Apolar binding sites

Synthetic heterocyclic and modified amino acid derivatives have been grouped in a class of thrombin inhibitors called peptidomimetics. An example of such a compound is argatroban, with a molecular mass of 532 Da. It blocks thrombin s active catalytic site by binding to the adjacent apolar binding site. This selective reversible inhibitor of thrombin has a K of 19 nM and blocks thrombin s role in coagulation and fibrinolysis (62). 

As was mentioned earlier, polyethylenimine has many amine nitrogens in it, and these have the potential to act as nucleophiles (e.g., in an aminolysis reaction). In addition, they are locally concentrated (Figs. 2 and 3). Furthermore, polymers with attached aliphatic acyl groups provide apolar binding sites in proximity to amine residues of the polymer. One might expect, therefore, to find progressively enhanced rates of... 

Collectively, the direct thrombin inhibitors are prototypically represented by hirudin, the antithrombotic molecule found in the saliva of the medicinal leech (Hirudo medicinalis), This protein is a 65 amino acid molecule that forms a highly stable but noncovalent complex with thrombin (7). With two domains, the NH2-terminal core domain and the COOH-terminal tail, the hirudin molecule inhibits the catalytic site and the anion-binding exosite in a two-step process. The first step is an ionic interaction that leads to a rearrangement of the thrombin-hirudin complex to form a tighter bond that is stoi-chiometrically I I and irreversible. The apolar-binding site may also be involved in hirudin binding. This complex and... 

To help characterize the interactions of some drugs and metabolites with HSA and their competition for its limited number of binding sites, we and others have studied the influence of those substrates on its acetylation by p-nitrophenyl acetate (29,30,31,32,33). This reaction is very fast and results in the irreversible acetylation of a particular tyrosine residue (i.e. 411) which is located in a major apolar binding site of that protein (34). Because this reaction is so fast under most conditions, competing reactions are not significant and formation of p-nitrophenolate is a convenient reflection of the reaction rate (35). 

Berliner L J, Shen Y Y L (1977). Physical evidence for an apolar binding site near the catalytic centre of human a-thrombin. Biochem. 16 4622-4626. 

Figure 4.10. Schematic representation of the co-operativity displayed by binding sites in the formation of organic host-guest complexes. S=hinge, M=metal ion, A=apolar binding site, B=polar binding site, L=lipophilic guest molecule. Reproduced from [182].

Many enzymes contain deeply buried apolar binding sites within their globular structures. We were interested in using dendritically shielded cyclophanes to mimic such behaviour and this led to the synthesis of a new class of molecules the dendrophanes dendniic cyc ophanes).[ 6] The synthesis of dendrophanes such as 2 (Fig. 2) was... 

The hydrophobic domain formed by the vinyl backbone represents an apolar binding site for the more apolar substrates. Mirejovsky (15) has recently proposed that the conformational changes are attributed to an inherent tendency of imidazole to aggregate resulting in an inefficient utilization of imidazole and a decrease in catalytic efficiency. Therefore, it is difficult to conceive, as suggested by the author, how the apolar interactions of poly(l-methyl-5-vinyl-imidazole) and substrate increases the observed catalytic efficiency. 

The binding site environments of certain LBPs (the NPAs in particular) appear to be extremely, and so far inexplicably, apolar. 

It has been found that the catalytic activity of PKC is enhanced by a lipid component of the cell membrane, namely phosphatidylserine. This activity is further stimulated by sn-1,2-diacylglycerol. Oleic acid also activates the enzyme in the presence of 1,2-diacylglycerol, and thus it is presumed to mimic phosphatidylserine. In order to identify that modulating binding site for oleic acid on PKC, a photoaffinity analogue was devised. A carbene generating photophore, diazirine was placed in the apolar terminus of the unsaturated fatty acid ligand (30, Fig. 12). The synthesis and the photochemical activation properties were reported by Ruhmann and Wentrup [113]. 

Finally, L-type /2-solenoids have an unusual inverted arch (Fig. 12). Stacking of these arches makes a groove which forms the center of the binding site for polysaccharides or pectins (Jenkins and Pickersgill, 2001). In contradistinction to the other arches, the arc residues of inverted arches are interior-facing and are apolar, while residues in the -conformation which bound the arc, face the solvent and are mostly polar (Fig. 12). These arcs have ab conformations. Frequently the first arc residue is small, glycine or alanine, and the second position is occupied by a bulky apolar residue. 

The stability of crown-ether complexes depends on several factors these include cavity size of the ligand, cation diameter, spatial distribution of ring binding sites, the character of the hetero-atoms, the presence of additional binding sites and the type of solvent used. In apolar solutions it also depends on the nature of the anion. The effects of these parameters will be illustrated in the next sections. 

S6) and since the apolar product is not negatively charged, we felt that our catalyst with strong binding sites would be effective with this substrate. The hydrolysis was followed to 75% of completion, and a rate enhancement of 230-fold over the imidazole catalysed rate was found at pH 8.2 (Fig. 7). Constant activity after repeated treatments with excesses of substrate showed that the catalyst was regenerated. 

Local anesthetic activity is also shown by uncharged substances, suggesting a binding site in apolar regions of the channel protein or the surrounding lipid membrane. 

The most frequent protein in the plasma, at around 45 g is albumin. Due to its high concentration, it plays a crucial role in maintaining the blood s colloid osmotic pressure and represents an important amino acid reserve for the body. Albumin has binding sites for apolar substances and therefore functions as a transport protein for long-chain fatty acids, bilirubin, drugs, and some steroid hormones and vitamins. In addition, serum albumin binds Ca "" and Mg "" ions. It is the only important plasma protein that is not glycosylated. 

Several additional features of the model are noteworthy. First, it is possible to build it without straining chemical bonds or causing unfavorable steric interactions. The polyamine chain is sufficiently long to reach around a cluster, but it is not so long or so bulky as to cause excessive crowding near the surface of the cluster. The spaces at this surface between the polyamine chains (Fig. 11) are likely binding sites for small apolar molecules, since such molecules can be bound at the interface or partially penetrate into the domain of the hydrocarbon sphere in response to favorable apolar interactions. In the model shown in Fig. 11, three bound p-nitrophenyl caproate molecules have also been included to illustrate possible modes of binding. An arrow points to one of these small molecules. 

The modified polyethylenimines described so far are only a few of many possibilities. It is obvious that this polymer provides a remarkably versatile macromolecular matrix for the attachment of a wide variety of different types of functional groups. Furthermore, the polymer framework makes it possible to juxtapose a binding site, a catalytic group, and an apolar-aqueous interface in a locally compact array. Thus a wide range of local environments can be created on this macromolecular water-soluble catalyst. We hope to be able to exploit these to obtain a series of synthetic macromolecules with tailor-made catalytic properties. 

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