Activity Measurements of Proteinases Using Synthetic Substrates

In this assay, enzymes with chymotrypsin-like specificity readily bind the side chain of Phe at the primary substrate binding site (SI) and subsequently hydrolyze the adjacent amide bond linking the Phe residue to the p-NA moiety. On cleavage, the release of p-NA is measured by the increase in absorbance at 410 nm (e4l0 = 8480 M 1 cm-1) with time, using a recording spectrophotometer. 

Amino-group substituents Carboxyl-group substituents 

Activity Measurements of Proteinases Using Synthetic Substrates 

Brachyurin Serine Suc-AlaAlaProPhe-p-NA Suc-AlaAlaProArg-p-NA Suc-AlaAlaProPhe-SBzl Suc-AlaAlaProArg-SBz.l 

Cathepsin B Cysteine Z-Arg-p-NA Z-ArgArg-p-NA Z-PheArg-p-NA Z-ArgArg-AMC Z-PheArg-AMC 

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C2.1 Activity Measurements of Proteinases Using Synthetic Substrates C2.2 Peptidase Activity Assays Using Protein Substrates... 

The observed rate of production R is equal to the formation rate minus the enzyme decay rate, where X is biomass concentration, S is the concentration of the limiting substrate, E is the activity of the enz3rme, jj is the specific growth rate and K, K, K, Kg are unknown parameters which have to be estimated from experimental data With both a-amylase and proteinase production the cultivation was performed on synthetic media where the concentration of limiting substrate were known and measurable By contrast, in most exocellular enzyme productions complex media with an unknown concentration of unidentified limiting substrates are used The model can be simplified when the mass balance for the limiting substrate S is applied as follows ... 

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