Turnover numbers of an enzyme

The turnover number of an enzyme, is a measure of its maximal catalytic activity, is defined as the number of substrate molecules converted into product per enzyme molecule per unit time when the enzyme is saturated with substrate. The turnover number is also referred to as the molecular activity of the enzyme. For the simple Michaelis-Menten reaction (14.9) under conditions of initial velocity measurements, Provided the concentration of... 

Different enzymes exhibit different specific activities and turnover numbers. The specific activity is a measure of enzyme purity and is defined as the number of enzyme units per milligram of protein. During the purification of an enzyme, the specific activity increases, and it reaches its maximum when the enzyme is in the pure state. The turnover number of an enzyme is the maximal number of moles of substrate hydrolyzed per mole of enzyme per unit time [63], For example, carbonic anhydrase, found in red blood cells, is a very active enzyme with a turnover number of 36 X 106/min per enzyme molecule. It catalyzes a very important reaction of reversible hydration of dissolved carbon dioxide in blood to form carbonic acid [57, p. 220],... 

The turnover number of an enzyme is defined as the maximum number of moles of substrate reacted per mole of enzyme (or molecules per molecule) per minute under optimum conditions (i.e., saturating substrate concentration, optimum pH, etc). If 2 mg/cm3 of a pure enzyme (50,000 molecular weight, Michaelis constant Km = 0.03 mole/m3) catalyzes a reaction at a rate of 2.5 jumoles/nUksec when the substrate concentration is 5 x 10 3 moles/m3, determine the turnover number corresponding to this definition and the actual number of moles of substrate reacting per minute per mole of enzyme. 

Enzyme activity refers to the rate at which a particular enzyme catalyzes the conversion of a particular substrate (or substrates) to one or more products under a given set of conditions. Usually, activity refers to the contribution of many enzyme molecules (often expressed simply as activity per mg of protein or similar) but, in its simplest form, activity refers to the contribution of a single enzyme molecule. The turnover number of an enzyme-substrate combination refers to the number of substrate molecules metabolized in unit time (usually a period of 1 s) under a given set of conditions (see later). These definitions appear, at first glance, to be largely self-explanatory. However, many factors contribute to the final activity of an enzyme, and these must be considered during any assessment of such activity. 

Significance of the Turnover Number, kcat. The turnover number of an enzyme, kCM, is the maximum number of mol-... 

Answer The turnover number of an enzyme is the number of substrate molecules transformed per unit time by a single enzyme molecule (or a single catalytic site) when the enzyme is saturated with substrate ... 

The term cat can be substituted for k2 and is referred to as the turnover number of an enzyme (units of s l). The expression kcJkm is widely used as a measure of the catalytic efficiency of an enzyme and is termed the specificity constant or turnover number. Where [6] Km one can assume that all the enzyme is bound to substrate (i.e., [ 0] = [ES ). Under these conditions the maximal velocity of the reaction Fmax, is a function of... 

The maximal rate, V reveals the turnover number of an enzyme, which is the number of substrate molecules converted into product by an enzyme molecule in a unit time when the enzyme is fully saturated with substrate. It is equal to the kinetic constant k 2, which is also called k The maximal rate, V reveals the turnover number of an enzyme if the concentration of active sites [E]j is known, because... 

The maximal rate, reveals the turnover number of an enzyme,... 

This quantity is also known as the turnover number of an enzyme because it is the number of reaction processes (turnovers) that each active site catalyses per unit time. For the Michaelis-Menten mechanism it is obvious from eq. (14.12) that = However, for more complex mechanisms, may be a function of several rate constants. 

Before NMR spectroscopy and mass spectrometry revolutionized the structural elucidation of organic molecules, UV spectroscopy was an important technique and was used to identify the key chromophore of an unknown molecule. The importance of UV is much diminished nowadays, but it still retains its place in certain applications, such as the determination of kinetic parameters, (the Michaelis constant) and A cat (the turnover rate of an enzyme, in molecules per second), for a number of enzymic reactions and in the analysis of pharmaceuticals. 

Water molecules flow through an AQP-1 channel at the rate of about 109 s 1. For comparison, the highest known turnover number for an enzyme is that for catalase, 4 X 107 s-1, and many enzymes have turnover numbers between 1 s 1 and 104 s 1 (see Table 6-7). The low activation energy for passage of water through aquaporin channels (AG < 15 kJ/mol) suggests that water moves through the channels in a continuous... 

Because k2 describes the number of molecules of substrate converted to product per second per molecule of enzyme, it is called the turnover number of the enzyme. Generally, in more complex enzyme mechanisms, the expression for Kmax is complicated by k2 being replaced by an expression that is a ratio of sums of products of unitary rate constants this grouped expression is then called kcal. 

Vmax is the velocity of an enzyme-catalyzed reaction when the enzyme is saturated with all of its substrates and is equal to the product of the rate constant for the rate-limiting step of the reaction at substrate saturation (kCiU) times the total enzyme concentration, Ex, expressed as molar concentration of enzyme active sites. For the very simple enzyme reaction involving only one substrate described by Equation II-4, kCM = . Elowever, more realistic enzyme reactions involving two or more substrates, such as described by Equations II-11 and 11-12, require several elementary rate constants to describe their mechanisms. It is not usually possible to determine by steady-state kinetic analysis which elementary rate constant corresponds to kcat. Nonetheless, it is common to calculate kcat values for enzymes by dividing the experimentally determined Fmax, expressed in units of moles per liter of product formed per minute (or second), by the molar concentration of the enzyme active sites at which the maximal velocity was determined. The units of cat are reciprocal time (min -1 or sec - x) and the reciprocal of cat is the time required for one enzyme-catalyzed reaction to occur. kcat is also sometimes called the turnover number of the enzyme. 

It is possibly incorrect to consider biotin synthase an enzyme in the true sense of the word it has a turnover number of 1. It only catalyzes the synthesis of a single molecule of biotin from dethiohiotin before being inactivated. This is because the iron-sulfur cluster of the protein is the source of the sulfur that is incorporated into biotin. There is some evidence that the enzyme can be reactivated by incorporation of sulfur from cysteine, but in vitro addition of the enzymes believed to catalyze this reaction has no effect on the turnover number of the enzyme (Frey, 2001 Marquet et al., 2001). 

In noncompetitive inhibition, which also is reversible, the inhibitor and substrate can bind simultaneously to an enzyme molecule at different binding sites (see Figure 8.16). A noncompetitive inhibitor acts by decreasing the turnover number rather than by diminishing the proportion of enzyme molecules that are bound to substrate. Noncompetitive inhibition, in contrast with competitive inhibition, cannot be overcome by increasing the substrate concentration. A more complex pattern, called mixed inhibition, is produced when a single inhibitor both hinders the binding of substrate and decreases the turnover number of the enzyme. 

Detection limits in EIA are ultimately determined by how low one can measure the label s concentration via an activity assay. Sensitivity in such a kinetic determination is dependent upon the turnover number of the enzyme molecule and the method employed to detect the product of the catalyzed reaction. Purified urease obtained from Sigma Chemical Co. has considerably higher activity on a molar basis (international units per mole of enzyme) than the best available commercial preparations of some other common enzyme labels such as alkaline phosphatase, /8-galactosi-dase, peroxidase, - and glucose oxidase. This is due to the high mo-... 

When [5] the characteristic property of the turnover number for an enzyme can be invoked. This number provides information on how many times the enzyme performs its catalytic function per unit time, or how many times it forms the ES complex and is regenerated (turned over) by yielding product. The turnover number can be determined from Vmax and [Et]. We know that... 

Water molecules flow through an AQP-1 channel at the rate of about 10 s. For comparison, the highest known turnover number for an enzyme is that for catalase, 4 X 10 s, and many enzymes have turnover... 

The use of V ax in the medical literature to describe the maximal rate at which a certain amount of tissue converts substrate to product can be confusing. The best way to describe an increase in enzyme activity in a tissue is to say that the maximal capacity of the tissue has increased. In contrast, the term kcat has been developed to clearly describe the speed at which an enzyme can catalyse a reaction under conditions of saturating substrate concentration. The rate constant k a, the turnover number of the enzyme, has the units of min (micromoles of product formed per minute divided by the micromoles of active site). 

Enzyme activity refers in general to the catalytic ability of an enzyme to increase the rate of a reaction. The amazing rate (36 million molecules per minute) at which one molecule of carbonic anhydrase converts carbon dioxide to carbonic acid was mentioned earlier. This rate, called the turnover number, is one of the highest known for enzyme systems. More common turnover numbers for enzymes are closer to lOVmin, or 1000 reactions per minute. Nevertheless, even such low numbers dramatize the speed with which a small number of enzyme molecules can transform a large number of substrate molecules. i Table 10.3 gives the turnover numbers of several enzymes. 

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