Flux-generating enzymes

Figure 7.13 Physiological pathway for fatty acid oxidation. The pathway starts with the hormone-sensitive lipase in adipose tissue (the flux-generating step) and ends with the formation of acetyl-CoA in the various tissues. Acetyl-CoA is the substrate for the flux-generating enzyme, citrate synthase, for the Krebs cycle (Chapter 9). Heart, kidney and skeletal muscle are the major tissues for fatty acid oxidation but other tissues also oxidise them.

An extremely important role of iron is the synthesis of haem for formation of erythrocytes and also for proliferating cells for synthesis of the mitochondrial enzymes that contain haem (e.g. cytochromes). The flux-generating enzyme in the synthesis of haem is aminolevulinic acid synthase (ALS) (Figure 15.20). If the cellular iron concentration is low, the concentration of this enzyme is increased in an attempt to maintain the rate of synthesis. As with the other two proteins, the concentration of ALS is controlled at the level of translation in a similar manner to that for transferrin, i.e. by increased stability of the mRNA, which is achieved by the binding of the IRP to the mRNA. 

It may be identified as a nonequilibrium reaction in which the of the enzyme is considerably lower than the normal substrate concentration. The first reaction in glycolysis, catalyzed by hexokinase (Figure 17-2), is such a flux-generating step because its for glucose of 0.05 mmol/L is well below the normal blood glucose concentration of 5 mmol/L. 

One of the enzymes that catalyses a non-equilibrium reaction approaches saturation with substrate, so that it is the flux-generating step, (i.e. the beginning of the pathway). 

As indicated above, the flux-generating step for fatty acid synthesis is the conversion of acetyl-CoA to malonyl-CoA, catalysed by acetyl-CoA carboxylase. Consequently, regulation of the rate of synthesis is achieved via changes in the activity of this enzyme. The properties of the carboxylase identify three mechanisms for regulation allosteric regulation, reversible phosphorylation (an interconversion cycle) and changes in the concentration of the enzyme. (The principles underlying the first two mechanisms are discussed in Chapter 3.)... 

Figure 20.24 The physiological pathway of polypeptide synthesis. The flux-generating step is that catalysed by the aminoacyl-tRNA synthetases, indicated by the broad arrow. The assumption implicit in this interpretation is that the physiological pathway starts with the intracellular amino acids and ends with the peptide that is formed in the elongation and termination processes. For the majority of enzymes, the concentration of intracellular amino acids is higher than the K, for the synthetase (Chapter 3).

Any enzymic reaction that supplies substrate to a metabolic pathway. For all subsequent steps to maintain their steady-state concentrations, flux-generating reactions must exhibit zero-order kinetics. 

The reactions of Fig. 1 are not in contention. The enzymes and intermediates of Figs. 2 and 4 are probably present in most tissues, particularly where Aid activity is greater than that of TA. No secure evidence exists, however, that the individual reactions of either Figs. 1 or 2 are so linked that they constitute a metabolic pathway, with a flux generating step, ordered reactant, and end-product stoichiometry or fixed direction of operational flux. Nor is there a basis for belief that the PPP can operate as... 

PFK is the Main regulatory enzyme (flux generating step, first committed step) ... 

This reaction is physiologically iiTeversible and is the flux generating or first committed step of fatty acid biosynthesis. As expected it is regulated. In mammals acetyl-CoA carboxylase is a large enzyme existing as inactive protomers (560,000 MW, 4 subunits, one biotin), which can assemble into active filaments (4 -10 million MW). 

Flux-generating steps must be nonequilibrium because of the saturation with a substrate saturation with any component of a reaction (substrate, product, cofactor) places a nonequilibrium step in the enzymic mechanism, thus making the whole reaction irreversible—see Cleland (8). 

Generation of mutants is also a starting point in optimization experiments, and now is the time for metabolic engineering of the astaxanthin biosynthetic pathway. Researchers should be able to manage carbon fluxes within the cells and resolve competitions between enzymes such as phytoene desaturase and lycopene cyclase. 

It follows from the above that MPO may catalyze the formation of chlorinated products in media containing chloride ions. Recently, Hazen et al. [172] have shown that the same enzyme catalyzes lipid peroxidation and protein nitration in media containing physiologically relevant levels of nitrite ions. It was found that the interaction of activated monocytes with LDL in the presence of nitrite ions resulted in the nitration of apolipoprotein B-100 tyrosine residues and the generation of lipid peroxidation products 9-hydroxy-10,12-octadecadienoate and 9-hydroxy-10,12-octadecadienoic acid. In this case there might be two mechanisms of MPO catalytic activity. At low rates of nitric oxide flux, the process was inhibited by catalase and MPO inhibitors but not SOD, suggesting the MPO initiation. 

---