The Journal of Biological Chemistry

In order to elucidate whether or not the spectral and circular dichroic changes observed during the biodegradative threonine deaminase reaction are caused by the Schiff base formation between the enzyme-bound pyridoxal phosphate and the dehydrated intermediate, the role of substituents and the absolute configuration of the 0 carbon of substrates was investigated using various analogs of L-threonine. The enzyme was found to catalyze the cr,p elimination of L-allothreonine, DL-fhreoand DL-erythro-@-hydroxynorvaline, DL-threoand DL-erythro-P-phenylserine, DL-P-hydroxyvaline, L-@-chlorobutyrine, and L-P-chloroalanine in addition to L-threonine and L-serine. When L-threonine or L-Pchlorobutyrine was used as substrate, the absorption maximum of the enzyme-bound pyridoxal phosphate shifted from 415 nm to 434 and 442 nm, respectively, but it remained unchanged when L-allothreonine was used as substrate. L-PChloroalanine also caused a bathochromic shift of the absorption maximum, but L-serine did not induce such a shift. The positive circular dichroism of the enzyme-bound pyridoxal phosphate at 415 nm disappeared upon the addition of L-threonine or L-&chloroalanine. On the other hand, L-Pchlorobutyrine did induce a negative circular dichroism during the reaction, and L-allothreonine slightly diminished the magnitude of the positive circular dichroism. These results indicate that the substituents as well as the absolute configuration of the fl carbon of the substrate participate in the spectral and circular dichroic changes observed during the reaction. Thus, the shift of the absorption maximum and the circular dichroic change at 415 nm appear to occur in the step prior to the p elimination reaction. This conclusion was confirmed by a kinetic analysis of the spectral changes.