Open AccessPurification, crystallization and preliminary X‐ray analysis of Enterococcus faecium aminoglycoside‐2′′‐phosphotransferase‐Ib [APH(2′′)‐Ib]
Author(s) -
Walanj Rupa,
Young Paul,
Baker Heather M.,
Baker Edward N.,
Metcalf Peter,
Chow Joseph W.,
Lerner Stephen,
Vakulenko Sergei,
Smith Clyde A.
Publication year - 2005
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s174430910500775x
Subject(s) - enterococcus faecium , phosphotransferase , crystallization , microbiology and biotechnology , aminoglycoside , enterococcus , chemistry , biology , antibiotics , biochemistry , enzyme , organic chemistry
Bacterial resistance to the aminoglycoside antibiotics is primarily the result of deactivation of the drugs. Three families of enzymes are responsible for this activity, with one such family being the aminoglycoside phosphotransferases (APHs). The gene encoding one of these enzymes, APH(2′′)‐Ib, has been cloned and the protein (comprising 299 amino‐acid residues) expressed in Escherichia coli , purified and crystallized in the presence of 16%( w / v ) PEG 3350 and gentamicin. The crystals belong to the monoclinic space group P 2 1 , with approximate unit‐cell parameters a = 79.7, b = 58.8, c = 81.4 Å, β = 98.4°, and preliminary X‐ray diffraction analysis is consistent with the presence of two molecules in the asymmetric unit. Synchrotron diffraction data to approximately 2.65 Å resolution were collected from a native APH(2′′)‐Ib crystal at beamline BL9‐2 at SSRL (Stanford, CA, USA). Selenium‐substituted crystals have also been produced and structure determination is proceeding.