The gene encoding melamine deaminase (TriA) from Pseudomonas sp. strain NRR
L B-12227 was identified, cloned into Escherichia coli, sequenced, and expr
essed for in vitro study of enzyme activity. Melamine deaminase displaced t
wo of the three amino groups from melamine, producing ammeline and ammelide
as sequential products. The first deamination reaction occurred more than
10 times faster than the second. Ammelide did not inhibit the first or seco
nd deamination reaction, suggesting that the lower rate of ammeline hydroly
sis was due to differential substrate turnover rather than product inhibiti
on. Remarkably, melamine deaminase is 98% identical to the enzyme atrazine
chlorohydrolase (AtzA) from Pseudomonas sp. strain ADP. Each enzyme consist
s of 475 amino acids and differs by only 9 amino acids. AtzA was shown to e
xclusively catalyze dehalogenation of halo-substituted triazine ring compou
nds and had no activity with melamine and ammeline. Similarly, melamine dea
minase had no detectable activity with the halo-triazine substrates. Melami
ne deaminase was active in deamination of a substrate that was structurally
identical to atrazine, except for the substitution of an amino group for t
he chlorine atom. Moreover, melamine deaminase and AtzA are found in bacter
ia that grow on melamine and atrazine compounds, respectively. These data s
trongly suggest that the 9 amino acid differences between melamine deaminas
e and AtzA represent a short evolutionary pathway connecting enzymes cataly
zing physiologically relevant deamination and dehalogenation reactions, res
pectively.