Metabolic pathway, flux control

Hofmeyr, J.H. 1989. Control-pattern analysis of metabolic pathways. Flux and concentration control in linear pathways. Eur. J. Biochem. 186 343-354. 

In order to develop a rational approach to improving rates of metabolite production, it is necessary to consider the fate of the nutrients that are required for its synthesis. However, overcoming the major flux control points within a metabolic pathway may not lead to metabolite overproduction if the energetic consequences of the alteration are unfavourable to the organism. 

In both intermediate and maximum rates of respiration, control is distributed between several different steps, including the activity of the adenine nucleotide translocator (Groen et al., 1983). It is now recognized that the idea of a simple rate-limiting step for a metabolic pathway is simplistic and that control is shared by all steps although to different extents (Kacserand Bums, 1978 Fell, 1992). Each step in a pathway has a flux control coefficient (FCC) defined as ... 

Metabolic control analysis (MCA) assigns a flux control coefficient (FCC) to each step in the pathway and considers the sum of the coefficients. Competing pathway components may have negative FCCs. To measure FCCs, a variety of experimental techniques including radio isotopomers and pulse chase experiments are necessary in a tissue culture system. Perturbation of the system, for example, with over-expression of various genes can be applied iteratively to understand and optimize product accumulation. 

After measuring the fluxes through the metabolic network, it is necessary to determine the extent to which each pathway or enzyme controls the net fluxes. Metabolic control analysis (MCA) is a technique used to elucidate how flux control is distributed in a metabolic network, thereby providing the information for identification of potential targets for metabolic engineering [8],... 

J. L. Galazzo and J. E. Bailey, Fermentation pathway kinetics and metabolic flux control in suspended and immobilized Saccharomyces cerevisiae. Enzyme Microb. Technol. 12(3), 162 172 (1990). 

It is crucial that the flux of metabohtes through metabolic pathways be finely controlled to meet the needs of the organism at all points in time and under a variety of physiological and enviromnental conditions. Since enzymes catalyze basically aU of the reactions in metabolic pathways, it will come as no surprise to learn that control is often exerted at the level of the enzymes. There are two basic ways to do that the first way is to control the amount of an enzyme that is present, either by controlling its rate of synthesis or its rate of degradation, or both the second way is to control the activity of the enzyme. This can happen in a number of ways, frequently by interaction of an enzyme with a small molecule. 

The formalized application of metabolic control analysis deals with several parameters (a) The flux control coefficient is defined as the fractional change in pathway flux... 

FIGURE 15-34 Flux control coefficient, C, in a branched metabolic pathway. In this simple pathway, the intermediate B has two alternative fates. To the extent that reaction B —> E draws B away from the pathway A —> D, it controls that pathway, which will result in a negative flux control coefficient for the enzyme that catalyzes step B —> E. Note that the sum of all four coefficients equals 1.0, as it must. 

Holms, W.H. (1986) The central metabolic pathways of Escherichia coli relationship between flux and control at a branch point, efficiency of conversion to biomass, and excretion of acetate. Curr. Top. Cell. Regul 28, 69-106. 

Metabolic control analysis This produces a quantitative representation of the degree of flux control exercised by specific enzymes, metabolites, or effectors in the metabolic pathway. 

Fell, D. A. (1998). Increasing the flux in metabolic pathways A metabolic control analysis perspective. Biotechnol Bioeng, 58(2-3), 121-124. 

The list of examples detailed above is by no means exhaustive. Several additional examples of the structure-based control of translation, such as repressor protein binding sequences on the mRNA transcripts and ribozymes [39], are among others that could be listed. The intentions of this chapter are not to enumerate examples of how structural characteristics of mRNA influence its translation. What should be evident from the preceding discussion is that mRNA structure is a critical determinant of translation, and synthetic DNA technology offers the possibility to alter and probe the effects of mRNA sequence on its structure and the resulting translation efficiency. Further developments in this area will be immensely valuable to metabolic engineering as it will enable practitioners to fine-tune the fluxes of desired metabolic pathways through modulation of protein levels. 

In simple metabolic pathways, with only one independent flux, the value of Cj is usually between zero and 1. Partial inhibition of an enzyme with zero flux control does not affect the flux, while an enzyme with a flux control coefficient of 1 is completely rate-limiting. Flux control coefficients between 0 and 1 have often been found (Groen et al. 1982 Ruijter et al. 1991 Snoep et al. 1996). In ideal metabolic pathways, for example, in the absence of metabolite channelling and coenzyme sequestration (Kholodenko et al. 1995), the sum of the flux control coefficients of all enzymes in the pathway must equal 1 ... 

Connectivity theorems allow to relate the control coefficients (systemic properties) to the elasticity coefficients (properties of the network s enzymes individually as if in isolation) (Westerhoff and Van Dam 1987 Heinrich and Schuster 1996 Fell 1997). The connectivity theorems have given us a strong insight into the functioning of metabolic pathways. For example, it follows directly from these theorems that enzymes that are very sensitive to the concentrations of metabolites, such as substrates, products and allosteric effectors, tend to have little control over the flux. This is illustrated by overproduction of phosphofructokinase in bakers yeast, an enzyme often referred to textbooks as rate-limiting. Yet, overproduction of phosphofructokinase does not lead to a significant flux increase, since the cell compensates by lowering the level of its allosteric effector fructose 2,6-bisphosphate (Schaaff et al. 1989 Davies and Brindle 1992). 

---