Effect of Trinitrophenylation of Amino Groups on Biological Activities of Ricin D

Abstract

Modification of amino groups with 2,4, ii-trinitrohenzenesulfonic acid (TN%) remarkably reduced the toxicity of ricin D to mice, whereas cy toagglu t ina t ing and carbohydra te -b inding ac t iv i t i es were fu l ly re ta ined unt i l the modi f ica t ion of 8 m o l e s ( 7 3 96) of the f ree amino groups , indicating that the free amino groups exposed on the surface of ricin D molecule contribute to the toxic action but are not essential for the binding to oligosaccharides on cell surface. Three trinitrophenyl (TNP) amino groups out of eight in ricin D were found to be distributed to the A chain and five TNP-amino groups were deduced to be distributed to the B chain. The inh ib i tory ac t iv i ty of the TNP-r ic in D to ce l l f ree pro te in synthes is was more than 50% of that of native ricin D, indicating that amino groups in the A chain do not contribute much to the inhibition of protein synthesis. Digestibility of the TNP-r ic in D by protease and i t s incorpora t ion in to ce l l s a l so were the same as the native ricin D. T h e s e r e s u l t s d e m o n s t r a t e t h a t t h e t r i n i t r o p h e n y l a t i o n i s o n e o f t h e u s e f u l means of reducing the toxicity of ricin D without impairing the B chain’s ability to bind to the cell surface and the A chain’s ability to inhibit protein synthesis. A possible contribution of amino groups to the transport mechanism of ricin D to the cytoplasm are discussed in the results.