Chymotrypsin octane solution

Proteins were lyophilized directly in the reaction vessels. Only in the case of insulin at LpH 7.5 was it necessary to lyophilize the protein from a large volume and transfer it to the reaction vessel. Insulin (20 mg) was lyophilized from a solution of 1 mM sodium phosphate buffer (40 ml), pH 7.5 and 40 mM sodium metaborate buffer (1 ml), pH 10. Ribonuclease and a-chymotrypsin were lyophilized from a solution of 40 mM sodium phosphate buffer (1 ml), pH 7.5 and 40 mM sodium metaborate buffer (1 mi), pH 10. Anhydrous octane (2 ml) was added to the protein and the medium was sonicated until the protein was finely dispersed, at which time [ Cjiodomethane (100 gl) was added. To prevent the loss of... 

FIG. 6 Dependence of the first-order rate constant (keat) for deacylation of N-trans-cinnamoy -a-chymotrypsin ( ), as well as rotational frequency, v (O), and hyperfine splitting constant, a (A) for spin-labeled a-chymotrypsin on degree of hydration (water-to-AOT molar ratio) in the system AOT (0.1 M)-water-octane. Spin label 2,2,5,5-tetramethyl-4-iodoacetamidopyrrolidine-l-oxyl. Dashed lines show keat and a values in aqueous solution. (From Ref. 41.)... 

FIG. 7 Dependence on the water/surfactant molar ratio of (a) the maximal reaction rate normalized to the enzyme concentration, V/E, of a-chymotrypsin-catalyzed hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide, and (b) the rotational frequency, v, of the spin label in the active site of the enzyme in the system AOT-water/glycerol-octane. Water/glycerol volume ratios 1—100 0 2—80 20, 3—50 50, 4—20 80, 5—6 94. Dashed lines show V/Eq and v values in aqueous solution. (From Ref. 42.)... 

FIG. 11 Combination of two factors regulating enzyme catalytic activity variation in the surfactant concentration and addition of water-miscible organic solvents, (a) Peroxidase in the system AOT-water/glycerol-octane at water/glycerol volume ratios (O) 100 0 ( ) 20 80. (b) a-Chymotrypsin in the system AOT-water/glycerol-octane at water/glycerol volume ratios (O) 100 0 ( ) 6 94. Dashed lines show the catal5dic activities of the enzymes in aqueous solution. (From Ref. 44.)... 

Some examples of DKR based on racemization of secondary alkyl amine via Shiff base were shown in Scheme 5.9. Schiffbase formation of a-amino carboxylic esters significantly increases the acidity of the a-proton in comparison to that of the parent amino acid, thus enabling enantiomer-selective hydrolysis and ammonoly-sis through DKR [23]. For example, chymotrypsin catalyses the hydrolysis of a Schiff base of phenylalanine ethyl ester and an aromatic aldehyde (Scheme 5.9, Equation 5.6) [23b]. In this particular case, natural phenylalanine precipitates, leaving the aldehyde and unhydrolysed enantiomeric ester in solution. Addition of l,4-diazabicyclo[2.2.2]octane (DABCO) (as the Bronsted base) allows DKR to take place by promoting racemization of the Schiffbase. Similarly, Novozym 435... 

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