The interactions between model polycations and polyanions have been studied using the fluorescent dye acridine orange as a probe. The dye emission at 540 nm is quenched by the addition of polyanions, a process which is reversed by the addition of certain polycations. Techniques have been developed which enable a determination of the stoichiometry of polyelectrolyte complexes so formed between three polyanions (carboxymethylcellulose, K-carrageenan and poly(styrenesu1fonate)) and two polycations [poly(L-lysine hydrobromide) and poly(p-xylylviologen dibromide)) . A one to one pairing of acidic sites with basic groups was observed over a wide range of temperature, pH and ionic strength. Deviations from unit stoichiometry in the presence of multi-charged metal ions have been interpreted as due to conformational changes in the polyanion. Results from viscosity measurements are included which suggest the neutralisation of charged groups in the polyelectrolyte complex.
It was shown that both the enzymes pepsin and a-chymotrypsin will displace the anionic dye Eosin Y from complexes of the dye with the polycations protamine or polybrene. The dyerelease was monitored fluorimetrically and was found to be strongly time-dependent. At equilibrium, pepsin released all the bound dye from both polycation-dye complexes when the mole ratio of pepsin to base moles of polycation were 1 : 10 for protamine and 1 : 13 for polybrene. a-Chymotrypsin did not release all the bound dye, but appeared to form definite complexes with the polycations. The existence of the complexes between the enzymes and the polycations was correlated with the fact that the polycations are inhibitors of the enzymes. Additionally, it has been shown that pepsin releases Acridine Orange (AO) from a poly(viny1 sulfate) (PVS)-A0 complex. The mole ratio of pepsin to base moles of PVS was found to be 1 : 2,5. Cooperative binding constants were also evaluated for each dye-polyion system.
An investigation has been made of the interactions of the enzyme papain with the polycations protamine, polybrene, poly(L-lysine), spermine, spermidine and the neutral polymer polyvinylpyrrolidone (PVP). At low concentrations, each behaves as an inhibitor of the enzyme. As their concentrations increased above a certain level, the activity of the systems increased, and their inhibition of the enzyme appeared to be less pronounced. When acting by themselves in the presence of the substrate haemoglobin, each of the polycations was a weak proteolytic catalyst with a ranking of catalytic effectiveness of protamine greater than polybrene greater than poly(L-lysine) greater than polyvinyl-pyrrolidone greater than spermidine greater than spermine. This effect could explain the anomalous inhibition of papain by these polycations. The interaction of papain with dansyl protamine (DNSP) and the extent of complex formation were studied using a fluorescence polarization technique and the results showed that there was a strong interaction occurred. The strength of binding was assessed by determination of the critical electrolyte concentration (0.2 M, NaNO3). The stoichiometry of the DNSP-papain complex was found to be 63 base moles of DNSP to one mole of papain.
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