Protein substrate sites

In free CDK2 the active site cleft is blocked by the T-loop and Thr 160 is buried (Figure 6.20a). Substrates cannot bind and Thr 160 cannot be phosphorylated consequently free CDK2 is inactive. The conformational changes induced by cyclin A binding not only expose the active site cleft so that ATP and protein substrates can bind but also rearrange essential active site residues to make the enzyme catalytically competent (Figure 6.20b). In addition Thr... 

FIGURE 7.1 Enzyme ortho- and allosterism as presented by Koshland [2], Steric hindrance whereby the competing molecules physically interfered with each other as they bound to the substrate site was differentiated from a direct interaction where only portions of the competing molecules interfered with each other. If no direct physical interaction between the molecules occurred, then the effects were solely due to effects transmitted through the protein structure (allosteric). 

HSFl phosphorylation must be sensitive to nonheat inducers of HSF-DNA binding activity because HSFl phosphorylation can be achieved at 37 °C by other inducers of the HS response. HSF 1 contains polypeptide sequences that could serve as substrates for well characterized protein kinases, but few of these are known to be heat inducible. One family of protein kinases, the S6 protein kinases, have already been shown to exhibit heat inducible activity however, their peak level of activity during HS occurs well after the maximal induction of HSF phosphorylation (Jurivich et al., 1991). Thus, other protein kinases are likely to be directly linked to the phosphorylation of HSF. Some of the putative protein phosphorylation sites on HSF include motifs for protein kinase C, casein kinase, and enterokinase. There are tyrosine sequences that match substrates for known tyrosine kinases, but whether these residues are accessible to phosphorylation is not established. 

It is not a trivial matter to get a converged value for these simulations, since in both we are forcing the substrate to vanish from the system a substantial mutation. But if one has copious computer time available and is careful, one has the potential for calculating such a value provided the substrate is not too large and there are not appreciable large-scale changes in the protein active site upon binding. 

The protein-based clotting process is a classic example of an enzyme cascade (see Figure 5.23). The clotting factors (which are designated with a Roman numeral, I to XIII) are synthesized in the liver and circulate in the blood as inactive precursors, strictiy, proenzymes. Most of the clotting factors are serine protease enzymes, that is they are enzymes which cleave other proteins (substrates) by a mechanism which involves a serine residue at the active site. 

R. J. Bloch, M. Velez, J. Krikorian, and D. Axelrod, Microfilaments and actin-associated proteins at sites of membrane-substrate attachment within acetylcholine receptor clusters, Exp. Cell Res. 182, 583-596 (1989). 

Fig. 6.15 FAC-MS chromatograms of dual indicators for protein kinase Ca [32]. (a) In the chromatograms, the red lines correspond to a void marker, the blue lines correspond to the substrate-site indicator chelerythrine chloride and the magenta lines correspond to the ATP-site indicator PDl53035. Arrows...

Figure 3/ for example/ places the lanosterol so as the 3f hydroxyl polar group lies over the propionate side chains. To reduce the complexity of this picture one can now replace the lanosterol structure by a surface canopy to represent the extent of the hydrophobic substrate binding site. There is also the facility to code this surface to signify the electronic properties of the substrates such as their electron density/ electrostatic potential/ or HOMO/LUMO values. Theoretical work of this type is currently suggesting quite remarkable complementarity of electron properties between bound substrates and protein binding sites. (10). 

The following are recent reviews on the molecular and physical properties of these transferases which transfer geranylgeranyl groups to defined sites on protein substrates. 

Figure 6. An example of inter-family target hopping between human and viral aspartyl proteases. The aspartyl protease active site is located at a homodimer interface in HIV and within a single domain in Cathepsin D, so sequence and structure alignments between these proteins cannot be constructed. By using an approach independent of sequence or structure homology to directly align the sites, SiteSorter finds that the HIV protease and Cathepsin D substrate sites are highly similar (identical chemical groups within 1 A are colored dark blue). It has been verified experimentally that Cathepsin D is susceptible to inhibition by HIV-protease inhibitors. ...

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