V170 AREA PLAYS AN ESSENTIAL ROLE IN THE BIOLOGICAL-ACTIVITY OF HUMANCILIARY NEUROTROPHIC FACTOR

The structure-function relationships of human ciliary neurotrophic fac tor (CNTF) were analyzed by mutagenic means. Amino acid substitutions at helix D caused marked changes in the biological activity of CNTF, s uggesting that the residues at helix D of CNTF participate in receptor recognition. In particular, both the cell survival-promoting activity and receptor binding ability of V170 mutant CNTF proteins correlated well with the hydrophobicity of amino acids at position 170. The reduc tion of hydrophobicity at position 170 resulted in a loss of biologica l activity, indicating that the hydrophobicity of V170 is essential fo r the receptor binding and cell survival-promoting activity. Substitut ions of R171 or D175 evoked very little folding ability and negated th e biological activity of CNTF. As R171 and D175 interact electrostatic ally with each other and with E75 and R72, respectively, these interac tions would be indispensable for stabilizing the whole CNTF protein an d for maintaining the structure of the receptor binding epitope.