PDBsum entry 1v0c

Transferase

PDB id: 1v0c

Contents

Protein chain

176 a.a. *

Ligands

ACO

KNC

Metals

_CA ×2

Waters ×158

* Residue conservation analysis

PDB id:

1v0c

Name:

Transferase

Title:

Structure of aac(6')-ib in complex with kanamycin c and acetylcoa.

Structure:

Aac(6')-ib. Chain: a. Synonym: fluroquinolone acetylating aminoglycoside acetyltransferase, aac6-ib. Engineered: yes. Mutation: yes

Source:

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

Resolution:

2.20Å R-factor: 0.179 R-free: 0.225

Authors:

M.W.Vetting,C.H.Park,S.S.Hedge,D.C.Hooper,J.S.Blanchard

Key ref:

M.W.Vetting et al. (2008). Mechanistic and structural analysis of aminoglycoside N-acetyltransferase AAC(6')-Ib and its bifunctional, fluoroquinolone-active AAC(6')-Ib-cr variant. Biochemistry, 47, 9825-9835. PubMed id: 18710261

Date:

20-Mar-08 Release date: 02-Sep-08

Protein chain

Q6SJ71  (Q6SJ71_ECOLX) -  AAC(6')-Ib from Escherichia coli

Seq: 199 a.a.

Struc: 176 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.2.3.1.82 - aminoglycoside 6'-N-acetyltransferase.
• Reaction: kanamycin B + acetyl-CoA = N(6')-acetylkanamycin B + CoA + H⁺

kanamycin B (Bound ligand (Het Group name = ACO) corresponds exactly) + acetyl-CoA (Bound ligand (Het Group name = KNC) matches with 94.12% similarity) = N(6')-acetylkanamycin B + CoA + H(+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

Biochemistry 47:9825-9835 (2008)

PubMed id: 18710261

Mechanistic and structural analysis of aminoglycoside N-acetyltransferase AAC(6')-Ib and its bifunctional, fluoroquinolone-active AAC(6')-Ib-cr variant.

M.W.Vetting, C.H.Park, S.S.Hegde, G.A.Jacoby, D.C.Hooper, J.S.Blanchard.

ABSTRACT

Enzymatic modification of aminoglycoside antibiotics mediated by regioselective aminoglycoside N-acetyltransferases is the predominant cause of bacterial resistance to aminoglycosides. A recently discovered bifunctional aminoglycoside acetyltransferase (AAC(6')-Ib variant, AAC(6')-Ib-cr) has been shown to catalyze the acetylation of fluoroquinolones as well as aminoglycosides. We have expressed and purified AAC(6')-Ib-wt and its bifunctional variant AAC(6')-Ib-cr in Escherichia coli and characterized their kinetic and chemical mechanism. Initial velocity and dead-end inhibition studies support an ordered sequential mechanism for the enzyme(s). The three-dimensional structure of AAC(6')-Ib-wt was determined in various complexes with donor and acceptor ligands to resolutions greater than 2.2 A. Observation of the direct, and optimally positioned, interaction between the 6'-NH 2 and Asp115 suggests that Asp115 acts as a general base to accept a proton in the reaction. The structure of AAC(6')-Ib-wt permits the construction of a molecular model of the interactions of fluoroquinolones with the AAC(6')-Ib-cr variant. The model suggests that a major contribution to the fluoroquinolone acetylation activity comes from the Asp179Tyr mutation, where Tyr179 makes pi-stacking interactions with the quinolone ring facilitating quinolone binding. The model also suggests that fluoroquinolones and aminoglycosides have different binding modes. On the basis of kinetic properties, the pH dependence of the kinetic parameters, and structural information, we propose an acid/base-assisted reaction catalyzed by AAC(6')-Ib-wt and the AAC(6')-Ib-cr variant involving a ternary complex.