Melamine deaminase and atrazine chlorohydrolase: 98 percent identical but functionally different.
The gene encoding melamine deaminase (TriA) from Pseudomonas sp. strain NRRL B-12227 was identified, cloned into Escherichia coli, sequenced, and expressed for in vitro study of enzyme activity. Melamine deaminase displaced two 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 second deamination reaction, suggesting that the lower rate of ammeline hydrolysis was due to differential substrate turnover rather than product inhibition. Remarkably, melamine deaminase is 98% identical to the enzyme atrazine chlorohydrolase (AtzA) from Pseudomonas sp. strain ADP. Each enzyme consists of 475 amino acids and differs by only 9 amino acids. AtzA was shown to exclusively catalyze dehalogenation of halo-substituted triazine ring compounds and had no activity with melamine and ammeline. Similarly, melamine deaminase had no detectable activity with the halo-triazine substrates. Melamine deaminase was active in deamination of a substrate that was structurally identical to atrazine, except for the substitution of an amino group for the chlorine atom. Moreover, melamine deaminase and AtzA are found in bacteria that grow on melamine and atrazine compounds, respectively. These data strongly suggest that the 9 amino acid differences between melamine deaminase and AtzA represent a short evolutionary pathway connecting enzymes catalyzing physiologically relevant deamination and dehalogenation reactions, respectively.
Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, St. Paul, Minnesota 55108, USA.
J L
JL
Seffernick
M L
ML
de Souza
M J
MJ
Sadowsky
L P
LP
Wackett
D000595:Amino Acid Sequence; D000619:Aminohydrolases; D001667:Binding, Competitive; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006867:Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D008969:Molecular Sequence Data; D011506:Proteins / Q000235:genetics* / Q000378:metabolism; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D014227:Triazines / Q000378:metabolism*
D011506:Proteins; D014227:Triazines; D006867:Hydrolases; D000619:Aminohydrolases; C424706:melamine deaminase; C100894:atrazine chlorohydrolase; C011907:melamine