Two interferons alpha influence each other during their interaction with the extracellular domain of human type interferon receptor subunit 2.
The interaction between two human interferons alpha (IFN-alphas) and the extracellular (EC) domain of human type I IFN receptor subunit 2 (IFNAR2) was analyzed. Previous experiments using Daudi cells showed that IFN-alpha21b and some IFN-alpha hybrids (made from IFN-alpha2c and 21b) competed poorly for the IFN-alpha2b binding site. This study examined the causes of the poor competition between these IFN-alphas. IFN-alpha2c and the IFN hybrid CM3 {IFN-alpha21b(1-75)(81-95)/IFN-alpha2c(76-80) (96-166), Y86K} were selected for this study based on their cell binding and biological properties. Competitive binding ELISA, native electrophoresis followed by Western blot, electrospray ionization mass spectrometry (ESI-MS), surface plasmon resonance biosensor (SPR) analysis, as well as neutralization of antiproliferative activities on Daudi cells in the presence of soluble IFNAR2-EC show evidence that each of the described IFN-alpha subtypes affected the binding of the other IFN-alpha to IFNAR2-EC by affecting the stability of the complex, i.e., dissociation of the complex. Moreover, native electrophoresis with different IFNAR2-EC mutants showed that IFN-alpha2c and CM3 utilize different amino acids in the binding domain of IFNAR2-EC. In addition to that, analytical ultracentrifugation (AUC) revealed differences in the oligomeric state of the two studied interferons. Our results demonstrated that two individual IFN-alphas interact differentially with IFNAR2-EC and influence each other during this interaction. This study contributes to the understanding of the mutual interaction between multiple IFN-alpha subtypes during the competition for binding to the receptor.
Schmeisser
Hana
H
National Institute of Allergy and Infectious Diseases and National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, Maryland 20892, USA. hschmeisser@niaid.nih.gov
Gorshkova
Inna
I
Brown
Patrick H
PH
Kontsek
Peter
P
Schuck
Peter
P
Zoon
Kathryn C
KC
D015153:Blotting, Western; D045744:Cell Line, Tumor; D004586:Electrophoresis; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D016898:Interferon-alpha / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D053633:Receptor, Interferon alpha-beta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D020349:Surface Plasmon Resonance
D016898:Interferon-alpha; D053633:Receptor, Interferon alpha-beta