Number and function of sulphydryl groups of N-acetylneuraminate lyase

The reaction between DTNB and the SH groups of N-acetylneuraminate lyase has been investigated in the presence and absence of pyruvic acid, substrate of the enzyme. It was found that DTNB inactivates N-acetylneuraminate lyase, while pyruvic acid protects the enzyme against this inactivation. When the enzyme was fully inactivated, two SH groups have reacted with DTNB. This result supports previous suggestions, that there is one cystein residue per active site responsible for enzyme activity. In the presence of SDS, approx. 6 SH groups reacted with DTNB suggesting the existence of 3 SH groups per enzyme subunit.