Enzyme kinetics definitions

The usual starting point in enzyme kinetics is the Michaelis-Menten equation for the reaction rate v. This also seems a convenient starting point for interpretation of pressure effects on enzyme mechanisms. It will be shown that this formalism may be deceptive if the definitions and interpretations have not been made clear from the beginning. For the mechanism... 

Since Dr. Goldanskii says he is not familiar with Douzou s work, perhaps I can make some response to the question of Dr. Thomas. Douzou s observations definitely do not illustrate a Goldanskii effect. In my understanding, the essential point from Douzou experiments, focusing on enzyme kinetics at temperatures from above 0°C to substantially below, is that the kinetic parameters change in a continuous way. 

The basic principles and definitions used frequently in enzyme kinetic analysis are discussed below. 

Reductive Reactions. A number of pesticides are susceptible to reductive reactions under anaerobic conditions, depending on the substituents present on the molecule. Reductive reactions can be either chemically or enzymatically mediated. Because biologically generated reductants, eg, cysteine and porphyrins, are frequendy the electron donors for both chemical and enzymatic reactions, results from sterile controls are not necessarily conclusive in distinguishing between the two mechanisms. The only definitive means of distinguishing between chemical vs biological (enzymatic) reactions is to determine whether the reaction rate is consistent with enzyme kinetics. The most common reductive reactions are the reduction of nitro substituents and reductive dechlorination. 

In this chapter, some principles of the kinetics of enzymatic reactions are discussed. A more detailed description of enzyme kinetics is covered in a number of textbooks and articles[45, 101-1041, First of all, a few general definitions will be given. 

One convenient source of information about enzymes is the Enzyme Handbook [3] which provides for each enzyme the chemical reaction catalyzed, the kinetic constants for the enzyme, a definition of activity units, and references for the preparation and properties of the enzyme. 

Somewhat similar to enzyme kinetics, but definitely not the same, is the area of microbial kinetics. Here one is concerned with reactions between entities that may not be of the same level of organization (i.e., not atom-atom, atom-molecule, etc,). Indeed, microbial kinetics is more concerned with interactions between populations of living organisms, and some of the problems seem more akin to population dynamics than to chemical kinetics. In fact, in much of the discussion below we are concerned with population-changing processes. First, we need to define a few terms. 

The basic principles and definitions that are used frequently in enzyme kinetic analysis are also the foim-dation of chemical kinetics in physical chemistry. Three key concepts in this area are as follows. 

Many of the early studies were conducted with enzymes from fermentation, particularly invertase, which catalyzes the hydrolysis of sucrose to monosaccharides D-glucose and D-fmctose. With the introduction of the concept of hydrogen ion concentration, expressed by the logarithmic scale of pH (Sorensen, 1909), Michaelis and Menten (1913) realized the necessity for carrying out definitive experiments with invertase. They controlled the pH of the reaction medium by using acetate buffer, allowed for the mutarotation of the product and measured initial reaction rates at different substrate concentrations. Michaelis and Menten described their experiments by a simple kinetic law which afforded a foundation for a subsequent rapid development of numerous kinetic models for enzyme-catalyzed reactions. Although the contribution of previous workers, especially Henri (1902, 1903), was substantial, Michaelis and Menten are regarded as the founders of modern enzyme kinetics due to the definitive nature of their experiments and the viability of their kinetic theory. 

At this point, it is appropriate to introduce the definitions of basic statistical parameters which are extensively covered in textbooks on biochemical statistics (Mandelhall ei a/., 1981 Ratkowsky, 1983 Wiener eta/., 1991 Comish-Bowden, 1995 Daniel, 1995 Rosner, 1996 Zhr, 1996 Punch Allison, 2000) and applied here to enzyme kinetics. 

Although graphical analysis is a quick and useful way to visualize enzyme kinetic data, for any definitive work, the data must be subjected to statistical analysis so that the precision of the kinetic constants can be evaluated. However, there are good reasons why plotting methods are essential. The human eye is much less easily deceived than any computer program and is capable of detecting unexpected behavior even if nothing currently available is found in the literature. 

Graphical analysis must always precede the statistical analysis. It is imperative to keep short the time elapsed between data acquisition and data analysis, and in most cases, it is advisable to perform the graphical analysis even while the experiment is still in progress. Wien the data clearly define the nature ofthe rate or binding equation, statistical analysis is not needed to do this. Nevertheless, for a definitive work, statistical methods are necessary for parameter estimation as well as for model discrimination (Senear Bolen, 1992). Computer programs are now available for even the most sophisticated problems in enzyme kinetics (see Section 18.2.4). 

Unit definition One restriction unit is usually defined as the amount of enzyme required to digest completely 1 fig of DNA (typically phage A) in 1 hr. Because the definition relies on the activity at an end point of the reaction, it is distinguished from the usual kinetic definition of enzyme units based on the initial reaction rate. 

Viewed in this way, the best definition of rate enhancement depends upon the relationship between enzyme and substrate concentrations and the enzyme s kinetic parameters. 

Acid-catalyzed matrices, kinetics of controlled release, 170-179 Active targeting, definition, 276 Adenosine deaminase, activity of polyethylene glycol modified enzymes, 98-99 Adjuvax... 

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