Structure and Activity of Recombinant Human Interferon-γ Analogs

Abstract

We have prepared interferon-gamma (IFN-gamma) analogs to study the structural role of particular amino acids in relation to their effects on antiviral activity. Three IFN-gamma analogs were prepared on the basis of predicted secondary structure. In two of the analogs, [Gln25]IFN-gamma and [Thr45]IFN-gamma, changes were made at residue 25 (Asn to Gln) and at residue 45 (Met to Thr), respectively. [Gln25Lys78]IFN-gamma had two changes, at residue 25 (Asn to Gln) and residue 78 (Asn to Lys). Another analog, [Cys-Tyr-Cys]IFN-gamma, incorporated Cys-Tyr-Cys at the amino terminus. Comparison of the structure and activity of these analogs with that of the natural sequence protein suggested that residues 25 and 78 are at the protein surface and play an important role in antiviral activity. The residue at position 45 was found to be important for maintaining the protein structure, as assessed by circular dichroism spectroscopy. The addition of Cys-Tyr-Cys resulted in a small perturbation of protein structure and a small decrease in antiviral activity.

Construction and analysis of a profile library characterizing groups of structurally known proteins

“…These regions were spatially close in the 3D structure (data not shown). From the database information only, we could not con-clude whether these regions were related to the activity of IFNy, but the residue at position 45, immediate upstream of the first found block, was reported to be important in maintaining the protein structure by an experiment using IFNy analogues (Hsu et al, 1986). Generally, the regions covered by the StrProf profiles were much longer than those of PROSITE patterns.…”

Two conserved tryptophan residues of tumor necrosis factor and lymphotoxin are not involved in the biological activity

“…Cys 69 can still be found in mLT. These cysteine residues in TNF form a disulfide bridge, that is probably not necessary for maintaining the overall conformation as shown by CD spectra [7,8]. Fluorescence spectra and quenching, however, indicated that, although the biological activity was not reduced significantly, the local conformation around the two tryptophan residues was affected after removing this disulfide bond [9].…”