NADH kinase

ATP NADH 2 -phosphotransferase ATP NADH2 2 -phosphotransferase DPNH kinase NADH kinase NADH2 kinase 

Additional information 2 ( 2 , amplification of NADH without the influence of coexisting NAD , by phosphorylating NADH into NADPH and putting this NAPH through an NADP -NADPH cycling system [2]) [2] 

Additional information 1 ( 1 , presence of 2 or more subunits of different 

S Additional information 1 ( 1 , no activity with dihydrostreptomycin 3 a,6-diphosphate, dihydrostreptomycin 3 a-phosphate [1]) [1] 

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S ATP + NADH <1> (<1>, the enzyme has primarily NADH kinase activity in mitochondria, mitochondrial NADPH is largely provided by the POS5 NADH kinase [2]) (Reversibility <1> [2]) [2]... 

S Additional information <1> (<1>, POS5 NADH kinase is required for mitochondrial stability with a critical role in detoxification of reactive oxygen species [1]) [1]... 

Strand, M.K. Stuart, G.R. Longley, M.J. Graziewicz, M.A. Dominick, O.G. Copeland, W.C. POS5 gene of Saccharomyces cerevisiae encodes a mitochondrial NADH kinase required for stability of mitochondrial DNA. Eukar-yot. Cell, 2, 809-820 (2003)... 

Outten, C.E. Culotta, V.C. A novel NADH kinase is the mitochondrial source of NADPH in Saccharomyces cerevisiae. EMBO J., 22, 2015-2024 (2003)... 

Ohno, T Suzuki, T. Horiuchi, T. Specific amplification of NADH using NADH kinase in a reaction mixture containing excess NAD. Biosci. Biotech-nol. Biochem., 58, 976-977 (1994)... 

Iwahashi, Y. Nakamura, T. Localization of the NADH kinase in the inner membrane of yeast mitochondria. J. Biochem., 105, 916-921 (1989)... 

The loss of NADH is followed for determination of the en2yme creatine kinase. 

Enzymes, measured in clinical laboratories, for which kits are available include y-glutamyl transferase (GGT), alanine transferase [9000-86-6] (ALT), aldolase, a-amylase [9000-90-2] aspartate aminotransferase [9000-97-9], creatine kinase and its isoenzymes, galactose-l-phosphate uridyl transferase, Hpase, malate dehydrogenase [9001 -64-3], 5 -nucleotidase, phosphohexose isomerase, and pymvate kinase [9001-59-6]. One example is the measurement of aspartate aminotransferase, where the reaction is followed by monitoring the loss of NADH ... 

Figure 2. Mechanism of PDH. The three different subunits of the PDH complex in the mitochondrial matrix (E, pyruvate decarboxylase E2, dihydrolipoamide acyltrans-ferase Ej, dihydrolipoamide dehydrogenase) catalyze the oxidative decarboxylation of pyruvate to acetyl-CoA and CO2. E, decarboxylates pyruvate and transfers the acetyl-group to lipoamide. Lipoamide is linked to the group of a lysine residue to E2 to form a flexible chain which rotates between the active sites of E, E2, and E3. E2 then transfers the acetyl-group from lipoamide to CoASH leaving the lipoamide in the reduced form. This in turn is oxidized by E3, which is an NAD-dependent (low potential) flavoprotein, completing the catalytic cycle. PDH activity is controlled in two ways by product inhibition by NADH and acetyl-CoA formed from pyruvate (or by P-oxidation), and by inactivation by phosphorylation of Ej by a specific ATP-de-pendent protein kinase associated with the complex, or activation by dephosphorylation by a specific phosphoprotein phosphatase. The phosphatase is activated by increases in the concentration of Ca in the matrix. The combination of insulin with its cell surface receptor activates PDH by activating the phosphatase by an unknown mechanism.

CK catalyzes the reversible phosphorylation of creatine in the presence of ATP and magnesium. When creatine phosphate is the substrate, the resulting creatine can be measured as the ninhydrin fluorescent compound, as in the continuous flow Auto Analyzer method. Kinetic methods based on coupled enzymatic reactions are also popular. Tanzer and Gilvarg (40) developed a kinetic method using the two exogenous enzymes pyruvate kinase and lactate dehydrogenase to measure the CK rate by following the oxidation of NADH. In this procedure the main reaction is run in a less favorable direction. 

Glutamine synthetase 6.3.1.2 C L-Glutamate NADH ADP Pyruvate Pyruvate kinase Lactate dehydrogenase... 

PDH is a multi-enzyme complex consisting of three separate enzyme units pyruvate decarboxylase, transacetylase and dihydrolipoyl dehydrogenase. Serine residues within the decarboxylase subunit are the target for a kinase which causes inhibition of the PDH the inhibition can be rescued by a phosphatase. The PDH kinase (PDH-K) is itself activated, and the phosphatase reciprocally inhibited, by NADH and acetyl-CoA. Figure 3.12(a and b) show the role and control of PDH. 

There are many examples of phosphorylation/dephosphorylation control of enzymes found in carbohydrate, fat and amino acid metabolism and most are ultimately under the control of a hormone induced second messenger usually, cytosolic cyclic AMP (cAMP). PDH is one of the relatively few mitochondrial enzymes to show covalent modification control, but PDH kinase and PDH phosphatase are controlled primarily by allosteric effects of NADH, acetyl-CoA and calcium ions rather than cAMP (see Table 6.6). 

Glycerol 3-phosphate can arise in two ways, either (i) from glycerol, via the enzyme glycerol kinase or (ii) from dihydroxyacetone phosphate, which is produced in glycolysis, by reduction with NADH, catalysed by glycerol-3-phosphate dehydrogenase ... 

Interconversion processes (see p. 120) also play an important role. They are shown here in detail using the example of the PDH complex (see p. 134). The inactivating protein kinase [la] is inhibited by the substrate pyruvate and is activated by the products acetyl-CoA and NADH+H. The protein phosphatase [Ibj—like isodtrate dehydrogenase [3] and the ODH complex [4j-is activated by Ca. This is particularly important during muscle contraction, when large amounts of ATP are needed. Insulin also activates the PDH complex (through inhibition of phosphorylation) and thereby promotes the breakdown of glucose and its conversion into fatty acids. 

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