Studies on the Essential Groups of the Alkaline Phosphatase from Ostrea cucullate

The alkaline phosphatase from Ostrea cucullate were selectively modified by DTT,PCMB,NBS,acetic anhydride,succinic anhydride and formaldehyde,after that the changes in their activities have been detected. The enzyme was modified by PCMB in the concentrate of 0.4 mmol·L~(-1) for 30 min, and the enzyme activity was not influenced, indicating that the -SH group is not essentical to the enzyme's faction. Modification of dithiothreitol (DTT) on the enzyme resulted in 98% inhibition of the enzyme activity in the concentration of 2.5 mmol·L~(-1) of DTT. The result shows that the disulfide bonds are essential to the enzyme activity.By chemical modification with N-bromosuccinimide,the enzyme can be totally inactivated,followed by the UV absorption spectrum change,suggesting that the residue of tryptophan is essential for the activity and may be at the active site of the enzyme.The results of the chemical modification of the enzyme by acetic anhydride, succinic anhydride and formaldehyde demonstrate that the amino group is one of the enzyme's functional groups. Studies of inactivation of the enzyme by chemical modification demonstrated that Ser,Lys and Trp residues are essential functional groups of the enzyme.Partial disulfide bonds are essential for the function of this enzyme.