ABSTRACT
The use of pH indicators to monitor hydrolase-catalyzed reactions is described. The formation of acid following an enzyme-mediated hydrolysis causes a drop in the pH that can be visualized by a change in the color of the indicator-containing solution. The best indicators are those showing a color transition within the operational pH range of the hydrolases, like bromothymol blue and phenol red. The enantioselectivity of lipases and esterases can be estimated using single isomers under the same conditions and comparing the color turnover for each one. The method has been tested to quickly evaluate the enantioselectivity of a lipase towards a set of ester substrates and applied to the hierarchical screening of a library of thermophilic esterases.
Subject(s)
Chemistry, Organic/methods , Enzymes/metabolism , Hydrolases/chemistry , Indicators and Reagents/chemistry , Acetonitriles/chemistry , Acetonitriles/metabolism , Bromthymol Blue/chemistry , Burkholderia cepacia/enzymology , Catalysis , Chemistry, Organic/instrumentation , Colorimetry/methods , Enzymes/chemistry , Esterases/chemistry , Esterases/metabolism , Hydrogen-Ion Concentration , Hydrolases/metabolism , Peptide Library , Phenolsulfonphthalein/chemistry , Phosphates/chemistry , Potassium Compounds/chemistry , Sensitivity and Specificity , StereoisomerismABSTRACT
The unique ways in which enzymes are differentiated from other catalysts translate into special advantages. Understanding these advantages is the key toward better matching of biocatalysts needs in industrial chemistry. Specific cases where enzymes and biotransformations have been used successfully at the production scale are examined, permitting the realities of using biocatalysts to be separated from the misconceptions and myths. Five such misconceptions will be examined in the context of examples of some commercially-successful biocatalytic processes.
