Enzymes molecular activity

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... 

Molecular characteristics of luciferase. A molecule of the luciferase of G. polyedra comprises three homologous domains (Li et al., 1997 Li and Hastings, 1998). The full-length luciferase (135 kDa) and each of the individual domains are most active at pH 6.3, and they show very little activity at pH 8.0. Morishita et al. (2002) prepared a recombinant Pyrocystis lunula luciferase consisting of mainly the third domain. This recombinant enzyme catalyzed the light emission of luciferin (luminescence A.max 474 nm) and the enzyme was active at pH 8.0. The recombinant enzyme of the third domain of G. polyedra luciferase was crystallized and its X-ray structure was determined (Schultz et al., 2005). A -barrel pocket putatively for substrate binding and catalysis was identified in the structure, and... 

The protein from D. desulfuricans has been characterized by Mbss-bauer and EPR spectroscopy 224). The enzyme has a molecular mass of approximately 150 kDa (three different subunits 88, 29, and 16 kDa) and contains three different types of redox-active centers four c-type hemes, nonheme iron arranged as two [4Fe-4S] centers, and a molybdopterin site (Mo-bound to two MGD). Selenium was also chemically detected. The enzyme specific activity is 78 units per mg of protein. 

The enzymes are protein molecules having globular structure, as a rule. The molecular masses of the different enzymes have values between ten thousands and hundred thousands. The enzyme s active site, which, as a rule, consists of a nonproteinic organic compound containing metal ions of variable valency (iron, copper, molybdenum, etc.) is linked to the protein globule by covalent or hydrogen bonds. The catalytic action of the enzymes is due to electron transfer from these ions to the substrate. The protein part of the enzyme secures a suitable disposition of the substrate relative to the active site and is responsible for the high selectivity of catalytic action. 

Plotting 1/V versus 1/[S], one obtains a straight line having a slope of Km/Vmax with a y-axis intercept of l/VmAX and an x-intercept of - 1/Km as shown in Figure 2.13. Lineweaver-Burk plots of enzyme activity in the presence of an inhibitor can distinguish the type of inhibitor. Competitive inhibitors have a molecular structure similar to that of the substrate and will alter Km but not VnrAX because they compete with the substrate for binding at the enzyme s active site but do not change the enzyme s affinity for substrate. Noncompetitive inhibitors bear no structural similarity to the substrate but bind the free enzyme or enzyme-substrate... 

Apart from electron promoters a large number of electron mediators have long been investigated to make redox enzymes electrochemically active on the electrode surface. In the line of this research electron mediators such as ferrocene and its derivatives have successfully been incorporated into an enzyme sensor for glucose [3]. The mediator was easily accessible to both glucose oxidase and an electron tunnelling pathway could be formed within the enzyme molecule [4]. The present authors [5,6] and Lowe and Foulds [7] used a conducting polymer as a molecular wire to connect a redox enzyme molecule to the electrode surface. 

Covalent modification of enzymes (molecular weight of several hundreds or thousands) by the incorporation of inorganic phosphate in the form of P03 (formula weight = 85), seems to represent a small chemical change in the enzyme yet is an important control mechanism of enzyme activity. Explain how phosphorylation can exert its controlling effect on the activity of the enzyme. 

Enzyme Molecular weight pi Collagenase activity M- min ) Metal content (moljmol protein ) ... 

Thermophilic enzymes are active and stable at high temperature (> 60 C) but they are generally inactive and extremely stable at low temperature (< 25 C). The molecular basis has not been elaborated to explain such thermophilicity. In general, thermophilic enzymes do not denature at high temperature and their activity is higher due to the Qio rule where a 10°C increase results in a doubling of chemical activity. 

Attempts have also been made to exploit the relatively high molecular activity of cod trypsin at low temperatures by incorporating the enzyme into herring "fermentations that proceed at 10°C. The preparation of brine-fermented round herring (matjes) is limited to certain seasons because of the balance of digestive enzymes in the fish at this time. Other studies have indicated that proteinases are important components in matje fermentation 41),... 

Dixon and Webb present an extensive consideration of activation mechanisms involving the reversible binding of an activator (less often termed an agonist ) to the enzyme. Nonessential activation refers to enzyme-de-pendent processes that can convert substrate(s) to prod-uct(s) in the absence of the activator, albeit at a slower rate. Essential activators are molecular entities that are required by the enzyme in the catalysis of a reaction. In a sense, essential activators are similar to second (or third) substrates, albeit they are not converted to products. An example of an essential activator might be an enzyme that requires the binding of a metal ion for catalysis to proceed. Below are a few cases of essential activation. 

CYP2El s substrates are small molecular weight aromatic hydrocarbons or their methyl derivatives, as the enzyme s active site is relative small and restricted. In a recently published study from Lewis et al. (235) eight alkyl benzenes, which undergo oxidative metabolism via human CYP2E1, were used to... 

Here Et is the total enzyme, namely, the free enzyme E plus enzyme-substrate complex ES. The equation holds only at substrate saturation, that is, when the substrate concentration is high enough that essentially all of the enzyme has been converted into the intermediate ES. The process is first order in enzyme but is zero order in substrate. The rate constant k is a measure of the speed at which the enzyme operates. When the concentration [E]t is given in moles per liter of active sites (actual molar concentration multiplied by the number of active sites per mole) the constant k is known as the turnover number, the molecular activity, or kcat. The symbol fccat is also used in place of k in Eq. 9-6 for complex rate expressions in which fccat cannot represent a single rate constant but is an algebraic expression that contains a number of different constants. 

---