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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
1
|
| pubmed:dateCreated |
1994-12-16
|
| pubmed:abstractText |
Assimilatory nitrate reductase from Chlorella vulgaris catalyzes the rate-limiting step, the conversion of nitrate to nitrite, in nitrate assimilation. Initial rate studies of nitrate reductase activity, performed under optimum conditions of constant ionic strength (mu = 0.2) and pH (8.0) and using NADH as reductant, indicated the absence of substrate inhibition at NADH concentrations below 300 microM and NO3- concentrations less than 3 mM. Chlorella nitrate reductase exhibited a marked preference for NADH (Vmax = 9.2 mumol NADH/min/nmol heme and Km = 2.3 microM) as the physiological electron donor but could also utilize alpha-NADH (Vmax = 5.6 mumol NADH/min/nmol heme and Km = 131 microM) and NADPH (Vmax = 0.6 mumol NADPH/min/nmol heme and Km = 910 microM) though with significantly decreased efficiency. Examination of various NADH-analogs indicated that reduced nicotinamide hypoxanthine dinucleotide (NHDH) was used most efficiently (Vmax = 9.3 mumol NHDH/min/nmol heme and Km = 7.9 microM), while reduced nicotinamide mononucleotide (NMNH) was utilized least efficiently (Vmax = 0.07 mumol NMNH/min/nmol heme and Km = 676 microM). Overall, modifications to the nicotinamide moiety or the addition of a phosphate group were observed to result in the most significant decreases in Vmax, indicating poor reducing substrates. Product inhibition studies indicated both NAD+ (Ki = 2.2 mM) and NADP+ (Ki = 10.5 mM) to be competitive inhibitors of Chlorella NR. A variety of NAD+ analogs were also determined to act as competitive inhibitors with varying degrees of efficiency. 3-Pyridinealdehyde adenine dinucleotide was the most efficient inhibitor (Ki = 0.74 mM) while nicotinamide was the least efficient (Ki = 18.1 mM). Overall, changing substituents on the nicotinamide ring or its complete deletion produced the most effective inhibitors compared to NAD+. In contrast, changes in the adenine or ribose moieties produced less effective inhibitors when compared to NAD+. These results represent the most comprehensive analysis of the effect of modifications of the physiological reductant (NADH) and product (NAD+) on nitrate reductase activity.
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| pubmed:grant | |
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Nov
|
| pubmed:issn |
0003-9861
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| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
15
|
| pubmed:volume |
315
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
48-53
|
| pubmed:dateRevised |
2007-11-14
|
| pubmed:meshHeading |
pubmed-meshheading:7979404-Binding, Competitive,
pubmed-meshheading:7979404-Chlorella,
pubmed-meshheading:7979404-Kinetics,
pubmed-meshheading:7979404-NAD,
pubmed-meshheading:7979404-NADP,
pubmed-meshheading:7979404-Nitrate Reductases,
pubmed-meshheading:7979404-Oxidation-Reduction,
pubmed-meshheading:7979404-Stereoisomerism,
pubmed-meshheading:7979404-Structure-Activity Relationship,
pubmed-meshheading:7979404-Substrate Specificity
|
| pubmed:year |
1994
|
| pubmed:articleTitle |
Assimilatory nitrate reductase: reduction and inhibition by NADH/NAD+ analogs.
|
| pubmed:affiliation |
Department of Biochemistry and Molecular Biology, University of South Florida, College of Medicine, Tampa.
|
| pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, U.S. Gov't, P.H.S.
|