The conformational transition associated with the activation of chymotrypsinogen to chymotrypsin

Optical rotatory dispersion measurements in the spectral interval 217 to 500 m μ, coupled with circular dichroism spectra in the 215 to 325 m μ region, demonstrate that the enzyme, α-chymotrypsin, and the zymogen, chymotrypsinogen (A), possess different conformations. In the far ultraviolet, a negative Cotton effect (optical rotatory dispersion) and an ellipticity band (circular dichroism) suggest the existence of right-handed α-helical segments in chymotrypsin. The character and intensity of these bands are sufficiently different in chymotrypsinogen to allow the supposition that one consequence of activation may be an increase in helix content. At longer wavelengths, Cotton effects and ellipticity bands are observed which reveal substantial differences between the two forms of the protein in the nature of interactions with neighbors of at least several side chains, including tryptophan and cystine residues. The complex optical rotatory dispersion curves may be analyzed for approximate helical content by several conventional means. As estimated by the coefficient b 0 of the Moffitt-Yang equation, and by the absolute value of the rotation at 235 m μ, the helix contents are virtually identical at about 13%. Inspection of the form of the curves in the far ultraviolet reveals, however, that they depart measurably from those expected of simple mixtures of α-helices and random coils as observed in synthetic polyamino acids and proteins of high helix content. The uncertain background rotation, probably arising from optically active side-chain transitions, is less troublesome in the circular dichroism spectrum. The circular dichroism spectrum of chymotrypsin comprises four ellipticity bands, that of chymotrypsinogen three bands, both in the spectral interval 220 to 325 m μ. The negative circular dichroism band centered near 230 m μ in chymotrypsin is not discerned in chymotrypsinogen. Side-chain transitions give rise to ellipticity bands in each protein at 260, 290 and 300 m μ. All three bands are of greater magnitude in chymotrypsin than in chymotrypsinogen. The disulfide chromophore is a possible source of the 260 m μ band, while tryptophan residues are probably responsible for the two bands of longer wavelength. Since the band at 230 m μ. is about 6 m μ. displaced from the characteristic 224 m μ α-helix band, it is reasonable to infer that side-chain transitions overlap the main peptide (helical) transitions in this spectral region. Estimates of helicity from t