ENZYME-POLYELECTROLYTE COMPLEXES IN WATER-ETHANOL MIXTURES - NEGATIVELY CHARGED GROUPS ARTIFICIALLY INTRODUCED INTO ALPHA-CHYMOTRYPSIN PROVIDE ADDITIONAL ACTIVATION AND STABILIZATION EFFECTS
Ev. Kudryashova et al., ENZYME-POLYELECTROLYTE COMPLEXES IN WATER-ETHANOL MIXTURES - NEGATIVELY CHARGED GROUPS ARTIFICIALLY INTRODUCED INTO ALPHA-CHYMOTRYPSIN PROVIDE ADDITIONAL ACTIVATION AND STABILIZATION EFFECTS, Biotechnology and bioengineering, 55(2), 1997, pp. 267-277
Formation of noncovalent complexes between alpha-chymotrypsin (CT) and
a polyelectrolyte, polybrene (PB), has been shown to produce two majo
r effects on enzymatic reactions in binary mixtures of polar organic c
osolvents with water. (i) At moderate concentrations of organic cosolv
ents (10% to 30% v/v), enzymatic activity of CT is higher than in aque
ous solutions, and this activation effect is more significant for CT i
n complex with PB (5- to 7-fold) than for free enzyme (1.5- to 2.5-fol
d). (ii) The range of cosolvent concentrations that the enzyme tolerat
es without complete loss of catalytic activity is much broader. For en
hancement of enzyme stability in the complex with the polycation, the
number of negatively charged groups in the protein has been artificial
ly increased by using chemical modification with pyromellitic and succ
inic anhydrides. Additional activation effect at moderate concentratio
ns of ethanol and enhanced resistance of the enzyme toward inactivatio
n at high concentrations of the organic solvent have been observed for
the modified preparations of CT in the complex with PB as compared wi
th an analogous complex of the native enzyme. Structural changes behin
d alterations in enzyme activity in water-ethanol mixtures have been s
tudied by the method of circular dichroism (CD). Protein conformation
of all CT preparations has not changed significantly up to 30% v/v of
ethanol where activation effects in enzymatic catalysis were most pron
ounced. At higher concentrations of ethanol, structural changes in the
protein have been observed for different forms of CT that were well c
orrelated with a decrease in enzymatic activity. (C) 1997 John Wiley &
Sons, Inc.