Protease formats

Coagulation Factors II, III, VII, IX, X, XI, and Xlla fragments, thrombin, and plasmin are classified as serine proteases because each possesses a serine residue with neighboring histidine and asparagine residues at its enzymatically active site (Table 3). Factors II, VII, IX, and X, Protein C, Protein S, and Protein Z are dependent on the presence of vitamin K [84-80-0] for their formation as biologically functionally active procoagulant glycoproteins. 

Protein G. This vitamin K-dependent glycoproteia serine protease zymogen is produced ia the Hver. It is an anticoagulant with species specificity (19—21). Proteia C is activated to Proteia by thrombomodulin, a proteia that resides on the surface of endothefial cells, plus thrombin ia the presence of calcium. In its active form, Proteia selectively iaactivates, by proteolytic degradation. Factors V, Va, VIII, and Villa. In this reaction the efficiency of Proteia is enhanced by complex formation with free Proteia S. la additioa, Proteia activates tissue plasminogen activator, which... 

Protein residues, eg, soft-boiled egg yolk, are difficult stains to handle. If the stains are not totally denatured, proteases can decompose them. There are commercial proteases with a high temperature optimum (60°C) that can remove most protein soils in a dishwasher (63). Patents on the use of Upases in ADDs have claimed that Upases can reduce the formation of spots and films on glasses (62,64—66) however, no commercial appUcation of Upases in ADDs has been implemented. 

BLOOD CLOTTING. The formation of blood clots is the result of a series of zymogen activations (Figure 15.5). The amplification achieved by this cascade of enzymatic activations allows blood clotting to occur rapidly in response to injury. Seven of the clotting factors in their active form are serine proteases ... 

Tissue-type plasminogen activator (tPA) is a glycoprotein (68 kDa), synthesized by endothelial and tumor-cells. As a serine protease, tPA hydrolyses Arg561-Val562 peptide bond in plasminogen, resulting in plasmin formation. It needs cofactors for efficient plasminogen activation. 

CarriUo A, Stewart KD, Sham HL, Norbeck DW, Kohlbrenner WE, Leonard JM, Kempf DJ, Molla A (1998) In vitro selection and characterization of human immunodeficiency virus type 1 variants with increased resistance to ABT-378, a novel protease inhibitor. J Virol 72 7532-7541 Chambers TJ, Nestorowicz A, Amberg SM, Rice CM (1993) Mutagenesis of the yellow fever virus NS2B protein effects on proteolytic processing, NS2B-NS3 complex formation, and viral replication. J Virol 67 6797-6807... 

Lipases are the enzymes for which a number of examples of a promiscuous activity have been reported. Thus, in addition to their original activity comprising hydrolysis of lipids and, generally, catalysis of the hydrolysis or formation of carboxylic esters [107], lipases have been found to catalyze not only the carbon-nitrogen bond hydrolysis/formation (in this case, acting as proteases) but also the carbon-carbon bond-forming reactions. The first example of a lipase-catalyzed Michael addition to 2-(trifluoromethyl)propenoic acid was described as early as in 1986 [108]. Michael addition of secondary amines to acrylonitrile is up to 100-fold faster in the presence of various preparations of the hpase from Candida antariica (CAL-B) than in the absence of a biocatalyst (Scheme 5.20) [109]. 

A number of iron-containing, ascorbate-requiring hydroxylases share a common reaction mechanism in which hydroxylation of the substrate is linked to decarboxylation of a-ketoglutarate (Figure 28-11). Many of these enzymes are involved in the modification of precursor proteins. Proline and lysine hydroxylases are required for the postsynthetic modification of procollagen to collagen, and prohne hydroxylase is also required in formation of osteocalcin and the Clq component of complement. Aspartate P-hydroxylase is required for the postsynthetic modification of the precursor of protein C, the vitamin K-dependent protease which hydrolyzes activated factor V in the blood clotting cascade. TrimethyUysine and y-butyrobetaine hydroxylases are required for the synthesis of carnitine. 

Because of their very complex chemical structures and heterogeneity, melanins are difficult to extract, separate, and characterize from tissues. Eumelanins are insoluble in water and organic solvents. They can be extracted from tissues with strong chemicals that are capable of removing lipids, proteins, and other tissue components but also lead to the formation of degradation products. Enzymatic procedures were developed for the isolation of eumelanins from mammalian hair and irises. The first step is sequential digestion with protease, proteinase K, and papaine in the presence... 

To check if PemB is surface exposed, E. chrysanthemi cells were subjected to proteolysis. Treatment of the cell suspension with trypsin, proteinase K or chimotrypsin at a concentration of 0.1 to 1 mg/ml for 1 h did not cause PemB proteolysis or its liberation into the medium. Cell pre-treatment with EDTA-lysozyme, which renders the periplasmic proteins accessible to proteases, gave no effect. PemB was also resistant to proteolytic digestion in extract of cells disrupted by sonication or in a French press. Only addition of Triton X-100 (up to 0.1%) causing formation of the micelles with PemB lead to a quick proteolyis of this protein (data not shown). In another approach to analyse the PemB exposition, bacterial cells were labelled with sulfo-NHS-biotin. This compound is unable to cross membranes and biotinylation... 

Although a few mechanisms have so far been proposed to explain the antimicrobial properties exhibited by proanthocyanidins (e.g., inhibition of extracellular enzymes) [86], Jones et al. [83] postulated that their ability to bind bacterial cell coat polymers and their abihty to inhibit cell-associated proteolysis might be considered responsible for the observed activity (Table 1). Accordingly, despite the formation of complexes with cell coat polymers, proanthocyanidins penetrated to the cell wall in sufficient concentration to react with one or more ultra-structural components and to selectively inhibit cell wall synthesis. Decreased proteolysis in these strains may also reflect a reduction of the export of proteases from the cell in the presence of proanthocyanidins [83]. 

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