Protein slope

The region before the first temperature break is characteristic of the buried Trp residues, the second region going from the first to the second temperature breaks is characteristic of the surface Trp residue and the one that begins at the second break is characteristic of the solvent. The value of bi is a linear-weighted combination of each temperature of the protein slope of the first tryptophan bui and the solvent slope of the second ... 

The presented algorithm was applied to 4 proteins (lysozyme, ribonuclease A, ovomucid and bovine pancreatic trypsin inhibitor) containing 51 titratable residues with experimentally known pKaS [32, 33]. Fig. 2 shows the correlation between the experimental and calculated pKaS. The linear correlation coefficient is r = 0.952 the slope of the line is A = 1.028 and the intercept is B = -0.104. This shows that the overall agreement between the experimental and predicted pKaS is good. 

Isoelectric Focusing. Isoelectric focusing is a technique used for protein separation, by driving proteins to a pH where they have no mobiUty. Resolution depends on the slope of a pH gradient that can be achieved in a gel. 

The ionic species of the mobile phase will also affect the separation. This is shown in Table 4.3 by the difference in resolution values for magnesium chloride buffer compared to sodium sulfate buffer. In addition, calibration curves for proteins in potassium phosphate buffers are shallower than those generated in sodium phosphate buffers. The slope of the curve in Sorenson buffer (containing both Na and ) is midway between the slopes generated with either cation alone (1). Table 4.4 illustrates the impact of different buffer conditions on mass recovery for six sample proteins. In this case, the mass recovery of proteins (1,4) is higher with sodium or potassium phosphate buffers (pH 6.9) than with Tris-HCl buffers (pH 7.8). 

FIGURE 3.3 The enthalpy change, ATT, for a reaction can be determined from the slope of a plot of R In versus l/T. To illns-trate the method, the values of the data points on either side of the 327.5 K (54.5 C) data point have been nsed to calculate ATT at 54.5 C. Regression analysis would normally be preferable. (Adapted from Brandts, ]. F., 1964. Tim thermo-dynamics of protein denatnration. I. The denatnration of ehy-motrypsinogen. om Q.7A of the American Chemical Society m 429 -430L)... 

If BjX is plotted against B (the Scatchard plot) it should give a straight line (Fig. 5.2(b)) with the slope (1/A d) giving K and the intercept on the abscissa providing the maximal binding (.Smax) expressed as fmol per mg tissue protein. The steeper the slope, the higher the affinity. 

Equation (20-80) requires a mass transfer coefficient k to calculate Cu, and a relation between protein concentration and osmotic pressure. Pure water flux obtained from a plot of flux versus pressure is used to calculate membrane resistance (t ically small). The LMH/psi slope is referred to as the NWP (normal water permeability). The membrane plus fouling resistances are determined after removing the reversible polarization layer through a buffer flush. To illustrate the components of the osmotic flux model. Fig. 20-63 shows flux versus TMP curves corresponding to just the membrane in buffer (Rfouimg = 0, = 0),... 

A general feature of optimum sample preparation is that maximum recovery of the analyte is observed. Consider a graph of recovery vs. variation in one experimental condition. Figure 5 shows such a graph, with temperature as the experimental variable. The curve exhibits a maximum and a decline on either side of the maximum. The assay will be most reproducible at the point of zero slope, i.e., at the maximum recovery, because small variations in conditions will not affect the result. In hydrolysis of a protein to its constituent amino acids, for example, it will be found that at very high temperatures or long hydrolysis times, degradation of the product amino acids occurs, while at low temperatures or short hydrolysis times, the protein... 

The number of binding sites can be determined in this model by a plot of d Ink /dlnm at constant temperature, pH, and ion valency. To do that, it may be assumed that dlny /dlnm is approximately zero. The actual value is -0.04 for 0.1 to 0.5 M sodium chloride and less at lower concentrations. To a first approximation, the stoichiometry of water molecules released by binding protein could be determined from the slope of the plot of dink /dlnm vs. m. However, especially at low salt concentration and near the isoelectric point, the slope of such plots is nonlinear. The nonlinearity may be due to hydrophobic interaction between stationary phase and protein or a large change of ionic hydration on binding.34... 

Here m is the slope value and [ii]app is the apparent total enzyme concentration, typically estimated from protein assays and other methods (Copeland, 1994). Note from Equation (7.13) that when our estimate of enzyme concentration is incorrect, the slope of the best fit line of IC50 as a function of [E] will not be 1/2, as theoretically expected. Nevertheless, the v-intercept estimate of K pp is unaffected by inaccuracies in [ ]. In fact we can combine Equations (7.12) and (7.13) to provide an accurate determination of [ /]T from the slope of plots such as those shown in Figure 7.2. The true value of [ii]T is related to the apparent value [ TPP as... 

The permeation profiles of the proteins through the polymer membrane can be approximated by two straight lines representing regions that were in the dark and under UV irradiation as shown in Figure 6. The permeation coefficients can be calculated from the slope of these straight lines by using the equation... 

On the basis of spot position and intensity, by assuming different critical interdistance Ax o values, experimental points can be obtained and fitted by a straight line (Eq. 4.13) whose slope represents a statistical estimation of m, the estimated number of single components. The values estimated for each ID strip were added to obtain the total number of proteins (Pietrogrande et al., 2002). 

Figure 15.8 (a) Time course of the activity restoration of formalin-treated RNase A during incubation at 50°C (0-2h) and 65°C (2-4h) in TAE buffer, pH 7.0. (b) Time course of the activity restoration of formalin-treated RNase A during incubation at 65°C in TAE buffers of various pH values. All RNase A preparations were freed of excess formaldehyde by dialysis prior to the assay. The RNase A activity was determined with a colorimetric assay using cytidine 2,3,-cyclophosphate as the substrate as described by Crook et al.54 Note that the slopes of the curves decrease with incubation time at 65°C, which is near the denaturation temperature of native RNase A. This loss of activity is likely due to the competing effect of protein denaturation of the recovered RNaseA at this temperature. See Rait et al.10 for details. 

Enzymes can be used not only for the determination of substrates but also for the analysis of enzyme inhibitors. In this type of sensors the response of the detectable species will decrease in the presence of the analyte. The inhibitor may affect the vmax or KM values. Competitive inhibitors, which bind to the same active site than the substrate, will increase the KM value, reflected by a change on the slope of the Lineweaver-Burke plot but will not change vmax. Non-competitive inhibitors, i.e. those that bind to another site of the protein, do not affect KM but produce a decrease in vmax. For instance, the acetylcholinesterase enzyme is inhibited by carbamate and organophosphate pesticides and has been widely used for the development of optical fiber sensors for these compounds based on different chemical transduction schemes (hydrolysis of a colored substrate, pH changes). 

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