Production and subsequent second-order decomposition of protein disulfide anions: Lengthy collisions between proteins

Abstract

By a technique of radiation chemistry, one can convert small groups on a protein (surface disulfide bonds) into reactive centers (disulfide anion radicals) in less than 100 μs. The reactive centers on different molecules then proceed to react with each other and are destroyed. The anion radical is formed by electron transfer from transient CO 2 − generated by pulse radiolysis (Adams et al., 1972). The formation and decay of the disulfide anion radical were studied by monitoring its absorption at 410 nm. The transient disulfide anion absorption is a reliable qualitative but not quantitative test for the presence of surface disulfide groups. The area of this reactive center is very small compared to the total surface area of the protein. Nevertheless the rate of this reaction for 13 proteins ranges from 2% to 50% of the diffusion controlled rate of 3 × 10 9 m −1 s −1. Billiard ball type collisions during which the protein molecules briefly touch each other do not account for these fast rates. We conclude that the collision has substantial duration. A radially symmetric potential holds the molecules together but allows them to roll on each others' surfaces. The model of Sole & Stockmayer (1973) can be applied to this reaction, showing that the collision duration is typically 10 to 100 μs. This conclusion, valid for 12 of 13 proteins studied, seems to be very general. Whenever two proteins approach each other, attractive forces hold them together long enough for mutual rotation to bring the reactive groups into contact, a necessary but sometimes not sufficient condition for their reaction.