Inactivation of Rhodanese by Pyridoxal 5′‐Phosphate

Abstract

Pyridoxal 5'-phosphate and other aromatic aldehydes inactivate rhodanese. This inactivation reaches higher extents if the enzyme is in the sulfur-free form. The identification of the reactive residue as an amino group has been made by spectrophotometric determination of the 5'-phosphorylated pyridoxyl derivative of the enzyme. The inactivation increases with pyridoxal 5'-phosphate concentration and can be partially removed by adding thiosulfate or valine. Prolonged dialysis against phosphate buffer also leads to the enzyme reactivation. The absorption spectra of the pyridoxal phosphate-rhodanese complex show a peak at 410 nm related to the Schiff base and a shoulder in the 330nm region which is probably due to the reaction between pyridoxal 5'-phosphate and both the amino and thiol groups of the enzyme that appear reasonably close to each other. The relationship between loss of activity and pyridoxal 5'-phosphate binding to the enzyme shows that complete inactivation pyridoxal 5'-phosphate.