The kinetics of conformational changes of α₂‐macroglobulin determined by time resolved X‐ray solution scattering

Abstract

The rate of gross conformational change of a,-macroglobulin (a,M) during its proteinase trapping was directly determined for the first time using time-resolved X-ray solution scattering. Decrease of radius of gyration was observed under pseudo-first-order conditions with excess proteinases, which exhibited a monophasic timecourse. The rate constants were 0.5 f 0.1 s-' and 0.8 f 0.2 s-' for the reaction with chymotrypsin and trypsin, respectively. There was no concentration dependence of the observed rate constants. Therefore, the rate-limiting step of the gross conformational change was not the bimolecular encounter reaction between a,M and proteinases, which requires a new proposal of pre-trapping of proteinases before the gross conformational change.

Role of α2‐macroglobulin in regulating amyloid β‐protein neurotoxicity: protective or detrimental factor?

“…The reaction with endoproteases generates the activated α 2 M (α 2 M*), inducing a major conformational change in the inhibitor and exposing receptor recognition sites (Figler et al . 1991; Arakawa et al . 1994).…”

Stopped-Flow Apparatus for X-ray Scattering and XAFS

“…Stopped-flow X-ray scattering (SFXS) methods have been applied to the elucidation of protein folding (Eliezer et al, 1993;, subunit reorientation such as allosteric transitions (Kihara et al, 1987;Tsuruta et al, 1990Tsuruta et al, , 1994, conformational change (Arakawa et al, 1994), and subunit association and dissociation (Moody et al, 1980;Fowler et al, 1983;Inoko, Kihara & Koch, 1983;Kihara et al, 1987;Wang et al, 1989;Sato et al, 1994). In particular, it is useful to obtain X-ray scattering patterns of intermediate or transient states in the kinetic pathway.…”

“…Since then, protease trapping of macroglobulin (Arakawa et al, 1994) and coupling of ATP hydrolysis of proton ATPase with its dissociation into monomers (Sato et al, 1994) have been investigated.…”