Enzymes adsorption

Modeling of enzyme adsorption isotherms on an inorganic support using the Scatchard equation Equation (1) allow one to determine the binding constant. 

In this communication a study of the catalytic behavior of the immobilized Rhizomucor miehei lipase in the transesterification reaction to biodiesel production has been reported. The main drawbacks associated to the current biodiesel production by basic homogeneous catalysis could be overcome by using immobilized lipases. Immobilization by adsorption and entrapment have been used as methods to prepare the heterogeneous biocatalyst. Zeolites and related materials have been used as inorganic lipase supports. To promote the enzyme adsorption, the surface of the supports have been functionalized by synthesis procedures or by post-treatments. While, the enzyme entrapping procedure has been carried out by sol-gel method in order to obtain the biocatalyst protected by a mesoporous matrix and to reduce its leaching after several catalytic uses. 

Moreover, the catalytic results clearly show that the lipase immobilization procedure strongly influences the final activity of the enzyme. Adsorption and entrapping procedures allow to preserve the open and active conformation of the enzyme whit respect to electrostatic ones. Using the entrapped lipase, the enzyme leaching can be avoided and the biodiesel total productivity strongly increases if compared with the one obtained by the free enzyme. 

Quiquampoix H, Servagent-Noinville S, Baron M-H (2002) Enzyme adsorption on mineral surfaces and consequences for the catalytic activity. In Bums RG, Dick RP (eds) Enzymes in the environment. Activity, ecology and applications. Marcel Dekker, New York, USA, pp 285-306... 

This broad class of hydrolases constitutes a special category of enzymes which bind to and conduct their catalytic functions at the interface between the aqueous solution and the surface of membranes, vesicles, or emulsions. In order to explain the kinetics of lipolysis, one must determine the rates and affinities that govern enzyme adsorption to the interface of insoluble lipid structures -. One must also account for the special properties of the lipid surface as well as for the ability of enzymes to scooC along the lipid surface. See specific enzyme Micelle Interfacial Catalysis... 

Proteins, enzymes Adsorption and electroreduction of proteins at mercury electrodes have been reviewed by several workers. Our short summary is partly based on the paper by Honeychurch [94]. Kuznetsov et al. [95] have suggested that the segments of the protein that interact with the surface, belong to relatively hydrophobic regions and they either denature or unfold following irreversible adsorption on mercury surface. Proteins spread on mercury electrodes [96] and form a layer of 8-10 A in thickness, corresponding to the single polypeptide chain. 

Results of enzyme adsorption and tests in the MSR are given in Table 1. 

The carrier materials used for enzyme adsorption must have a highly porous character and a pore size distribution, which should facihtate a free diffusion of enzyme into the carrier. Furthermore, substrate and product should also be able to diffuse freely. This is especially important in the case of large protein substrates, where diffusion into the pore can be a problem (e.g. whey protein and casein). 

Fig. 24.4 Mechanism of enzyme inactivation at an aqueous-organic interface. Step 1 reversible enzyme adsorption to the interface and concomitant enzyme structural rearrangement at the interface. Step 2 unfolding of enzyme molecule at the interface. Step 3 desorption of inactivated/un-folded enzyme molecules from the interface. Step 4 irreversible aggregation and precipitation of inactivated enzyme. (From [34])...

Tietjen,T., and Wetzel, R. G. (2003). Extracellular enzyme-clay mineral complexes Enzyme adsorption, alteration of enzyme activity, and protection from photodegradation. Aq. Ecology 37, 331-339. 

Using a cylindrical internal reflectance (CIRcle) cell and GC-IR data collection software, it was determined for both lysozyme and BPN, that most of the enzyme adsorption occurred within ten seconds after injection. Nearly an order-of-magnitude more BPN adsorbed on the hydrophobic surface than the hydrophilic one, while lysozyme adsorbed somewhat more strongly to the hydrophilic Ge surface. Over time periods of about one day, the lysozyme layer continued to increase somewhat in thickness, while BPN maintained its initial coverage. 

Starch can be enzymically converted in the presence of pigment. The conversion follows a similar time-temperature cycle as in neat starch conversion. The pigment will adsorb a portion of the enzyme adsorption can be minimized by the addition of sodium silicate to the mixture prior to the addition of the enzyme (Vanderbilt process). Even with silicate treatment, a higher quantity of enzyme will be required to reach a specific viscosity target. Other coating components, such as latex and lubricants, have to be added after the conversion. The Vanderbilt process is now rarely used for the preparation of coating binder. 

Fig. 14.25. (A). The scheme of enzyme adsorption of the electrode with different lipid interlayers. (B). The relative oxygen reduction rate vs. the distance between the electrode and the enzyme for lactase adsorbed (1) on soot, (2) on cholesterol, (4) on lecithin. The curves a and b are calculated for barrier heights of 4 and 5 eV, respectively. (Reprinted from J. O M Bockris, M. Szklonzyk, and Szucs, in Electropharmacology, G. M. Eckert, F. Gutmann, and H. Keyzar, eds., Figs. 25,29, 30,1990. Reproduced with permission of CRC Press.)...

Immobilization by adsorption. This is the simplest method and consists in enzyme adsorption due to electrostatic, hydrophobic or dispersive forces on the electrode surface. The disadvantage is the high probability of enzyme desorption and denaturing. 

The adsorption of horseradish peroxidase (HRD) on gold and polymer modified resonator surfaces was studied by quartz crystal microbalance. It was shown that the HRP adsorption reached saturation in 5-10 min. The rate constants and the maximum enzyme adsorption were determined. The thickness of HRP/PSS bilayer was calculated. 

Woodward, J. Immobilized enzymes adsorption and covalent coupling. In Immobilized Cells and Enzymes A Practical Approach Woodward, J., Ed. JRL Oxford, 1985 3-17. 

Brunauer-Emmet-Teller (BET) estimated surface areas [23], For example, from Figure 5.9, graphite felt electrodes show poor volume-normalized ORR current density compared to carbon nanofibers and multiwaUed carbon nanotube (MWCNT)-based electrodes. However, the results also reveal that CNTs and porous carbon tubes exhibit dramaticaUy lower ORR current densities when normalized to B ET surface area, while graphite felt electrodes perform better, perhaps indicative of agglomeration of the carbon tubes, preventing enzyme adsorption over the entire area. Further research on methods to permit dispersion of nano-tubes, while retaining electrical conductivity and adsorption of enzymes oriented for DET, is warranted. 

Because cellobiohydrolases and endoglucanases act on insoluble cellulose, the rate of enzyme adsorption correlates to the rate of enzyme-substrate complex formation. Since P-glucosidase acts upon soluble cellobiose, MichaeUs-Menten kinetics can be used to model its activity. This is an example of the uti-... 

The presence of various functional surface groups and the high conductivity and porosity of carbon material permit effective enzyme adsorption. Glucose oxidase has been irreversibly adsorbed to a graphite electrode by drying a concentrated enzyme solution on the surface (Ikeda et al., 1984). In the presence of p-benzoquinone an electrocatalytic current was observed at 500 mV vs SCE. The measuring signal was... 

Figure 1 shows clearly that the total process can be represented by Equation 2 for several bulk enzyme concentrations. However, the evolution of the surface radioactivity of the tritiated enzyme under the oligomer film (Figure 2) indicates that enzyme activity and adsorption of the enzyme at the interface are not interrelated—namely, the enzyme kinetics at 1 min are identical at 10 min, when most of the enzyme adsorption takes place. 

The evolution of the enzyme adsorption was studied with tritiated molecules at pH 2-11 (Figure 5). Even for a low adsorption of enzyme molecules, maximum activity can be obtained. Thus, either the molecules initially adsorbed contribute to the kinetics, or the enzyme required for activity is not permanently adsorbed. If it were, the small quantities would be immeasurable with our techniques and indistinguishable from the ordinary adsorption or film penetration. 

Influence of Subphase Temperature. Enzymic activity at the interface increases clearly with subphase temperature (Figure 6). When comparing enzymic adsorption with enzymic activity (Figure 7), the rate of enzymic cleavage increases when the number of macromolecules of the enzyme that reaches the surface increases. The two processes are parallel but are not immediately related. 

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