Reactivating effect of some naturally occurring dithiol compounds on threonine deaminase

Abstract

The biodegradative threonine deaminases have been purified to homogeneity and crystallized from the extracts of Escherichia coli (E. coli) and Clostridium tetranomorphum, respectively. It has been demonstrated that deaminases are composed of four identical subunits, each monomer containing one mole of pyridoxal phosphate. The deamination reaction mechanism studied using the E. coli enzyme is compatible with the mechanism derived from the pyridoxal-catalyzed α,β-elimination reaction in the non-enzymic model system, although direct evidence for the participation of pyridoxal phosphate in the enzyme reaction is lacking. Spectral and circular dichroic changes of the enzyme-bound pyridoxal phosphate after addition of the substrate, L-threonine, were postulated to represent the formation of a complex between pyridoxal phosphate and the dehydrated reaction intermediate. Optical studies indicate that optical changes occur prior to the β-elimination reaction. Kinetic analyses have indicated that the enzyme exists as an inactive form under the conditions of these optical experiments. These phenomena could be interpreted as indicating that the conversion of this inactive enzyme into an active form occurs after addition of L-threonine. These optical changes are observed during the reaction and this conversion step appears to be the rate-limiting step of the overall reaction under the conditions employed.