THE 3-DIMENSIONAL HIGH-RESOLUTION STRUCTURE OF HUMAN INTERFERON ALPHA-2A DETERMINED BY HETERONUCLEAR NMR-SPECTROSCOPY IN SOLUTION

The solution structure of recombinant human interferon alpha-2a (Rofer on-A) has been determined by :multidimensional heteronuclear NMR spect roscopy. The calculations using simulated annealing produced a family of 24 convergent structures which satisfy the experimental restraints comprising 1541 NOE-derived inter-proton distances, 187 dihedral restr aints, 66 pairs of hydrogen bond restraints, and sir, upper and lower limits for two disulfide bridges. The fractional labeling of methyl gr oups allowed their direct and unambiguous stereospecific assignment wh ich proved to be essential for obtaining a high resolution of the stru ctures. A best fit superposition of residues 10 to 47, 50 to 101 and 1 11 to 157 gives an rms deviation of 0.62 Angstrom for the backbone hea vy atoms and 1.39 Angstrom for all heavy atoms of these segments. The dominant feature of the structure is a cluster of five alpha-helices, four of which are arranged to form a left-handed helix bundle with an up-up-down-down topology and two overhand connections. The interpretat ion of heteronuclear N-15-[H-1] NOE data shows the co-existence of fle xible regions within an otherwise rigid framework of the protein. Four stretches of pronounced flexibility can be located: Cys1-Ser8, Gly44- Ala50, Ile100-Lys112, and Ser160-Glu165. Among the structurally relate d four-helical bundle cytokines, the structure of IFN alpha-2a is most similar to that of human interferon alpha-2b and murine interferon-be ta. From this structural information and mutagenesis data, areas on th e surface of the protein are identified which seem to be important :in receptor interactions. (C) 1997 Academic Press Limited.