Uniform labeling of a recombinant antibody Fv‐fragment with 15N and 13C for heteronuclear NMR spectroscopy
Authors:
- John CavanaghJCJohn Cavanagh
- Siobhan McManusSMSiobhan McManus
- 5 August 1991
- journal article
- Published by Wiley in FEBS Letters
- Vol. 287 (1-2) , 185-188
Abstract
The expression of functional antibody fragments in Escherichia coli enables a detailed analysis by NMR spectroscopy. This is demonstrated with the uniform labeling of an Fv‐fragment (25 kDa) comprising the antigen binding site of an anitbody against 2‐phenyloxazolone with 15N and 13C. The antigen‐complexed Fv‐fragment was analysed for a potential assignment by heteronuclear multi‐dimensional NMR spectroscopy. For almost all backbone amides 15N/1H crosspeaks and for 80% of them TOCSY crosspeaks were observed. In a 13C‐edited‐HCCH‐2D experiment 17 out of 18 threonine spin‐systems were identified. Thus detailed assignments are possible, but some amino acid specific labeling in addition to uniform labeling will be required for complete assignments of Fv‐fragments.Keywords
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