UNIFORM LABELING OF A RECOMBINANT ANTIBODY FV-FRAGMENT WITH N-15 AND C-13 FOR HETERONUCLEAR NMR-SPECTROSCOPY

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 antibody against 2-phenyloxazolone with N-15 and C-13. The antigen-complexed Fv-fragment was analysed for a potential assignment by heteronuclear multi-dimensional NMR spectroscopy. For almost all backbone amides N-15/H-1 crosspeaks and for 80% of them TOCSY crosspeaks were observed. In a C-13-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.