Poly(ethyleneimine) stabilizes proteins during long-time storage and has been shown to be very effective in preventing oxidation and aggregation of lactate dehydrogenase [1, 2]. Due to its cationic character it is likely that it can interact with the protein molecule and form a complex, which could explain part of its stabilizing effect. Interactions between poly(ethyleneimine) (PEI) and porcine muscle lactate dehydrogenase (LDH) were therefore studied using static and dynamic light scattering and intrinsic fluorescence spectroscopy. Time-related aggregation of the enzyme was significantly reduced in the presence of the polymer. PEI of molecular weights 2000 and 25 000 were found to complex with the enzyme without significant change in the particle hydrodynamic radius, i.e. the polymer is bound in a flat conformation at the enzyme surface. Interactions with the high molecular weight PEI (2.6*106), on the other hand, resulted in a particle of a size significantly larger than the polymer, indicating that several LDH molecules may bind within the large, branched, polymer structure.

Although it was not found possible to quantitatively determine the degree of binding, estimation of the binding constants by intrinsic fluorescence measurements revealed weak binding between the enzyme and the polymer.

The observed effect of PEI on aggregation probably results from coating of the enzyme surface by PEI molecules to yield a polycationic species; this inhibits protein-protein contacts by electrostatic repulsion which in turn hinders aggregation. The presence of PEI in the close proximity of the enzyme surface could also hinder or slow down deleterious inactivation processes by decreasing the mass transfer rates and also hinder oxygen transfer and oxidation.

[1] Andersson, M. M. and Hatti-Kaul, R., J. Biotechnol. 72, 21-31, (1999)

[2] Andersson, M. M.; Breccia J. D. and Hatti-Kaul R. (1999) (not published)