R. Hegde et al., THE VARIANTS OF THE PROTEIN TOXINS ABRIN AND RICIN - A USEFUL GUIDE TO UNDERSTANDING THE PROCESSING EVENTS IN THE TOXIN TRANSPORT, European journal of biochemistry, 215(2), 1993, pp. 411-419
Kinetic data on inhibition of protein synthesis in thymocyte by three
abrins and ricin have been obtained. The intrinsic efficiencies of A c
hains of four toxins to inactivate ribosomes, as analyzed by k(i)-vers
us-concentration plots were abrin II, III > ricin > abrin I. The lag t
imes were 90, 66, 75 and 105 min at a 0.0744 nM concentration of each
of abrin I, II, III and ficin, respectively. To account for the observ
ed differences in the dose-dependent lag time, functional and structur
al variables of toxins such as binding efficiency of B chains to recep
tors and low-pH-induced structural alterations have been analyzed. The
association constants obtained by stopped flow studies showed that ab
rin-I (4.13 X 10(5) M-1 s-1) association with putative receptor (4-met
hylumbelliferyl-alpha-D-galactoside) is nearly two times more often th
an abrin III (2.6 X 10(5) M-1 s-1) at 20-degrees-C. Equilibrium bindin
g constants of abrin I and II to thymocyte at 37-degrees-C were 2.26 X
10(7) M-1 and 2.8 X 107 M-1 respectively. pH-induced structural alter
ations as studied by a parallel enhancement in 8-anilino-L-naphthalene
sulfonate fluorescence revealed a high degree of qualitative similari
ty. These results taken with a nearly identical concentration-independ
ent lag time (minimum lag of 41-42 min) indicated that the binding eff
iciencies and internalization efficiencies of these toxins are the sam
e and that the observed difference in the dose-dependent lag time is c
ausally related to the proposed processing event. The rates of reducti
on of inter-subunit disulfide bond, an obligatory step in the intoxica
tion process, have been measured and compared under a variety of condi
tions. Intersubunit disulfide reduction of abrin I is fourfold faster
than that of abrin II at pH 7.2. The rate of disulfide reduction in ab
rin I could be decreased 11-fold by adding lactose, compared to that w
ithout lactose. The observed differences in the efficiencies of A chai
ns, the dose-dependent lag period, the modulating effect of lactose on
the rates of disulfide reduction and similarity in binding properties
make the variants a valuable tool to probe the processing events in t
oxin transport in detail.