A-Interferon Structure and Natural Killer Cell Stimulatory Activity1
[CANCER RESEARCH 50. 5328-5332, September 1, 1990] a-Interferon Structure and Natural Killer Cell Stimulatory Activity1 Bo-Liang Li,2 Xiao-Xia Zhao,3 Xin-Yuan Liu,2 Hyon Suk Kim, Karel Raska, Jr., John R. Ortaldo, Barbara Schwartz, and Sidney Pestka4 Departments of Molecular Genetics and Microbiology [B.-L. L., X-X. Z.. X-Y. L., B. S., S. P.] and Pathology [H. S. A., A'. R.J, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, Sew Jersey 08854, and National Cancer Institute, Frederick Cancer Research Facility, Frederick, Maryland 217011J. R.O.I ABSTRACT chased from New England BioLabs, T4 DNA ligase from International Biotechnologies Inc., polynucleotide kinase and deoxyribonucleotide Expression vectors for human a-interferon (Hu-IFN-a) .11. a site- triphosphates from P-L Biochemicals, and [«-"SjATP and [7-12P]ATP specific mutant |Ser"6]Hu-IFN-aJl, and Hu-IFN-aJ/C or Hu-IFN-aC/ from Amersham Corporation. J hybrids were constructed and expressed in Escherichìacoli. These Oligonucleotide-directed Site-specific Mutagenesis. M13mp8 Rf interferons and others were purified by immunoaffinity chromatography DN A was digested with £coRIand Hindi (Fig. 1) and then precipitated with a monoclonal antibody against human a-interferon. Their antiviral by equal volumes of 13% polyethylene glycol/1.6 M NaCl (9). The activity and ability to stimulate natural killer cell activity were determined precipitate was dissolved in 0.01 M Tris •HCl,pH 7.5-1.0 IHMEDTA) in comparison to several other human interferons. These results provide and ligated to the J689 fragment from the Hu-IFN-aJl gene (1, 10, some insight into structure-activity relationships for stimulating natural 11) under the following conditions: 100 ng of M13mpl8 Rf DNA killer cells and confirm our previous conclusions that antiviral activity vector fragment (EcoRl/Hincl\), 10-40 ng of fragment J689, and 0.9 cannot be used to predict other activities for an individual IFN-a species.
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