycle. These results are discussed in terms of ATPase and respiratory-chain structure and function during the cell cycle of Alcaligenes eutrophus.

scholarly journals The napEDABC gene cluster encoding the periplasmic nitrate reductase system of Thiosphaera pantotropha

1995 ◽  
Vol 309 (3) ◽  
pp. 983-992 ◽  
Author(s):  
B C Berks ◽  
D J Richardson ◽  
A Reilly ◽  
A C Willis ◽  
S J Ferguson
Keyword(s):  
Nitrate Reductase ◽  
Sequence Similarity ◽  
Alcaligenes Eutrophus ◽  
Integral Membrane Protein ◽  
Periplasmic Nitrate Reductase ◽  
Thiosphaera Pantotropha ◽  
Cysteine Motif ◽  
Membrane Bound ◽  
K 12 ◽  
Nitrate Reductases

The napEDABC locus coding for the periplasmic nitrate reductase of Thiosphaera pantotropha has been cloned and sequenced. The large and small subunits of the enzyme are coded by napA and napB. The sequence of NapA indicates that this protein binds the GMP-conjugated form of the molybdopterin cofactor. Cysteine-181 is proposed to ligate the molybdenum atom. It is inferred that the active site of the periplasmic nitrate reductase is structurally related to those of the molybdenum-dependent formate dehydrogenases and bacterial assimilatory nitrate reductases, but is distinct from that of the membrane-bound respiratory nitrate reductases. A four-cysteine motif at the N-terminus of NapA binds a [4Fe-4S] cluster. The DNA- and protein-derived primary sequence of NapB confirm that this protein is a dihaem c-type cytochrome and, together with spectroscopic data, indicate that both NapB haems have bis-histidine ligation. napC is predicted to code for a membrane-anchored tetrahaem c-type cytochrome that shows sequence similarity to the NirT cytochrome c family. NapC may be the direct electron donor to the NapAB complex. napD is predicted to encode a soluble cytoplasmic protein and napE a monotopic integral membrane protein, napDABC genes can be discerned at the aeg-46.5 locus of Escherichia coli K-12, suggesting that this operon encodes a periplasmic nitrate reductase system, while napD and napC are identified adjacent to the napAB genes of Alcaligenes eutrophus H16.

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