CRYSTAL-STRUCTURE OF DIMERIC HUMAN CILIARY NEUROTROPHIC FACTOR DETERMINED BY MAD PHASING

Ciliary neurotrophic factor (CNTF) promotes the survival and differentiation of developing motor neurons and is a potential therapeutic for treating neurodegeneration and nerve injury, The crystal structure of human CNTF has been determined at 2.4 Angstrom resolution using multi-wavelength anomalous diffraction (MAD) phasing from a single Yb3+ ion. The structure reveals that CNTF is dimeric, with a novel anti-parallel arrangement of the subunits, not previously observed for other cytokines, Each subunit adopts a double crossover four-helix bundle fold, in which two helices contribute to the dimer interface, whilst two different helices show pronounced kinks, Analysis of the electrostatic surface of CNTF identified residues within these kinked helices that may contact the CNTF receptor-alpha. Solution experiments show that CNTF dimerizes at concentrations >40 mu M. Such dimers are likely to be relevant to the storage of CNTF in the peripheral nerve given the high concentrations present in this tissue, However, it is unlikely that they play a role in engaging the three distinct receptor subunits that comprise the CNTF receptor, given the low concentration of extracellular CNTF and its high potency.