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.
Clemente-Jiménez
Josefa María
JM
Departamento de Química-Física, Bioquímica y Química Inorgánica Edificio C.I.T.E. I, Universidad de Almería, Almería 04120, Spain.
Martínez-Rogríguez
Sergio
S
Mingorance-Cazorla
Lydia
L
De La Escalera-Hueso
Santiago
S
Las Heras-Vázquez
Francisco Javier
FJ
Rodríguez-Vico
Felipe
F
D016960:Agrobacterium tumefaciens / Q000737:chemistry / Q000145:classification / Q000201:enzymology* / Q000235:genetics*; D000581:Amidohydrolases / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D000595:Amino Acid Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D003067:Coenzymes / Q000737:chemistry; D004789:Enzyme Activation; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000737:chemistry*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D013045:Species Specificity; D013379:Substrate Specificity; D013696:Temperature
D003067:Coenzymes; D008670:Metals; D011994:Recombinant Proteins; D000581:Amidohydrolases; C020047:dihydropyrimidinase