3O7U

Crystal structure of Cytosine Deaminase from Escherichia Coli complexed with zinc and phosphono-cytosine

Summary for 3O7U

• Entry DOI: 10.2210/pdb3o7u/pdb
• Related: 1K6W
• Descriptor: Cytosine deaminase, (2S)-1-[3-{[(2R)-2-hydroxypropyl]oxy}-2,2-bis({[(2R)-2-hydroxypropyl]oxy}methyl)propoxy]propan-2-ol, ZINC ION, ... (7 entities in total)
• Functional Keywords: (beta-alpha)8 barrel, hydrolase, hydrolase-hydrolase inhibitor complex, hydrolase/hydrolase inhibitor
• Biological source: Escherichia coli
• Total number of polymer chains: 1
• Total formula weight: 49055.94

Authors

Fedorov, A.A.,Fedorov, E.V.,Hall, R.S.,Raushel, F.M.,Almo, S.C. (deposition date: 2010-07-31, release date: 2011-06-15, Last modification date: 2023-10-04)

Primary citation

Hall, R.S.,Fedorov, A.A.,Xu, C.,Fedorov, E.V.,Almo, S.C.,Raushel, F.M.
Three-dimensional structure and catalytic mechanism of Cytosine deaminase.
Biochemistry, 50:5077-5085, 2011

PubMed Abstract: Cytosine deaminase (CDA) from E. coli is a member of the amidohydrolase superfamily. The structure of the zinc-activated enzyme was determined in the presence of phosphonocytosine, a mimic of the tetrahedral reaction intermediate. This compound inhibits the deamination of cytosine with a K(i) of 52 nM. The zinc- and iron-containing enzymes were characterized to determine the effect of the divalent cations on activation of the hydrolytic water. Fe-CDA loses activity at low pH with a kinetic pK(a) of 6.0, and Zn-CDA has a kinetic pK(a) of 7.3. Mutation of Gln-156 decreased the catalytic activity by more than 5 orders of magnitude, supporting its role in substrate binding. Mutation of Glu-217, Asp-313, and His-246 significantly decreased catalytic activity supporting the role of these three residues in activation of the hydrolytic water molecule and facilitation of proton transfer reactions. A library of potential substrates was used to probe the structural determinants responsible for catalytic activity. CDA was able to catalyze the deamination of isocytosine and the hydrolysis of 3-oxauracil. Large inverse solvent isotope effects were obtained on k(cat) and k(cat)/K(m), consistent with the formation of a low-barrier hydrogen bond during the conversion of cytosine to uracil. A chemical mechanism for substrate deamination by CDA was proposed.

PubMed: 21545144
DOI: 10.1021/bi200483k

Experimental method

X-RAY DIFFRACTION (1.708 Å)