CHAOTROPIC AGENTS CAUSE DISAGGREGATION AND ENHANCED ACTIVITY OF PASTEURELLA-HAEMOLYTICA LEUKOTOXIN

Pasteurella haemolytica biotype A, serotype 1 grown to late logarithmi c growth phase in cell culture medium (RPMI 1640) produced highly aggr egated leukotoxin. The multimer mass of the highly aggregated leukotox in in 0.1 M sodium phosphate buffer pH 7.0 as determined by gel filtra tion chromatography on Sephacryl S400HR was approximate to 8000 kDa. R esuspension of leukotoxin in phosphate buffer containing various chaot ropic agents resulted in partial disaggregation of leukotoxin and enha nced leukotoxic activity. 3M guanidine disaggregated leukotoxin to a m ultimer mass of approximate to 800 kDa and enhanced leukotoxic activit y 3 to 20-fold. Tn 6 M urea or 1 M sodium thiocyanate, leukotoxin mult imers were observed ranging in mass from 8000 kDa to 400 kDa, and acti vity enhancement was less than that for leukotoxin in 3 M guanidine. S everal detergents were tested for enhancement of leukotoxic activity, but only 1% Tween 20 enhanced leukotoxic activity (4-fold), whereas 1. 25% octylglucoside, 10 mM CHAPS, and 5 mM deoxycholate diminished and 1% Triton X-100 abolished leukotoxic activity.