Crystallization of a pentapeptide-repeat protein by reductive cyclic pentylation of free amines with glutaraldehyde
Open Access
- 18 April 2009
- journal article
- research article
- Published by International Union of Crystallography (IUCr) in Acta crystallographica. Section D, Structural biology
- Vol. 65 (5), 462-469
- https://doi.org/10.1107/s0907444909008324
Abstract
The pentapeptide-repeat protein EfsQnr from Enterococcus faecalis protects DNA gyrase from inhibition by fluoroquinolones. EfsQnr was cloned and purified to homogeneity, but failed to produce diffraction-quality crystals in initial crystallization screens. Treatment of EfsQnr with glutaraldehyde and the strong reducing agent borane-dimethylamine resulted in a derivatized protein which produced crystals that diffracted to 1.6 A resolution; their structure was subsequently determined by single-wavelength anomalous dispersion. Analysis of the derivatized protein using Fourier transform ion cyclotron resonance mass spectrometry indicated a mass increase of 68 Da per free amino group. Electron-density maps about a limited number of structurally ordered lysines indicated that the modification was a cyclic pentylation of free amines, producing piperidine groups.Keywords
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