Rate-controlling enzymes

Decomposition of the complex to the product and free enzyme is assumed irreversible, and rate controlling ... 

The polyelectrolyte covalently functionalized with reactive groups may be viewed as an enzyme-like functional polymer or as a molecular reaction system in the sense that it has both reactive centers and reaction rate-controlling microenvironments bound together on the same macromolecule. 

While metabolic engineers traditionally sought the rate-limiting enzyme to unlock flow through a pathway, now they understand that there may be many points of control and feedback with the metabolic network, and seek to empirically determine the dynamics of the interactions between rate controllers and other factors. For example, the sizes of metabolic precursor pools and the catabolism or sequestration of products affect accumulation as well as flux through the pathway. 

Pyruvate kinase (PK) is one of the three postulated rate-controlling enzymes of glycolysis. The high-energy phosphate of phosphoenolpyruvate is transferred to ADP by this enzyme, which requires for its activity both monovalent and divalent cations. Enolpyruvate formed in this reaction is converted spontaneously to the keto form of pyruvate with the synthesis of one ATP molecule. PK has four isozymes in mammals M, M2, L, and R. The M2 type, which is considered to be the prototype, is the only form detected in early fetal tissues and is expressed in many adult tissues. This form is progressively replaced by the M( type in the skeletal muscle, heart, and brain by the L type in the liver and by the R type in red blood cells during development or differentiation (M26). The M, and M2 isozymes display Michaelis-Menten kinetics with respect to phosphoenolpyruvate. The Mj isozyme is not affected by fructose-1,6-diphosphate (F-1,6-DP) and the M2 is al-losterically activated by this compound. Type L and R exhibit cooperatively in... 

The first step is catalysed by the tetrahydrobiopterin-dependent enzyme tyrosine hydroxylase (tyrosine 3-monooxygenase), which is regulated by end-product feedback is the rate controlling step in this pathway. A second hydroxylation reaction, that of dopamine to noradrenaline (norepinephrine) (dopamine [3 oxygenase) requires ascorbate (vitamin C). The final reaction is the conversion of noradrenaline (norepinephrine) to adrenaline (epinephrine). This is a methylation step catalysed by phenylethanolamine-jV-methyl transferase (PNMT) in which S-adenosylmethionine (SAM) acts as the methyl group donor. Contrast this with catechol-O-methyl transferase (COMT) which takes part in catecholamine degradation (Section 4.6). 

The simplest possible enzymatic reaction scheme was proposed in 1913 by Michealis and Menten. They assumed the molecule undergoing reaction (the substrate, S) is adsorbed reversibly on a specific site of the enzyme E to form a complex ES whose decomposition into product P is rate controlling. The scheme resembles that for unimolecular decomposition (see Chapter 14). 

The major routes for the synthesis and metabolism of noradrenaline in adrenergic nerves [375], together with the names of the enzymes concerned, are shown in Figure 3.1. Under normal conditions the rate controlling step in noradrenaline synthesis is the first, and the tissue noradrenaline content can be markedly lowered by inhibition of tyrosine hydroxylase [376]. Tissue noradrenaline levels can also be lowered, but to a lesser extent, by inhibition of dopamine-(3-oxidase [377, 378]. However, the noradrenaline depletion produced by guanethidine is unlikely to result from inhibition of synthesis, since intra-cisternal injection of guanethidine does not prevent the accumulation of noradrenaline which follows brain monoamine oxidase inhibition, even though it does cause depletion of brain noradrenaline [323]. 

In mechanistic studies, this form of equation appears whenever the rate-controlling step of a reaction is viewed to involve the association of reactant with some quantity that is present in limited but fixed amounts for example, the association of reactant with enzyme to form a complex, or the association of gaseous reactant with an active site on the catalyst surface. 

Possibly a signaling molecule which can help fight weight gain utilized by the key enzymes that control fat oxidation, at extraordinarily high rates... 

When H2O deacetylates the acyl-enzyme, phenylacetic acid is formed. When nucleophiles other than H2O deacylate the acyl-enzyme, a new condensation product, in this case phenylacetyl-O-R or phenylacetyl-NH-R is formed. By definition the hydrolysis of these condensation products can be catalyzed by the same enzyme that catalyzes their formation in equation 10.1. Thus, when the acyl-enzyme is formed from phenylacetyl-glycine or phenylacetyl-O-Me, this gives rise to an alternative process to produce Penicillin G, in addition to the thermodynamically controlled (= equilibrium controlled) condensation of phenylacetic acid and 6-aminopenicillanic acid (6-APA). This reaction that involves an activated side chain is a kinetically controlled (= rate controlled) process where the hydrolase acts as a transferase (Kasche, 1986 1989). 

At one extreme diffusivity may be so low that chemical reaction takes place only at suface active sites. In that case p is equal to the fraction of active sites on the surface of the catalyst. Such a polymer-supported phase transfer catalyst would have extremely low activity. At the other extreme when diffusion is much faster than chemical reaction p = 1. In that case the observed reaction rate equals the intrinsic reaction rate. Between the extremes a combination of intraparticle diffusion rates and intrinsic rates controls the observed reaction rates as shown in Fig. 2, which profiles the reactant concentration as a function of distance from the center of a spherical catalyst particle located at the right axis, When both diffusion and intrinsic reactivity control overall reaction rates, there is a gradient of reactant concentration from CAS at the surface, to a lower concentration at the center of the particle. The reactant is consumed as it diffuses into the particle. With diffusional limitations the active sites nearest the surface have the highest turnover numbers. The overall process of simultaneous diffusion and chemical reaction in a spherical particle has been described mathematically for the cases of ion exchange catalysis,63 65) and catalysis by enzymes immobilized in gels 66-67). Many experimental parameters influence the balance between intraparticle diffusional and intrinsic reactivity control of reaction rates with polymer-supported phase transfer catalysts, as shown in Fig. 1. 

Angiotensin-I converting enzyme (ACE) controls blood pressure by catalyzing the hydrolysis of two amino acids (His-Leu) at the C-terminus of angiotensin-I to produce a vasoconstrictor, angiotensin-II. The enzyme can also hydrolyze a synthetic substrate, hippuryl-L-histidyl-L-leucine (HHL), to hippuric acid (HA). At four different concentrations of HHL solutions (pH 8.3), the initial rates of HA formation (pmolmin" ) are obtained as shown in Table P3.8. Several small peptides (e.g., Ile-Lys-Tyr) can reversibly inhibit the ACE activity. The reaction rates of HA formation in the presence of 1.5 and 2.5 pmoll of an inhibitory peptide (Ile-Lys-Tyr) are also given in the table. 

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) ... 

As indicated in Fig. 28-11, the chromosome can be divided into four major operons, the short one that produces repressor, and the early left, early right, and late operons. The early operons code largely for replication and recombination enzymes and control proteins. The late operon is concerned with production of proteins needed for assembly of the virus particles and must be transcribed at an even higher rate hence the need for the product of gene Q. Within the late operon, genes A to F are involved in packaging of K DNA and in formation of heads, while genes S to / are... 

The role of the side chain in both catalytic and noncatalytic (75) enzyme-substrate interactions has been investigated in considerable detail (reviewed in reference 2), and a mechanism has been suggested which may explain the rate-controlling effect of side chains on the catalytic activity of / -lactamase (see Section IV,B). 

The rate of cholesterol biosynthesis appears to be regulated primarily by the activity of HMG-CoA reductase. This key enzyme is controlled by the rate of enzyme synthesis and degradation and by phosphorylation-dephosphorylation reactions. Synthesis of the mRNA for the reductase is inhibited by cholesterol delivered to cells by means of low-density lipoproteins (LDLs). 

Figure A7.1 The effect of an enzyme on the minimum energy pathway of a simple enzyme catalysed reaction involving a single substrate. (TS = transition state.) The heights of the energy barriers TSi, ts2 and ts3 will vary depending on which step in the enzyme controlled route is the rate controlling step...

Factors affecting the rate of synthesis include the level of induction or repression of the gene encoding the enzyme (see Topics G3 and G4 and also the rate of degradation of the mRNA produced from that gene. Many key enzymes at control points in metabolic pathways have particularly short-lived mRNAs and the rate of enzyme synthesis is thus readily controlled by factors that affect the rate of gene transcription. 

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