Catalytic analysis of a recombinant D-hydantoinase from Agrobacterium tumefaciens.

The D-hydantoinase gene of a wild strain of Agrobacterium tumefaciens BQL9 had 99.78% nucleotide sequence identity with other available Agrobacterium genes. The resulting amino acid sequence showed two important substitutions affecting two alpha-helixes in the secondary structure of the protein. The union of Mn2+ to the protein was essential for activating the enzyme and was independent of the temperature. D-Hydantoinase only was inactivated in the presence of 70 mM EDTA and at over 40 degrees C. The enzyme showed both hydantoinase and pyrimidinase activities, but only with the D-enantiomers of the substrates. Activity was greater for substrates with apolar groups in the number 5 carbon atom of the hydantoin. The native structure of the N-terminal end of this D-hydantoinase proved to be indispensable to its enzymatic activity.