3W0Q: Crystal Structure Of A Thermostable Mutant Of Aminoglycoside Phosphotransferase Aph(4)-ia (n203a), Ternary Complex With Amp-pnp And Hygromycin B

Aminoglycoside 4-phosphotransferase-Ia (APH(4)-Ia)/Hygromycin B phosphotransferase (Hph) inactivates the aminoglycoside antibiotic hygromycin B (hygB) via phosphorylation. The crystal structure of the binary complex of APH(4)-Ia with hygB was recently reported. To characterize substrate recognition by the enzyme, we determined the crystal structure of the ternary complex of non-hydrolyzable ATP analog AMP-PNP and hygB with wild-type, thermostable Hph mutant Hph5, and apo-mutant enzyme forms. The comparison between the ternary complex and apo structures revealed that Hph undergoes domain movement upon binding of AMP-PNP and hygB. This was about half amount of the case of APH(9)-Ia. We also determined the crystal structures of mutants in which the conserved, catalytically important residues Asp198 and Asn203, and the non-conserved Asn202, were converted to Ala, revealing the importance of Asn202 for catalysis. Hph5 contains five amino acid substitutions that alter its thermostability by 16 degrees C; its structure revealed that 4/5 mutations in Hph5 are located in the hydrophobic core and appear to increase thermostability by strengthening hydrophobic interactions.
PDB ID: 3W0QDownload
MMDB ID: 112361
PDB Deposition Date: 2012/11/2
Updated in MMDB: 2013/08
Experimental Method:
x-ray diffraction
Resolution: 1.8  Å
Source Organism:
Similar Structures:
Biological Unit for 3W0Q: monomeric; determined by author and by software (PISA)
Molecular Components in 3W0Q
Label Count Molecule
Protein (1 molecule)
Hygromycin-b 4-o-kinase(Gene symbol: PWD4IncI1_10)
Molecule annotation
Chemicals (2 molecules)
* Click molecule labels to explore molecular sequence information.

Citing MMDB