Characterization of two fluorescent tryptophans in recombinant human granulocyte-colony stimulating factor: comparison of native sequence protein and tryptophan-deficient mutants.

摘要: In order to probe the role of individual tryptophans granulocyte-colony stimulating factor (G-CSF) inpH and guanidine HCl-induced fluorescence changes, site-directed mutagenesis was used generate mutants replacing Trp118, Trp58, or both with phenylalanine. Neither Trp Phe mutation affected folding activity recombinant G-CSF, material expressed in yeast behaved identically that inEscherichia coli. All G-CSF species responded topH HCl qualitatively same manner. Trp58 has a maximum at 350 nm is quenched greater extent by addition HCl, indicating it fully solvent-exposed. Trp118 344 nm, less solvent-accessible than Trp58. The analog which have been replaced phenylalanine shows only tyrosine fluorescence, peak 304 decreases increasingpH. intensity this much native sequence protein single tryptophan mutants, energy transfer taking place from these molecules. Below neutralpH [Phe58]G-CSF [Phe118]G-CSF, might be more efficient recipient tyrosine(s).