This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.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 the gentamicin complex crystallized in the presence of 16%(w/v) PEG 3350 and gentamicin. The crystals belong to the monoclinic space group P21, with approximate unit-cell parameters a = 79.7, b = 58.8, c = 81.4 , beta = 98.4 , and x-ray diffraction analysis is consistent with the presence of two molecules in the asymmetric unit. Synchrotron diffraction data to approximately 2.65 resolution were collected from a native APH(2 )-Ib crystal at beamline BL9-2. Data from selenium-substituted crystals has also been collected and the structure solved by MAD methods. The AMP and ATP-bound forms of APH(2 )-Ib have been crystallized in a trigonal space group P31, with cell dimensions a = b = 127.9, c = 58.1 , which diffract to much higher resolution. The structural analysis of these complexes is underway.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001209-28
Application #
7598096
Study Section
Special Emphasis Panel (ZRG1-BPC-E (40))
Project Start
2007-03-01
Project End
2008-02-29
Budget Start
2007-03-01
Budget End
2008-02-29
Support Year
28
Fiscal Year
2007
Total Cost
$1,191
Indirect Cost
Name
Stanford University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
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