1UX1

Bacillus subtilis cytidine deaminase with a Cys53His and an Arg56Gln substitution

Summary for 1UX1

• Entry DOI: 10.2210/pdb1ux1/pdb
• Related: 1JTK 1UWZ 1UX0
• Descriptor: CYTIDINE DEAMINASE, TETRAHYDRODEOXYURIDINE, ZINC ION, ... (5 entities in total)
• Functional Keywords: cytidine deaminase, cdd, tetramer, zinc binding, pyrimidine metabolism, salvage, hydrolase
• Biological source: BACILLUS SUBTILIS
• Total number of polymer chains: 4
• Total formula weight: 60804.46

Authors

Johansson, E.,Neuhard, J.,Willemoes, M.,Larsen, S. (deposition date: 2004-02-18, release date: 2004-05-20, Last modification date: 2023-12-13)

Primary citation

Johansson, E.,Neuhard, J.,Willemoes, M.,Larsen, S.
Structural, Kinetic, and Mutational Studies of the Zinc Ion Environment in Tetrameric Cytidine Deaminase
Biochemistry, 43:6020-, 2004

PubMed Abstract: The zinc-containing cytidine deaminase (CDA, EC 3.5.4.5) is a pyrimidine salvage enzyme catalyzing the hydrolytic deamination of cytidine and 2'-deoxycytidine forming uridine and 2'-deoxyuridine, respectively. Homodimeric CDA (D-CDA) and homotetrameric CDA (T-CDA) both contain one zinc ion per subunit coordinated to the catalytic water molecule. The zinc ligands in D-CDA are one histidine and two cysteine residues, whereas in T-CDA zinc is coordinated to three cysteines. Two of the zinc coordinating cysteines in T-CDA form hydrogen bonds to the conserved residue Arg56, and this residue together with the dipole moments from two alpha-helices partially neutralizes the additional negative charge in the active site, leading to a catalytic activity similar to D-CDA. Arg56 has been substituted by a glutamine (R56Q), the corresponding residue in D-CDA, an alanine (R56A), and an aspartate (R56D). Moreover, one of the zinc-liganding cysteines has been substituted by histidine to mimic D-CDA, alone (C53H) and in combination with R56Q (C53H/R56Q). R56A, R56Q, and C53H/R56Q contain the same amount of zinc as the wild-type enzyme. The zinc-binding capacity of R56D is reduced. Only R56A, R56Q, and C53H/R56Q yielded measurable CDA activity, R56A and R56Q with similar K(m) but decreased V(max) values compared to wild-type enzyme. Because of dissociation into its inactive subunits, it was impossible to determine the kinetic parameters for C53H/R56Q. R56A and C53H/R56Q display increased apparent pK(a) values compared to the wild-type enzyme and R56Q. On the basis of the structures of R56A, R56Q, and C53H/R56Q an explanation is provided of kinetic results and the apparent instability of C53H/R56Q.

PubMed: 15147186
DOI: 10.1021/BI035893X

Experimental method

X-RAY DIFFRACTION (2.36 Å)