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J. Bacteriol., Jan 1998, 152-158, Vol 180, No. 1
MJ Sadowsky, Z Tong, M de Souza and LP Wackett
Pseudomonas sp. strain ADP metabolizes atrazine to cyanuric acid via three
plasmid-encoded enzymes, AtzA, AtzB, and AtzC. The first enzyme, AtzA,
catalyzes the hydrolytic dechlorination of atrazine, yielding
hydroxyatrazine. The second enzyme, AtzB, catalyzes hydroxyatrazine
deamidation, yielding N-isopropylammelide. In this study, the third gene in
the atrazine catabolic pathway, atzC, was cloned from a Pseudomonas sp.
strain ADP cosmid library as a 25-kb EcoRI DNA fragment in Escherichia
coli. The atzC gene was further delimited by functional analysis following
transposon Tn5 mutagenesis and subcloned as a 2.0-kb EcoRI-AvaI fragment.
An E. coli strain containing this DNA fragment expressed
N-isopropylammelide isopropylamino hydrolase activity, metabolizing
N-isopropylammelide stoichiometrically to cyanuric acid and
N-isopropylamine. The 2.0-kb DNA fragment was sequenced and found to
contain a single open reading frame of 1,209 nucleotides, encoding a
protein of 403 amino acids. AtzC showed modest sequence identity of 29 and
25%, respectively, to cytosine deaminase and dihydroorotase, both members
of an amidohydrolase protein superfamily. The sequence of AtzC was compared
to that of E. coli cytosine deaminase in the regions containing the five
ligands to the catalytically important metal for the protein. Pairwise
comparison of the 35 amino acids showed 61% sequence identity and 85%
sequence similarity. AtzC is thus assigned to the amidohydrolase protein
family that includes cytosine deaminase, urease, adenine deaminase, and
phosphotriester hydrolase. Similar sequence comparisons of the most highly
conserved regions indicated that the AtzA and AtzB proteins also belong to
the same amidohydrolase family. Overall, the data suggest that AtzA, AtzB,
and AtzC diverged from a common ancestor and, by random events, have been
reconstituted onto an atrazine catabolic plasmid.
Copyright © 1998, American Society for Microbiology
AtzC is a new member of the amidohydrolase protein superfamily and is homologous to other atrazine-metabolizing enzymes [In Process Citation]
Department of Microbiology, Institute for Advanced Studies in Biological Process Technology, University of Minnesota, St. Paul 55108, USA. Sadowsky@soils.umn.edu
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