In support of the trap hypothesis. Chymotrypsin is not rigidly held in its complex with human .alpha.2-macroglobulin

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Abstract
Complexes (2:1) of chymotrypsin with human .alpha.2-macroglobulin have been prepared in the presence of 200 mM methylamine such that 90% of the chymotrypsin remains noncovalently bound to the .alpha.2-macroglobulin. Reaction of this complex with the active-site-directed spin-labeling reagent 4-[(ethoxyfluorophosphinyl)oxy]-2,2,6,6-tetramethylpiperidinyl-1-oxy results in nitroxide labeling of the active-site serine residue of the complexed chymotrypsin. Electron spin resonance (ESR) spectra of this complex were recorded at 275 K in buffer and at 263 K in 50% glycerol. At 263 K in 50% glycerol the spectrum is that expected for a rigid glass, whereas at room temperature the ESR spectrum shows that the chymotrypsin is only slightly immobilized compared with free spin-labeled chymotrypsin. These findings are discussed in relation to possible models of inhibition of protease activity by .alpha.2-macroglobulin. It is concluded that the trap mechanism of Barrett and Starkey [Barrett, A. J., and Starkey, P. M. (1973) Biochem. J. 133, 709-724] is the only model currently considered that can account for the present findings.