Hydrolysis proteinase activity

Bishop et al. (1984) have isolated a pectic polysaccharide (6 kDa) from tomato leaves that can elicit the induction of proteinase inhibitors. Upon further hydrolysis this pectic polysaccharide yields oligogalacturanan (degree of polymerisation from 2 to 6) that still possesses the proteinase inhibitor inducing activities. 

The classic pathway is triggered by the formation of factor Cl at IgG or IgM on the surface of microorganisms (left). Cl is an 18-part molecular complex with three different components (Clq, Clr, and Cls). Clq is shaped like a bunch of tulips, the flowers of which bind to the Fc region of antibodies (left). This activates Clr, a serine proteinase that initiates the cascade of the classic pathway. First, C4 is proteolytically activated into C4b, which in turn cleaves C2 into C2a and C2b. C4B and C2a together form C3 convertase [1], which finally catalyzes the cleavage of C3 into C3a and C3b. Small amounts of C3b also arise from non-enzymatic hydrolysis of C3. 

Latent forms of a neutral protease and an acid protease (pH optimum 5.3) from cartilage are activated by trypsin hydrolysis to give Ca2+-dependent enzymes that catalyze the hydrolysis of proteoglycan.406 Some Ca2+-dependent proteinases are isolated as proenzymes that can be converted to the active form by high [Ca2+] or by low [Ca2+] in the presence of a digestable substrate.407... 

Crystallographic studies of native cysteine proteinases and enzyme-inhibitor complexes have been used to interpret much or the kinetic data for cysteine protemsse-caUlyzed hydrolysis of amide bonds. Analysis of the crystal structures of papain [16]. caricain [38], actinidain [56], etc. shows that these structures are closely related. The active site of all these cysteine proteinases contains the Cys-25 sulfhydryl group in close proximity to the His-159 imidazole ring nitrogens, where the latter can abstract the sulfhydryl proton to facilitate attack on the substrate amide carbonyl group [17]. 

Boronic acids (5a) were among the first examples of low-molecular-weight, reversible inhibitors of serine proteinases [151, 152]. Significant inhibition was initially demonstrated against a-chymotrypsin. Unlike the carbonyl-derived reversible inhibitors, which require a polypeptide or peptide-like chain, activity was found with simple alkylboronic acids (e.g. the value for PhCH2CH2B(OH)2 with a-chymotrypsin was = 40 //M) [153], Weak inhibition of elastase (PPE) was first reported for a series of arylboronic acids, for example, (10-1) [123]. Some of the boron-based inhibitors Figure 2.5) were tested as either the difluoroboranes (5b) or as the pinacol boronate esters (5c). These modifications were employed because they were more readily synthesized and/or purified than the boronic acids. For both of these derivatives inhibition was shown to be due to in situ hydrolysis to the parent boronic acid (5a) [154, 155]. 

The first nonpeptide, irreversible inactivators of serine proteinases were phosphoryl fluoride derivatives such as diisopropylphosphofluoridate (see Figure 2.7) [168]. It was shown that the phosphofluoridates stoichiometri-cally inactivate serine proteinases, by coupling with the active-site serine hydroxyl group. They have since been used as a diagnostic tool for the classification of serine proteinases. Similarily, both aryl and aralkyl sulphonyl fluorides were determined to be inactivators, forming a sulphonyl enzyme derivative at Ser-195 which is resistant to hydrolysis. 

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