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Rapid screening for structural integrity of expressed proteins by heteronuclear NMR spectroscopy

✍ Scribed by Angela M. Gronenborn; G. Marius Clore

Publisher
Cold Spring Harbor Laboratory Press
Year
2008
Tongue
English
Weight
284 KB
Volume
5
Category
Article
ISSN
0961-8368

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✦ Synopsis

A simple and rapid method based on 15N labeling and 1H-15N heteronuclear single quantum coherence spectroscopy is presented to directly assess the structural integrity of overexpressed proteins in crude Escherichia coli extracts without the need for any purification. The method is demonstrated using two different expression systems and two different proteins, the B1 immunoglobulin-binding domain of streptococcal protein G (56 residues) and human interleukin-1 beta (153 residues). It is shown that high quality 1H-15N correlation spectra, recorded in as little as 15 min and displaying only cross-peaks arising from the overexpressed protein of interest, can be obtained from crude E. coli extracts.

πŸ“œ SIMILAR VOLUMES

Structural and dynamic characterization
Structural and dynamic characterization of the urea denatured state of the immunoglobulin binding domain of streptococcal protein G by multidimensional heteronuclear NMR spectroscopy
✍ M. Kirsten Frank; G. Marius Clore; Angela M. Gronenborn πŸ“‚ Article πŸ“… 1995 πŸ› Cold Spring Harbor Laboratory Press 🌐 English βš– 1009 KB

## Abstract The structure and dynamics of the urea‐denatured B1 immunoglobulin binding domain of streptococcal protein G (GB1) has been investigated by multidimensional heteronuclear NMR spectroscopy. Complete ^1^H, ^15^N, and ^13^C assignments are obtained by means of sequential through‐bond corre