PDBsum entry: 1wei

Hydrolase

PDB id: 1wei

Contents

Protein chain

220 a.a. *

Ligands

ADE

EDO ×3

SF4

Waters ×301

* Residue conservation analysis

PDB id:

1wei

Name:

Hydrolase

Title:

Catalytic domain of muty from escherichia coli k20a mutant complexed to adenine

Structure:

A/g-specific adenine glycosylase. Chain: a. Fragment: catalytic domain. Synonym: adenine glycosylase. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 562. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

1.45Å R-factor: 0.217 R-free: 0.232

Authors:

K.Hitomi,A.S.Arvai,J.A.Tainer

Key ref:

R.C.Manuel et al. (2004). Reaction intermediates in the catalytic mechanism of Escherichia coli MutY DNA glycosylase. J Biol Chem, 279, 46930-46939. PubMed id: 15326180 DOI: 10.1074/jbc.M403944200

Date:

25-May-04 Release date: 21-Sep-04

Protein chain

P17802  (MUTY_ECOLI) -  A/G-specific adenine glycosylase

Seq: 350 a.a.

Struc: 220 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

 Gene Ontology (GO) functional annotation 

• Cellular component: intracellular (1 term)
• Biological process: DNA repair (2 terms)
• Biochemical function: catalytic activity (5 terms)

DOI no: 10.1074/jbc.M403944200

J Biol Chem 279:46930-46939 (2004)

PubMed id: 15326180

Reaction intermediates in the catalytic mechanism of Escherichia coli MutY DNA glycosylase.

R.C.Manuel, K.Hitomi, A.S.Arvai, P.G.House, A.J.Kurtz, M.L.Dodson, A.K.McCullough, J.A.Tainer, R.S.Lloyd.

ABSTRACT

The Escherichia coli adenine DNA glycosylase, MutY, plays an important role in the maintenance of genomic stability by catalyzing the removal of adenine opposite 8-oxo-7,8-dihydroguanine or guanine in duplex DNA. Although the x-ray crystal structure of the catalytic domain of MutY revealed a mechanism for catalysis of the glycosyl bond, it appeared that several opportunistically positioned lysine side chains could participate in a secondary beta-elimination reaction. In this investigation, it is established via site-directed mutagenesis and the determination of a 1.35-A structure of MutY in complex with adenine that the abasic site (apurinic/apyrimidinic) lyase activity is alternatively regulated by two lysines, Lys142 and Lys20. Analyses of the crystallographic structure also suggest a role for Glu161 in the apurinic/apyrimidinic lyase chemistry. The beta-elimination reaction is structurally and chemically uncoupled from the initial glycosyl bond scission, indicating that this reaction occurs as a consequence of active site plasticity and slow dissociation of the product complex. MutY with either the K142A or K20A mutation still catalyzes beta and beta-delta elimination reactions, and both mutants can be trapped as covalent enzyme-DNA intermediates by chemical reduction. The trapping was observed to occur both pre- and post-phosphodiester bond scission, establishing that both of these intermediates have significant half-lives. Thus, the final spectrum of DNA products generated reflects the outcome of a delicate balance of closely related equilibrium constants.

Selected figure(s)

Figure 2.
FIG. 2. Detailed structural comparisons of the cMutY wild type and mutant enzymes. The wild type cMutY is shown in orange, K20A in blue, D138N in yellow, and K142A in green. A, conservation of the active site in the K20A mutant. B, contribution of Lys142 to the ordering of the Pro155-Gly156-Lys157-Lys158-Glu159 loop through a water. In the K142A mutants, Glu161, shown in a box, was disordered. C, the K20A mutant with adenine bound. Adenine is shown in pale green. D, adenine binding changes the position of the Glu161 side chain.

Figure 9.
FIG. 9. The catalytic mechanism of MutY. Schematic representation showing the proposed reaction mechanism of MutY for glycosylase (A) and lyase (B) activities. B illustrates the spectrum of AP lyase products, which are isolated as stable chemical species in the presence of a reducing agent.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2004, 279, 46930-46939) copyright 2004.

Figures were selected by an automated process.