Trypsin turnover number

The results of experiments in which the mutation was made were, however, a complete surprise. The Asp 189-Lys mutant was totally inactive with both Asp and Glu substrates. It was, as expected, also inactive toward Lys and Arg substrates. The mutant was, however, catalytically active with Phe and Tyr substrates, with the same low turnover number as wild-type trypsin. On the other hand, it showed a more than 5000-fold increase in kcat/f m with Leu substrates over wild type. The three-dimensional structure of this interesting mutant has not yet been determined, but the structure of a related mutant Asp 189-His shows the histidine side chain in an unexpected position, buried inside the protein. 

Turnover numbers of enzymes vary from <1 to 106 s . Trypsin, chymotrypsin, and many intracellular... 

There are also examples of nanoreactors which have increased enzyme activity when encapsulated compared with bulk conditions. PICsomes that are stable to salt and elevated temperatures have been successfully developed using PEG-h-P( Asp) and are able to encapsulate trypsin without altering the original enzymatic activity [25]. Furthermore, these specifically designed PICsomes maintained their vesicular architecture at salt concentrations of 300 mM NaCl and temperatures up to 70°C, with only a subtle inaease in their size. For example, encapsulated nucleoside hydrolase has a substrate turnover number (kat) up to six times higher than the free enzyme [59]. Other enzymes, such as GOx and HRP, showed an 85% inaease in activity afta encapsulation in polyelectrolyte capsules [112], In addition, laccase entrapped in linear-dendritic based nanoreactors showed superior activity compared with its activity in bulk, and also inaeased temperature stability (up to 70 C) [19]. Howeva, the intrinsic branched architecture of den-drimers offers insufficient fluidity and space for entrapping larger biomolecules. 

Figure 5. Turnover numbers of the trypsin-catalyzed hydrolysis of trimethylethoxysilane and condensation of trimethylsilanol at 10 C.

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