Purification and characterization of multiple components of human lymphoblastoid interferon-alpha

Area of Science:

• Biochemistry
• Immunology
• Molecular Biology

Background:

• Human interferon-alpha (IFN-alpha) is a crucial cytokine with diverse biological activities.
• Understanding the heterogeneity of IFN-alpha components is essential for its therapeutic applications.

Purpose of the Study:

• To purify and characterize multiple components of human interferon-alpha (IFN-alpha) derived from Namalwa cells.
• To investigate the biochemical and sequence properties of purified IFN-alpha variants.

Main Methods:

• Sequential immunoadsorbent affinity chromatography using monoclonal antibodies.
• Ultrafiltration and reversed-phase high-performance liquid chromatography (RP-HPLC) for purification.
• Sodium dodecyl polyacrylamide-gel electrophoresis (SDS-PAGE) for molecular weight determination.
• Amino-terminal amino acid sequencing.

Main Results:

• Twenty-two distinct IFN-alpha components were purified, exhibiting a wide range of specific activities across different cell lines.
• Apparent molecular weights varied between non-reducing and reducing SDS-PAGE, indicating potential disulfide bonds.
• Amino-terminal sequences were largely conserved but four components (f, i, l, m) showed unique sequences compared to cloned IFN-alpha genes.
• One component (pre-a) possessed a potential N-linked glycosylation site.

Conclusions:

• The study successfully purified and characterized multiple human IFN-alpha components, highlighting their heterogeneity.
• Identified unique amino-terminal sequences and a potential glycosylation site suggest post-translational modifications or distinct gene products.
• These findings contribute to a deeper understanding of IFN-alpha diversity and its implications.