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J R Andreesen

Publications and source records attributed to J R Andreesen.

67 records · Page 4Linked to original sources
PubMed1977

Some properties of formate dehydrogenase, accumulation and incorporation of 185W-tungsten into proteins of Clostridium formicoaceticum.

Formate dehydrogenase of Clostridium formicoaceticum used only methyl and benzyl viologen, but not NAD as electron acceptor. The S0.5 values were 0.9 X 10(-4) M for formate and 5.8 X 10(-3) M for methyl viologen. Using potassium phosphate buffer a pH-optimum of 7.9 was observed. The initial velocity of the formate dehydrogenase activity reached a maximum at 70 degrees C, whereas the activity was stable only up to 50 degrees C. The level of formate dehydrogenase in C. formicoaceticum was increased to its maximum when 10(-6) M selenite and 10(-7) M tungstate were added to a synthetic medium. Addition of molybdate instead of tungstate did not increase the level of formate dehydrogenase. 185W-tungsten was concentrated about 100-fold by C. formicoaceticum; molybdate had no major effect on the uptake of tungsten. 185W-tungsten was found almost exclusively in the soluble fluid and was predominantly recovered after chromatography in a protein of about 88000 molecular weight. Occasionally a labelled protein of low molecular weight was observed. Again molybdate added even in high molar excess did not influence the labelling pattern. No radioactivity peak could be obtained at the elution peak of formate dehydrogenase activity. The extreme instability of formate dehydrogenase prevented further purification.

Aldehyde Oxidoreductases↗
PubMed1977

Differentiation between Clostridium acidiurici and Clostridium cylindrosporum on the basis of specific metal requirements for formate dehydrogenase formation.

The formate dehydrogenases of Clostridium acidiurici and of C. cylindrosporum coupled the oxidation of formate with the reduction of viologen dyes. The basal activity level was about 0.85 mumoles/min X mg of protein for both species. The level of formate dehydrogenase of C. acidiurici increased 12-fold when 10(-7) M tungstate and selenite were present during growth. Molybdate exerted no effect. On the other hand, molybdate and selenite were required to increase the formate dehydrogenase of C. cylindrosporum, and tungstate exhibitedan antagonistic effect in this organism. Growth on hypoxanthine generally depended on the addition of bicarbonate. Supplementation with tungstate and selenite accellerated growth of C. acidiurici and increased again the level of formate dehydrogenase. The addition of both, molybdate and selenite was necessary to initiate growth of C. cyclindrosporum and to form an active formate dehydrogenase. The differences in the requirement for metal ion supplementation to form high levels of formate dehydrogenase and their involvement in hypoxanthine degradation can be used to differentiate between C. acidiurici and C. cylindrosporum.

Aldehyde Oxidoreductases↗
PubMed1976

Properties of enzymes from Clostridium thermoaceticum and Clostridium formicoaceticum.

Methylenetetrahydrofolate dehydrogenase from C. thermoaceticum and C. formicoaceticum have been purified to homogeneity and compared. The two enzymes are very similar physically, chemically, and kinetically, but he C. thermoaceticum enzyme has a higher thermostablility, which is an intrinsic property of the protein. Formate dehydrogenase enzymes from both bacteria require selenite and tungstate for formation and these enzymes also appear to have similar properties, although the C. thermoaceticum is stable at 70 degrees C for more than one hour. Acetate kinase from C. thermoaceticum appears to be under metabolic control. It can be concluded that enzymes from C. thermoaceticum, although they are more thermostable, are very similar to corresponding enzymes from mesophilic organisms.

Acetates↗
PubMed1974

Nicotinamide adenine dinucleotide phosphate-dependent formate dehydrogenase from Clostridium thermoaceticum: purification and properties.

The nicotinamide adenine dinucleotide phosphate (NADP)-dependent formate dehydrogenase in Clostridium thermoaceticum used, in addition to its natural electron acceptor, methyl and benzyl viologen. The enzyme was purified to a specific activity of 34 (micromoles per minute per milligram of protein) with NADP as electron acceptor. Disc gel electrophoresis of the purified enzyme yielded two major and two minor protein bands, and during centrifugation in sucrose gradients two components of apparent molecular weights of 270,000 and 320,000 were obtained, both having formate dehydrogenase activity. The enzyme preparation catalyzed the reduction of riboflavine 5'-phosphate flavine adenine dinucleotide and methyl viologen by using reduced NADP as a source of electrons. It also had reduced NADP oxidase activity. The enzyme was strongly inhibited by cyanide and ethylenediaminetetraacetic acid. It was also inhibited by hypophosphite, an inhibition that was reversed by formate. Sulfite inhibited the activity with NADP but not with methyl viologen as acceptor. The apparent K(m) at 55 C and pH 7.5 for formate was 2.27 x 10(-4) M with NADP and 0.83 x 10(-4) with methyl viologen as acceptor. The apparent K(m) for NADP was 1.09 x 10(-4) M and for methyl viologen was 2.35 x 10(-3) M. NADP showed substrate inhibition at 5 x 10(-3) M and higher concentrations. With NADP as electron acceptor, the enzyme had a broad pH optimum between 7 and 9.5. The apparent temperature optimum was 85 C. In the absence of substrates, the enzyme was stable at 70 C but was rapidly inactivated at temperatures above 73 C. The enzyme was very sensitive to oxygen but was stabilized by thiol-iron complexes and formate.

Aldehyde Oxidoreductases↗
PubMed1973

Fermentation of glucose, fructose, and xylose by Clostridium thermoaceticum: effect of metals on growth yield, enzymes, and the synthesis of acetate from CO 2 .

Clostridium thermoaceticum ferments xylose, fructose, and glucose with acetate as the only product. In fermentations with mixtures of the sugars, xylose is first fermented, then fructose, and last, glucose. Fructose inhibits the fermentation of glucose, and this inhibition appears to be due to a repression of the synthesis of an enzyme needed for glucose utilization. Addition of metals to the culture medium increases the cell yield drastically from about 7 to 18 g per liter, and Y(glucose) values between 40 and 50 are obtained. According to the postulated pathways of the fermentation of glucose and synthesis of acetate from CO(2) by C. thermoaceticum, 3 mol of ATP are available as energy for growth. Thus a Y(adenosine 5'-triphosphate) of 13 to 16 is obtained. Because the normal Y(ATP) value is 10.5, this could mean that an additional source of ATP is available by an unknown mechanism. The addition of metals also increases the nicotinamide adenine dinucleotide phosphate-dependent formate dehydrogenase activity, the overall reaction ((14)CO(2) --> acetate), and the incorporation of the methyl group of 5-methyltetrahydrofolate into acetate. These reactions are catalyzed very efficiently by cells harvested in early growth, whereas cells obtained at the end of a fermentation have very low formate dehydrogenase activity and capacity to incorporate CO(2) into acetate. The following enzymes involved in the synthesis of acetate from CO(2) and in the metabolism of pyruvate are present in extracts of C. thermoaceticum: 10-formyltetrahydrofolate synthetase, 5,10-methenyltetrahydrofolate cyclohydrolase, 5,10-methylenetetrahydrofolate dehydrogenase, 5,10-methylenetetrahydrofolate reductase, phosphate acetyltransferase, and acetate kinase. These enzymes are not or are very little affected by the addition of metals to the growth medium. The amount of corrinoids in cells from early growth is low, whereas it is high in cells harvested late in growth. The opposite is found for the activity of delta-aminolevulinate dehydratase, which is high at the beginning of growth and low at the end.

Acetates↗
PubMed1973

Formate dehydrogenase of Clostridium thermoaceticum: incorporation of selenium-75, and the effects of selenite, molybdate, and tungstate on the enzyme.

The formation of the nicotinamide adenine dinucleotide phosphate-dependent formate dehydrogenase in Clostridium thermoaceticum is stimulated by the presence of molybdate and selenite in the growth medium. The highest formate dehydrogenase activity was obtained with 2.5 x 10(-4) M Na(2)MoO(4) and 5 x 10(-5) Na(2)SeO(3). Tungstate but not vanadate could replace molybdate and stimulate the formation of formate dehydrogenase. Tungstate stimulated activity more than molybdate, and in combination with molybdate the stimulation of formation of formate dehydrogenase was additive. Formate dehydrogenase was isolated from cells grown in the presence of Na(2) (75)SeO(2), and a correlation was observed between bound (75)Se and enzyme activity.

Aldehyde Oxidoreductases↗
PubMedDate not supplied

Morpholine-induced formation of L-alanine dehydrogenase activity in Mycobacterium strain HE5.

An NAD-dependent, morpholine-stimulated L-alanine dehydrogenase activity was detected in crude extracts from morpholine-, pyrrolidine-, and piperidine-grown cells of Mycobacterium strain HE5. Addition of morpholine to the assay mixture resulted in an up to 4. 6-fold increase of L-alanine dehydrogenase activity when L-alanine was supplied at suboptimal concentration. L-alanine dehydrogenase was purified to near homogeneity using a four-step purification procedure. The native enzyme had a molecular mass of 160 kDa and contained one type of subunit with a molecular mass of 41 kDa, indicating a tetrameric structure. The sequence of 30 N-terminal amino acids was determined and showed a similarity of up to 81% to that of various alanine dehydrogenases. The pH optimum for the oxidative deamination of L-alanine, the only amino acid converted by the enzyme, was determined to be pH 10.1, and apparent Km values for L-alanine and NAD were 1.0 and 0.2 mM, respectively. Km values of 0. 6, 0.02, and 72 mM for pyruvate, NADH, and NH4+, respectively, were estimated at pH 8.7 for the reductive amination reaction.

Alanine Dehydrogenase↗