NADPH-Independent Reactions

P450 catalyzes the isomerization of a variety of fatty acid hydroperoxides, including 15-hydro-peroxyeicosatetraenoic acid (15-HPETE) and of the prostaglandin H2 (PGHj) endoperox-ide. A distinctive feature of some P450 fatty acid peroxide isomerases is their inability to accept electrons from NADPH and to activate molecular 

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The light independent reactions take place in the stroma with the help of ATP and NADPH. In a process called the Calvin-Benson cycle, or carbon fixation, carbon dioxide from the atmosphere is captured and converted into carbohydrates [135]. The reaction is catalyzed by the enzyme RuBisCO (ribulose-1,5-biphosphate... 

Like the examples mentioned above, most examples of metabolic flux analysis by metabolite balancing have redox balances as a central constraint used in the determination of the flux distribution. However, the redox balance is, especially under aerobic conditions, subject to uncertainties which make it less suitable for estimation of the fluxes. Part of the reason for this is to be found in futile cycles, e. g., oxidation of sulfides to disulfides, where reductive power is needed to reduce the disulfides. The net result of this reaction is reduction of molecular oxygen to water, and oxidation of NADPH to NADP+. Since the consumption rate of oxygen of these specific reactions is impossible to measure, the result may be that the NADPH consumption is underestimated. This is in accordance with the finding that when the NADPH-producing reactions are estimated independently of the NADPH-consuming reactions, there is usually a large excess of NADPH that needs to be oxidized by reactions not included in the network, e. g., futile cycles [11-13]. 

Photosystem I and NADPH Synthesis Photophosphorylation THE LIGHT-INDEPENDENT REACTIONS The Calvin Cycle Photorespiration... 

The incorporation of COz into carbohydrate by eukaryotic photosynthesizing organisms, a process that occurs within chloroplast stroma, is often referred to as the Calvin cycle. Because the reactions of the Calvin cycle can occur without light if sufficient ATP and NADPH are supplied, they have often been called the dark reactions. The name dark reactions is somewhat misleading, however. The Calvin cycle reactions typically occur only when the plant is illuminated, because ATP and NADPH are produced by the light reactions. Therefore light-independent reactions is a more appropriate term. Because of the types of reactions that occur in the Calvin cycle, it is also referred to as the reductive pentose phosphate cycle (RPP cycle) and the photo synthetic carbon reduction cycle (PCR cycle). 

The Z scheme is a mechanism whereby electrons are transferred from water to NADP +. This process produces the reducing agent NADPH required for fixing carbon dioxide in the light-independent reactions of photosynthesis. Removal of the electrons from water also results in the production of oxygen. As electrons flow from PSII to PSI, protons are... 

With very few exceptions, life directly or indirecdy depends on photosynthesis. The electric current is the flow of electrons from water to NADP+ a light-requiring process. The current continues in the light-independent reactions, with electrons flowing from NADPH to 6 phosphoglycerate, which ultimately yields glucose. 

Peroxisomes in the yeast S. cerevisiae are the only site of P-oxidation and furthermore, this yeast species is able to degrade a range of polyunsaturated fatty acids with double bonds at both even and uneven positions which requires the participation of 2,4-dienoyl-CoA reductase, an NADPH-requiring reaction. Oxidation of polyunsaturated fatty acids thus requires continued reduction of NADP to NADPH. Independent studies by ourselves (Van Roermund etal ) and others (Henke et have identified the peroxisomal isoform of NADP-linked isocitrate dehydrogenase as an essential component of such a NADP(H) redoxshuttle which requires full elucidation in the future. 

ADP to ATP, as illustrated in Figure 5.2. The reduction of CO2 and production of sugars occurs via light-independent reactions, in the Calvin Cycle, where NADPH and ATP are finally consumed. 

The reaction centres participate in the photochemical reactions called the light reactions in which ATP, NADPH and oxygen are generated. The synthesis of carbohydrate involves a second series of light-independent reactions called the dark reactions which are catalysed by stromal and cytosolic enzymes but require the availability of ATP and NADPH. 

Carbon dioxide is fixed by a series of light-independent reactions, called the dark reactions, which utilize the NADPH and ATP generated by light-dependent electron transport within the thylakoid membranes. The enzymes of the dark reactions reside mainly within the chloroplast stroma but it should be noted that sucrose synthesis from triose phosphate is a cytosolic process. 

Nitric oxide may also be an antioxidant by virtue of the feet that it can directly inhibit NADPH oxidase and thus prevent superoxide production (Clancy etaJ., 1992). This inhibition was reported to be independent of the reaction between nitric oxide and superoxide, which might be expected to be pro-oxidant (see Section 2.2.3). 

Mannose, fructose and glucose may all be assayed independently using hexokinase and the appropriate additional enzymes. All the reactions can be monitored by the increase in absorbance at 340 nm as NADI is reduced to NADPH. 

Inhibition studies involving ALR2 have indicated noncompetitive inhibition for virtually all compounds examined to date when the forward (reduction) reaction is monitored. This mode of inhibition is often interpreted as meaning that the inhibitor binds to a site on the enzyme that is independent of the catalytic site. Kinetic and competition studies have both led to this conclusion in the case of ALR2 [24,25]. The crystal structure of the enzyme complexed with both the NADPH cofactor and zopolrestat, however, clearly shows the inhibitor occupying the region directly above the nicotinamide of the NADPH and, therefore, the active site (Figures 5, 6, and 7). 

Calculation of enzyme activity The decrease in NADPH concentration, zf(NADPH)/min is calculated from the linear slope of the graph obtained and the appropriate absorption coefficient for NADPH. The true glutathione peroxidase-dependent reaction rate is obtained by subtracting the value of the non-enzymic and hydroperoxide-independent apparent rates. Glutathione peroxidase activity is frequently given simply in terms of d(NADPH)/min under standard conditions. A calculation of an activity value, A, that is likely to be more universally applicable, is given in by Flohe and Gumzler (1984). 

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