Studies on Conformational Changes in Glyceraldehyde-3-Phosphate Dehydrogenase Accompanying its Catalytic Action.

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Abstract
The optical rotatory dispersion of glyceraldehyde-3-phosphate dehydrogenase apoenzyme, holoenzyme (with NAD), and G3P-hemimercaptalenzyme were measured in the range: 300 [long dash]700 m[mu] at pH 7.7[long dash]8.4 at 23[degree]C in 0.15 M salt solution (0.001 M in EDTA). The dispersion parameters [lambda]c, K and b0 were calculated and used to estimate the helical content. A tentative interpretation of the observed changes in dispersion parameters in terms of conformational and other changes were made. The helical content (as represented by b0) of the apoenzyme is pH-dependent in the range: pH 7.7[long dash]8.4, but the solvation (K) is not. Even a low concentration (6 x 10-4 [image]) of the nonpolar compound a -thioglycerol affects both the helical content (b0) and the solvation of the apoenzyme (K). Stoichiometric binding of the cofactor NAD to the apoenzyme alters the helical content (b0), but not the solvation. Addition of excess NAD causes no further changes except in the amplitude of the anomalous rotation (at 650 m[mu]). Formation of the substrate G3p-enzyme complex from the apoenzyme changes both the helical content (b0) and the solvation (K). Anomalous rotation was observed in two wavelength ranges: one at about 305 m[mu], the other at about 650 m[mu]. The reference curve indicated that the former was due to a Zn-complex (probably a Cotton effect), while the latter might arise from an interaction involving the sulphur atom of cysteine.