Lactate dehydrogenase in interpolyelectrolyte complex. Function and stability

A new method for encapsulating enzymes by multilayer polyelectrolyte coatin g is proposed. The method,consists in a stepwise adsorption of polyelectrol ytes from solution onto protein aggregates formed by salting out the protei ns in highly concentrated salt solutions. Polystyrene sulfonate and fluores cence-labeled polyalylamine were used for capsule formation. The size of la ctate dehydrogenase aggregates covered by four layer pairs of electrolytes was 1-5 mu m, as indicated by fluorescence microscopy. The catalytic charac teristics and stability of pig muscle lactate dehydrogenase (EC 1.1.1.13) i ncapsulated in multilayer electrolyte complex obtained by this method were studied. It was found that the affinity of the substrate pyruvate for the e nzyme in the polyelectrolyte complex (K-M) did not essenstially change as c ompared with the free enzyme. Incapsulated lactate dehydrogenase showed the following features that distinguish it from the free form: (1) the lifetim e in diluted solutions increases from 30 min (without capsules) to 1-2 days tin capsules); (2) a higher stability to basic denaturation (up to pH 10); and (3) the absence of substrate inhibition of enzyme in the polyelectroly te complex. The changes in the catalytic characteristics of incapsulated la ctate dehydrogenase are discussed in terms of an increase in effective pK v alues of amino acid perturbed by polyelectrolyte coating of enzyme.