Conformation and activity of recombinant human fibroblast interferon-beta.
Conformation of highly purified recombinant human fibroblast interferon-beta (rHuIFN-beta) was correlated with its biological activity. The extent of ordered secondary structure was determined by circular dichroic (CD) spectroscopy in various buffer conditions to establish conditions of protein stability and its potential for helix formation. The highest "helicity" (about 50 +/- 5% of alpha-helices) and the highest antiviral activities (4-10 x 10(7) units/mg) were found in 50% ethylene glycol, 1 M NaCl and 0.05 M Na3PO4, pH 7.2 (Buffer I); 80 mM citric acid, 20 mM Na2HPO4, pH 2.9 (Buffer II); and 25 mM NH4OAc, 125 mM NaCl, pH 5.1 (Buffer III). Both helicity and antiviral activity of the IFN-beta decrease in parallel with denaturation by urea, heat, and/or by repeated cycles of freezing and thawing. Low pH (pH 2.9 Buffer II) exhibits a distinct stabilizing effect on the structure and antiviral activity of IFN-beta against heat denaturation.
Boublik
M
M
Roche Institute of Molecular Biology, Roche Research Center, Nutley, NJ 07110.
Moschera
J A
JA
Wei
C
C
Kung
H F
HF
D000595:Amino Acid Sequence; D002021:Buffers; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007370:Interferon Type I / Q000032:analysis / Q000494:pharmacology; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship
D002021:Buffers; D007370:Interferon Type I; D011994:Recombinant Proteins