PDBsum entry 2bkk

Transferase/peptide

PDB id: 2bkk

Contents

Protein chains

247 a.a. *

156 a.a. *

262 a.a. *

Ligands

ADP ×2

Metals

_MG ×4

Waters ×347

* Residue conservation analysis

PDB id:

2bkk

Name:

Transferase/peptide

Title:

Crystal structure of aminoglycoside phosphotransferase aph(3')-iiia in complex with the inhibitor ar_3a

Structure:

Aminoglycoside 3'-phosphotransferase. Chain: a, c. Synonym: aph iiia, kanamycin kinase type iii, neomycin-kanamycin phosphotransferase type iii, aph(3')iii. Engineered: yes. Mutation: yes. Designed ankyrin repeat inhibitor ar_3a. Chain: b, d. Engineered: yes

Source:

Enterococcus faecalis. Organism_taxid: 1351. Expressed in: escherichia coli. Expression_system_taxid: 469008. Other_details: streptococcus faecalis, staphylococcus aureus. Synthetic construct. Organism_taxid: 32630. Expression_system_taxid: 562

Biol. unit:

Dimer (from PDB file)

Resolution:

2.15Å R-factor: 0.200 R-free: 0.260

Authors:

A.Kohl,P.Amstutz,P.Parizek,H.K.Binz,C.Briand,G.Capitani,P.Forrer, A.Pluckthun,M.G.Grutter

Key ref:

A.Kohl et al. (2005). Allosteric inhibition of aminoglycoside phosphotransferase by a designed ankyrin repeat protein. Structure (Camb), 13, 1131-1141. PubMed id: 16084385 DOI: 10.1016/j.str.2005.04.020

Date:

16-Feb-05 Release date: 09-Aug-05

Protein chain

P0A3Y5  (KKA3_ENTFL) -  Aminoglycoside 3'-phosphotransferase from Enterococcus faecalis

Seq: 264 a.a.

Struc: 247 a.a.*

Protein chains

No UniProt id for this chain

Struc: 156 a.a.

Protein chain

P0A3Y5  (KKA3_ENTFL) -  Aminoglycoside 3'-phosphotransferase from Enterococcus faecalis

Seq: 264 a.a.

Struc: 262 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, C: E.C.2.7.1.95 - kanamycin kinase.
• Reaction: kanamycin A + ATP = kanamycin 3'-phosphate + ADP + H⁺

kanamycin A + ATP = kanamycin 3'-phosphate (Bound ligand (Het Group name = ADP) corresponds exactly) + ADP + H(+)

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

reference

DOI no: 10.1016/j.str.2005.04.020

Structure (Camb) 13:1131-1141 (2005)

PubMed id: 16084385

Allosteric inhibition of aminoglycoside phosphotransferase by a designed ankyrin repeat protein.

A.Kohl, P.Amstutz, P.Parizek, H.K.Binz, C.Briand, G.Capitani, P.Forrer, A.Plückthun, M.G.Grütter.

ABSTRACT

Aminoglycoside phosphotransferase (3')-IIIa (APH) is a bacterial kinase that confers antibiotic resistance to many pathogenic bacteria and shares structural homology with eukaryotic protein kinases. We report here the crystal structure of APH, trapped in an inactive conformation by a tailor-made inhibitory ankyrin repeat (AR) protein, at 2.15 A resolution. The inhibitor was selected from a combinatorial library of designed AR proteins. The AR protein binds the C-terminal lobe of APH and thereby stabilizes three alpha helices, which are necessary for substrate binding, in a significantly displaced conformation. BIAcore analysis and kinetic enzyme inhibition experiments are consistent with the proposed allosteric inhibition mechanism. In contrast to most small-molecule kinase inhibitors, the AR proteins are not restricted to active site binding, allowing for higher specificity. Inactive conformations of pharmaceutically relevant enzymes, as can be elucidated with the approach presented here, represent powerful starting points for rational drug design.

Selected figure(s)

Figure 2.
Figure 2. Crystal Structure of the mAPH in Complex with the AR Protein AR_3a
(A) Stereo view of the AB heterodimer of the mAPH/AR_3a complex. The mAPH is shown in orange, and the AR_3a is shown in light blue. AR_3a binds to the C-terminal lobe of the mAPH and stabilizes an inactive conformation.
(B) Asymmetric mAPH pseudo-homodimer found in the crystal of the mAPH/AR_3a complex.
(C) Symmetric wtAPH dimer found in the wtAPH crystal.
(D) Stereo view of the superposition of the mAPH hetereodimer AB (orange) and BC (light blue).
(E) wtAPH in the kanamycin A bound form (PDB: 1L8T1L8T). ADP and kanamycin A are labeled.
(F) mAPH/AR_3a AB heterodimer in the same orientation as the wtAPH in (E).

The above figure is reprinted by permission from Cell Press: Structure (Camb) (2005, 13, 1131-1141) copyright 2005.

Figure was selected by the author.