Enzyme kinetic control

Hence, sometimes phenomena associated with enzyme kinetics control the rate of biotransformations. If suitable enzymes are present in the microbial community, for example due to consumption of structurally related growth substrates, then we may see immediate degradation of compounds of interest like BQ when they are added to these metabolically competent microbial communities (Fig. 17.17). For such cases, if the abundance of the bacteria is varied, the rate of removal changes accordingly. Consequently, the removal of BQ could be described by a second-order rate law (Smith et al., 1978) ... 

M. Baldo, A. Grass and A. Raudino, Effect of interconversion between reactant configurational states of enzyme kinetics controlled by roto-translational diffusion motions, J. Chem. Phys., 93 (1990) 6034-6040. 

Kinases (Section 28 3) Enzymes that catalyze the transfer of phosphate from ATP to some other molecule Kinetically controlled reaction (Section 10 10) Reaction in which the major product is the one that is formed at the fastest rate... 

This system displays a two-enzyme kinetic model in which bioconversion is controlled by the interaction between the two reactions and the mass transfer. This situation offers a more realistic model for the conditions occurring in vivo, in which some pathways of intermediary metabolism consist of linear sequences of reactions. These pathways take place in highly organized compartments. 

P. J. Mulquiney and P. W. Kuchel, Model of 2,3 bisphosphoglycerate metabolism in the human erythrocyte based on detailed enzyme kinetic equations Computer simulation and metabolic control analysis. Biochem. J. 342 (3), 597 604 (1999). 

The various chemical mechanisms of enzyme action will not be discussed here but an overview of enzyme kinetics is essential to allow a full understanding of metabolic control. Enzymes accelerate biochemical reactions. The precise rate of reaction is influenced by a number of physiological (cellular) factors ... 

In a further study, Taniuchi et al. (1977) have shown that in the association of overlapping fragments of staphylococcal nuclease, two different species of active enzyme are formed. On the basis of the products of limited proteolysis, structures for the two species were deduced. In one case a structure is proposed in which fragment 1-126 assumes native-like structure over the sequence 1-48, and all of fragment 50-149 assumes native-like structure. In the other case the structure is one in which fragment 1-126 assumes native-like structure over the sequence 1-110, while that part of fragment 50-149 in the sequence interval 111-149 assumes native-like structure. The interest of these results is enhanced by the finding that the two active species initially form in relative concentrations substantially different from their equilibrium concentrations. Thus, both a mobile equilibrium and substantial kinetic control of the early products are evident. Taniuchi et al. did not reach a clear-cut mechanistic conclusion from their studies. 

A practical enzymatic procedure using alcalase as biocatalyst has been developed for the synthesis of hydrophilic peptides.Alcalase is an industrial alkaline protease from Bacillus licheniformis produced by Novozymes that has been used as a detergent and for silk degumming. The major enzyme component of alcalase is the serine protease subtilisin Carlsberg, which is one of the fully characterized bacterial proteases. Alcalase has better stability and activity in polar organic solvents, such as alcohols, acetonitrile, dimethylformamide, etc., than other proteases. In addition, alcalase has wide specificity and both l- and o-amino acids that are accepted as nucleophiles at the p-1 subsite. Therefore, alcalase is a suitable biocatalyst to catalyse peptide bond formation in organic solvents under kinetic control without any racemization of the amino acids (Scheme 5.1). 

Equations 2.26 and 2.27 carmot be solved analytically except for a series of limiting cases considered by Bartlett and Pratt [147,192]. Since fine control of film thickness and organization can be achieved with LbL self-assembled enzyme polyelectrolyte multilayers, these different cases of the kinetic case-diagram for amperometric enzyme electrodes could be tested [147]. For the enzyme multilayer with entrapped mediator in the mediator-limited kinetics (enzyme-mediator reaction rate-determining step), two kinetic cases deserve consideration in this system in both cases I and II, there is no substrate dependence since the kinetics are mediator limited and the current is potential dependent, since the mediator concentration is potential dependent. Since diffusion is fast as compared to enzyme kinetics, mediator and substrate are both approximately at their bulk concentrations throughout the film in case I. The current is first order in both mediator and enzyme concentration and k, the enzyme reoxidation rate. It increases linearly with film thickness since there is no... 

As discussed above, proteases are peptide bond hydrolases and act as catalysts in this reaction. Consequently, as catalysts they also have the potential to catalyze the reverse reaction, the formation of a peptide bond. Peptide synthesis with proteases can occur via one of two routes either in an equilibrium controlled or a kinetically controlled manner 60). In the kinetically controlled process, the enzyme acts as a transferase. The protease catalyzes the transfer of an acyl group to a nucleophile. This requires an activated substrate preferably in the form of an ester and a protected P carboxyl group. This process occurs through an acyl covalent intermediate. Hence, for kineticmly controlled reactions the eii me must go through an acyl intermediate in its mechanism and thus only serine and cysteine proteases are of use. In equilibrium controlled synthesis, the enzyme serves omy to expedite the rate at which the equilibrium is reached, however, the position of the equilibrium is unaffected by the protease. 

For example, Bachelard used [Mgtotai]/[ATPtotai ] = 1 in his rate studies, and he obtained a slightly sigmoidal plot of initial velocity versus substrate ATP concentration. This culminated in the erroneous proposal that brain hexokinase was allosterically activated by magnesium ions and by magnesium ion-adenosine triphosphate complex. Purich and Fromm demonstrated that failure to achieve adequate experimental control over the free magnesium ion concentration can wreak havoc on the examination of enzyme kinetic behavior. Indeed, these investigators were able to account fully for the effects obtained in the previous hexokinase study. ... 

In principle, there are two possible ways to measure this effect. First, there is the end-point measurement (steady-state mode), where the difference is calculated between the initial current of the endogenous respiration and the resulting current of the altered respiration, which is influenced by the tested substances. Second, by kinetic measurement the decrease or the acceleration, respectively, of the respiration with time is calculated from the first derivative of the currenttime curve. The first procedure has been most frequently used in microbial sensors. These biosensors with a relatively high concentration of biomass have a longer response time than that of enzyme sensors. Response times of comparable magnitude to those of enzyme sensors are reached only with kinetically controlled sensors. 

Is it the case that all components of our cellular life are thermodynamically stable Of course not. We have around us many compounds in our biochemistry that are not under thermodynamic control - think of important compounds such as adenonine triphosphate (ATP), phospholipids, RNA, DNA, proteins. .. Nowadays, these compounds are formed thanks to the action of enzymes, which are often specialized for catalyzing the synthesis of products under kinetic control. 

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