Enzymes calculations

The CD spectrum of the C188S mutant is essentially the same as that of the wild-type enzyme, which reflects that the tertiary structure of this mutant changed little compared to that of the wild-type enzyme. Calculation of the content of secondary structure of the mutant enzyme based on J-600S Secondary Structure Estimation system (JASCO) also showed that there is no significant change in the secondary structure of the mutant. The fact that the k value of this mutant is extremely small despite little change in conformation clearly indicates that Cysl88 is located in the active site. 

Figure 13.10 Price—Volume of use relationships for different classes of commercially important enzymes (calculated from data in Poldermans, 1989).

Intracellular Concentration of Enzymes To approximate the actual concentration of enzymes in a bacterial cell, assume that the cell contains equal concentrations of 1,000 different enzymes in solution in the cytosol and that each protein has a molecular weight of 100,000. Assume also that the bacterial cell is a cylinder (diameter 1.0 /xm, height 2.0 /rm), that the cytosol (specific gravity 1.20) is 20% soluble protein by weight, and that the soluble protein consists entirely of enzymes. Calculate the average molar concentration of each enzyme in this hypothetical cell. 

The amino acid composition of B. subtilis and E. coli enzymes, calculated for a minimum subunit of 33,500, show a greater divergence. Whereas the E. coli enzyme is not unsimilar to both the mitochondrial and cytoplasmic isozymes from eukaryotic sources, the B. subtilis protein is decidedly different. Interestingly, the E. coli enzyme possesses a molecular weight of about 67,000 as compared to the 117,000 of the B. subtilis protein (S7). Primary structure analyses will be required to ascertain the extent to which either of these prokaryotic enzymes are sequentially related to either of the eukaryotic forms of the enzyme. 

Figure 9.32 Structures, EC50 values for the activation of recombinant human enzyme, calculated LogP values, and unbound percentages for a series of glucokinase activators. ...

For really accurate enzyme calculations, we argue that detailed QM/MM calculations are needed. These should include single-point calculations with 600-1000 atom QM systems and estimations of QM/MM free energies, as well as extrapolations to coupled-cluster energies, as in the thorough studies by Thiel and coworkers on XO and aldehyde oxidoreductase. We have started similar studies on DMSOR and... 

Domain Residue DNA Binding Residues Auto- NAD modification Binding Residues Residues Native Enzyme Calculated Determined Residues (%) (%) ... 

Table 6.13 The microbial production of enzymes (calculated as the weight of the pure protein) in 1979.

A good relationship exists between inactivation and equivalents of reagent bound to the enzyme calculated by spectrophotometric method. 

DTT, and EDTA, (4) pepstatin and iodoacetate. The first tube would be expected to reflect the full proteolytic activity of all protein-ases except for metalloproteases the second would be expected to permit only the expression of non-thiol proteases the third would be expected to show no cathepsin D activity, and the fourth would be expected to block all thiol and carboxyl enzymes. Calculations based on these four values were used to estimate the results in Figure 2. 

Fig. 10. Concentration profiles of intermediate product in a membrane containing two successive enzymes calculated using the simplified Equation (20). First enzyme at maximum rate profiles and fluxes are dependent on the relative activity of second enzyme compared to first enzyme and diffusion fluxes it is seen that if the value Vmb -Vma is negative, the system produces p if this quantity is positive p is degraded by the system if it is zero the concentration profile in the membrane is horizontal and the concentration of p remains unmodified. (From Broun et al [42] and Thomas [44]).

Calculation of Conformational Free Energies for a Model of a Bilobal Enzyme Protein kinases catalyze the transfer of phosphate from adenosine triphosphate (ATP) to protein substrates and are regulatory elements of most known pathways of signal transduction. 

Table 2 shows the results of our preliminary calculations of the pKa of the Cys403 residue, for several different models of the enzyme, based on two structures available from the PDB. In the case of the YPT structure, a crystal water molecule is close to Cys403 and was included in some of the calculations as part of the protein (i.e. it was treated with the same internal dielectric as that of the protein). Simulations denoted as -I-H2O in Table 2, include a crystallographically resolved, buried water molecule, situated 3.2lA from... 

The problem with most quantum mechanical methods is that they scale badly. This means that, for instance, a calculation for twice as large a molecule does not require twice as much computer time and resources (this would be linear scaling), but rather 2" times as much, where n varies between about 3 for DFT calculations to 4 for Hartree-Fock and very large numbers for ab-initio techniques with explicit treatment of electron correlation. Thus, the size of the molecules that we can treat with conventional methods is limited. Linear scaling methods have been developed for ab-initio, DFT and semi-empirical methods, but only the latter are currently able to treat complete enzymes. There are two different approaches available. 

Application of the CCM to small sets (n < 6) of enzyme inhibitors revealed correlations between the inhibitory activity and the chirality measure of the inhibitors, calculated by Eq. (26) for the entire structure or for the substructure that interacts with the enzyme (pharmacophore) [41], This was done for arylammonium inhibitors of trypsin, Di-dopamine receptor inhibitors, and organophosphate inhibitors of trypsin, acetylcholine esterase, and butyrylcholine esterase. Because the CCM values are equal for opposite enantiomers, the method had to be applied separately to the two families of enantiomers (R- and S-enantiomers). 

This means that the methods developed for the calculation of physicochemical effects can also be used to deepen our understanding of biochemical rcaaions. Clearly, electronic effects within the substrate molecule arc not the only ones determining its reactivity, The binding of the substrate to the enzyme is also influenced... 

Molecular volumes are usually computed by a nonquantum mechanical method, which integrates the area inside a van der Waals or Connolly surface of some sort. Alternatively, molecular volume can be determined by choosing an isosurface of the electron density and determining the volume inside of that surface. Thus, one could find the isosurface that contains a certain percentage of the electron density. These properties are important due to their relationship to certain applications, such as determining whether a molecule will fit in the active site of an enzyme, predicting liquid densities, and determining the cavity size for solvation calculations. 

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