title: Rescue of the orphan enzyme isoguanine deaminase.
abstract: Cytosine deaminase (CDA) from Escherichia coli was shown to catalyze the deamination of isoguanine (2-oxoadenine) to xanthine. Isoguanine is an oxidation product of adenine in DNA that is mutagenic to the cell. The isoguanine deaminase activity in E. coli was partially purified by ammonium sulfate fractionation, gel filtration, and anion exchange chromatography. The active protein was identified by peptide mass fingerprint analysis as cytosine deaminase. The kinetic constants for the deamination of isoguanine at pH 7.7 are as follows: k(cat) = 49 s(-1), K(m) = 72 μM, and k(cat)/K(m) = 6.7 × 10(5) M(-1) s(-1). The kinetic constants for the deamination of cytosine are as follows: k(cat) = 45 s(-1), K(m) = 302 μM, and k(cat)/K(m) = 1.5 × 10(5) M(-1) s(-1). Under these reaction conditions, isoguanine is the better substrate for cytosine deaminase. The three-dimensional structure of CDA was determined with isoguanine in the active site.
authors: Department of Biochemistry and Biophysics and Department of Chemistry, Texas A&M University, College Station, Texas 77843, USA.
authors: Daniel S
authors: DS
authors: Hitchcock
authors: Alexander A
authors: AA
authors: Fedorov
authors: Elena V
authors: EV
authors: Fedorov
authors: Lawrence J
authors: LJ
authors: Dangott
authors: Steven C
authors: SC
authors: Almo
authors: Frank M
authors: FM
authors: Raushel
references: J Bacteriol. 1992 Oct;174(20):6321-5
references: 1328155
references: Biochemistry. 2006 Feb 21;45(7):1997-2005
references: 16475788
references: J Am Chem Soc. 2011 Feb 23;133(7):2080-3
references: 21275375
references: Nat Protoc. 2007;2(10):2366-82
references: 17947979
references: FEBS Lett. 1996 Aug 5;391(1-2):113-6
references: 8706896
references: J Bacteriol. 1988 Nov;170(11):5405-7
references: 3053667
references: J Bacteriol. 1998 Jan;180(1):152-8
references: 9422605
references: Nucleic Acids Res. 1997 Jan 15;25(2):304-11
references: 9016558
references: PLoS One. 2009;4(2):e4345
references: 19190775
references: Biochemistry. 2010 May 25;49(20):4374-82
references: 20415463
references: Mol Syst Biol. 2006;2:2006.0008
references: 16738554
references: Mol Microbiol. 1992 May;6(10):1335-44
references: 1640834
references: Biochemistry. 2005 May 3;44(17):6383-91
references: 15850372
references: Carcinogenesis. 1983 Aug;4(8):997-1000
references: 6409445
references: Arch Biochem Biophys. 1951 Jun;32(1):227-9
references: 14847690
references: Nature. 2007 Aug 16;448(7155):775-9
references: 17603473
references: Nature. 2007 Jun 21;447(7147):941-50
references: 17581577
references: Int J Radiat Biol. 2002 Jul;78(7):585-92
references: 12079537
references: Biochemistry. 1992 Jan 14;31(1):106-10
references: 1731864
references: Nucleic Acids Res. 1991 Jul 11;19(13):3629-32
references: 1649454
references: Biochemistry. 1997 Sep 16;36(37):11125-30
references: 9287155
references: J Mol Biol. 2002 Jan 25;315(4):687-97
references: 11812140
references: Proc Natl Acad Sci U S A. 2007 May 8;104(19):7821-6
references: 17483465
references: J Am Chem Soc. 2010 Feb 17;132(6):1762-3
references: 20088583
references: Nature. 1992 Jan 16;355(6357):273-5
references: 1309939
references: Nucleic Acids Symp Ser. 1995;(34):233-4
references: 8841637
references: Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):7022-5
references: 1495996