Resonance assignments, secondary structure and topology of leukaemia inhibitory factor in solution

The chemical shift assignments and secondary structure of a murine-human chimera, MH35, of leukaemia inhibitory factor (LIF), a 180-residue protein of molecular mass 20 kDa, have been determined from multidimensional heteronuclear NMR spectra acquired on a uniformly ¹³C,¹⁵N-labelled sample. Secondary structure elements were defined on the basis of chemical shifts, NH-C^αH coupling constants, medium-range NOEs and the location of slowly exchanging amide protons. The protein contains four α-helices, the relative orientations of which were determined on the basis of long-range, interhelical NOEs. The four helices are arranged in an up-up-down-down orientation, as found in other four-helical bundle cytokines. The overall topology of MH35-LIF is similar to that of the X-ray crystallographic structure for murine LIF [Robinson et al. (1994) Cell, 77, 1101-1116]. Differences between the X-ray structure and the solution structure are evident in the N-terminal tail, where the solution structure has a trans-Pro¹⁷ compared with the cis-Pro¹⁷ found in the crystal structure and the small antiparallel β-sheet encompassing residues in the N-terminus and CD loop in the crystal structure is less stable.