Furylacryloyl group

Chromophoric acyl group,4,5 The spectrum of the furylacryloyl group depends on the polarity of the surrounding medium, and also on the nature of the moiety to which it is attached. The spectrum of furylacryloyl-L-tyrosine ethyl ester changes slightly when it is bound to chymotrypsin. There are also further changes on formation of the acylenzyme and on the subsequent hydrolysis. The rate constants for acylation and deacylation and the dissociation constant of the Michaelis complex may be measured by the appropriate experiments. 

When the ester is mixed with the enzyme, there is an initial change in absorbance that is due to the formation of the Michaelis complex. The rate constant for this is beyond the time scale of stopped flow, but the magnitude of the change can be used to calculate the dissociation constant. The absorbance then changes exponentially as the acylenzyme accumulates. There are further changes in the spectrum of the furylacryloyl group as the ester is gradually hydrolyzed to the free acid. 

Use of the proflavin displacement method is far more convenient than use of the furylacryloyl group, since no special substrates have to be synthesized and one readily available compound can be used with all substrates. In general, it is better not to use modified substrates not only are they chemically inconvenient to synthesize, but they are always open to criticism on the grounds that the results could be artifacts. 

There remains model 4, and MacQuarrie and Bernhard 175) have utilized the full-site reactivity by iodoacetamide and half-site reactivity by FAP to provide support for this model. Thus, di(2-furylacryloyl) enzyme was prepared, and the two remaining sites were blocked with iodoacetate. Acyl groups were then removed from this derivative by arsenolysis, and the resulting dialkyl enzyme was tested for stoichiometry with FAP. Only one acyl group could be incorporated into the dialkyl enzyme. This result cannot be explained in terms of an induced asymmetry model, and indeed, can only be explained by a preexisting asymmetry model if there is a subunit rearrangement. In addition, alkylation of the enzyme with varying quantities of iodoacetate, followed by acylation of these derivatives with FAP, showed a 2 1 ratio of alkylation to acylation, independent... 

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