Reactive site residue

The reaction of the proteinase with the inhibitor occurs at a basic residue (Arg or Lys residue) in a loop that extends away from the globular inhibitor molecule. This basic residue, called the reactive site residue, is locked in the proteinase active site as the acyl enzyme. The reactive site... 

Proteinase inactivation occurs by a stoichiometric reaction between proteinase and inhibitor that results in the formation of a covalent ester bond between the reactive site residue of the inhibitor (Arg in antithrombin) and the active site residue (Ser in the proteinase). The proteinases thrombin, factor Xa, factor IXa, and, less effectively, factor Vila and factor XIa are all inactivated by antithrombin (Figure 36-16). Other SERPINS can inactivate procoagulant proteinases, heparin cofactor II can inactivate thrombin, and O I-proteinase inhibitor can inactivate factor Xa. An altered a i -proteinase inhibitor (a i -proteinase inhibitor... 

CHAPTER 36, FIGURE 7 A group of structurally similar protein inhibitors of the serine proteinases known as SERPINS (SERine Proteinase INhibitors). The structure shown is human antithrombin. The reference SERPIN, a j-proteinase inhibitor or a. -antitrypsin contains -30% a helix (9 helices) and 40% sheet (5 3 sheets). Other members of the SERPIN family contain both additional helices and p sheets. The reactive center loop of antithrombin, residues 378-396, contains the reactive site residues Arg and Ser . Upon reaction with the target proteinase or after cleavage by the target proteinase (a reaction that inactivates the inhibitor without inactivating the proteinase), the reactive center loop folds between the S3 and S5 sheets. 

Kaplan and DeGrado [9] to catalyze the oxidation of aminophenol by a two-electron transfer pathway, Scheme 5.4. The catalyst has been extensively characterized with regards to structure and the designed protein provided not only an active site cavity but also a channel through which the active site could be reached. The reaction mechanism and reactive site residues were determined. 

Importantly, the crystal structure of 34 complexed with N9 sialidase (Fig. 8) indicated differences in the orientation of the guanidino group in subsite S2, and in its interaction with the active site residues, compared to that of zanamivir (Babu et al. 2000). These differences have implications for cross-reactivity of 34 with zanamivir-resistant influenza viruses that have Glul 19 mutations in the sialidase S2 subsite (see Sect. 5.1). 

The primary site of action of OPs is AChE, with which they interact as suicide substrates (see also Section 10.2.2 and Chapter 2, Figure 2.9). Similar to other B-type esterases, AChE has a reactive serine residue located at its active site, and the serine hydroxyl is phosphorylated by organophosphates. Phosphorylation causes loss of AChE activity and, at best, the phosphorylated enzyme reactivates only slowly. The rate of reactivation of the phosphorylated enzyme depends on the nature of the X groups, being relatively rapid with methoxy groups (tso 1-2 h), but slower with larger... 

Lys [47,48] and the fluorescein-5-isothiocyanate (FITC) reactive site Lys [49,50], as well as a region between residue 157 and 300, which has been proposed as part of an energy transduction region [40,44]. 

However, diffusion of the reactive QM out of the enzyme active site is a major concern. For instance, a 2-acyloxy-5-nitrobenzylchloride does not modify any nucleophilic residue located within the enzyme active site but becomes attached to a tryptophan residue proximal to the active site of chymotrypsin or papain.23,24 The lack of inactivation could also be due to other factors the unmasked QM being poorly electrophilic, active site residues not being nucleophilic enough, or the covalent adduct being unstable. Cyclized acyloxybenzyl molecules of type a could well overcome the diffusion problem. They will retain both the electrophilic hydroxybenzyl species b, and then the tethered QM, in the active site throughout the lifetime of the acyl-enzyme (Scheme 11.1). This reasoning led us to synthesize functionalized... 

In the synthesis of polypeptides with biological activity on a crosslinked polymer support as pioneered by Merrifield (1 2) a strict control of the amino acid sequence requires that each of the consecutive reactions should go virtually to completion. Thus, for the preparation of a polypeptide with 60 amino acid residues, even an average conversion of 99% would contaminate the product with an unacceptable amount of "defect chains". Yet, it has been observed (13) that with a large excess of an amino acid reagent —Tn the solution reacting with a polymer-bound polypeptide, the reaction kinetics deviate significantly from the expected exponential approach to quantitative conversion, indicating that the reactive sites on the polymer are not equally reactive. 

As mentioned previously, additive treatments involve the application of a polymer to the fibre. This is usually prepared before application and contains reactive groups. However, it is also possible to form the polymer in situ within the fibres. The traditional approach is to apply the polymer after a subtractive oxidation treatment but environmental concern over A OX problems is increasing demand for additive treatments that can stand alone. There is no denying that the oxidative step can facilitate subsequent treatment with a polymer, since the scission of cystine disulphide bonds to yield cysteic acid residues provides useful reactive sites for crosslinking or anchoring the polymer. 

Another reactive site, called the T-site, makes a modest contribution to the overall hydrolytic activity of the protein (ca. 11%), and a lysine residue has been suggested as the catalyst. The position of the T-site might be in the subdomain IIA of HSA (Fig. 3.17), since there is evidence that Lys220 in sub-domain IIA could belong to an esterase site [119][120],... 

The hydrolysis of acetylsalicylic acid (aspirin) has also been described, and it involves the rapid acetylation of Lys199 [123], This reactive site also involves Trp214 and has been called the U-site [124], Drugs that inhibit the hydrolysis of substituted acetylsalicylic acids by HSA and decrease the fluorescence intensity of Trp214 are referred to as U-type drugs (e.g., sulfinpyrazone and warfarin). Because U-type drugs also bind to the site know as site I (which overlaps with subdomain IIA), the U-site and I-site are believed to overlap. Lysine residues also appear to be involved in the /3-lactamase activity of HSA [125],... 

Bowman-Birk protease inhibitors (BBIs) are among the best-studied serine protease inhibitors. They are found abundantly in dicotyledonous and monocotyledonous plants, with the former species expressing inhibitors of approximately 8 kDa in size with two reactive sites (double headed) and the latter expressing 8 kDa inhibitors with one reactive site and 16 kDa inhibitors with two reactive sites. Dicot BBIs feature 14 cysteine residues involved in disulfide bonds monocot BBIs have 10 cysteine residues. ... 

The reactive site of BBIs usually consists of a seven-residue loop held in position by a disulfide bond from cysteine residues at the termini of the loop. The loop itself forms the tip of a double-stranded /3-sheet extending... 

Figure 10 The global structures of Bowman-Birk inhibitors are diverse, but the reactive sites are structurally highly conserved. Reactive sites are oriented to the bottom right corner. The double-stranded /3-sheets neighboring the reactive sites are clearly visible, (a) 1 mvz, (b) 1 bbi, and (c) 1 h34. (d) The overlay of the backbone traces of the three BBIs ((a)-(c), colors as before) reveals their structural similarity. Additionally SFTI-1 (green, Ijbl) adopts the same fold in its active site. The residues (Arg in 1 mvz, Lys for other peptides) are also shown N and C denote the N- and C-termini of the truncated loops.

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