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cDNA cloning and expression of human complement component C2.

T Horiuchi1, K J Macon, V J Kidd

  • 1Department of Medicine, University of Alabama, Birmingham 35294.

Journal of Immunology (Baltimore, Md. : 1950)
|March 15, 1989
PubMed
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Researchers isolated a full-length cDNA clone for human complement component C2 (C2). This clone, when expressed in COS cells, produced a protein with native C2 activity and antigenicity, confirming its functional relevance.

Area of Science:

  • Molecular Biology
  • Immunology
  • Genetics

Background:

  • The complement system is crucial for innate and adaptive immunity.
  • Complement component C2 (C2) plays a vital role in the classical and lectin complement pathways.
  • Understanding C2 gene and protein structure is essential for studying complement-mediated functions.

Purpose of the Study:

  • To isolate and characterize a full-length cDNA clone for human complement component C2 (C2).
  • To confirm the functional expression of the isolated C2 cDNA.
  • To compare the novel C2 cDNA sequence with previously reported sequences.

Main Methods:

  • Screening of a human liver cDNA library using antibodies and cDNA probes.
  • DNA sequencing of the isolated C2 cDNA clone (lambda C2HL5-3).

Related Experiment Videos

  • Cloning the full-length C2 cDNA into an expression vector and transfecting COS cells.
  • Assessing protein expression and activity via Western blot, antigenicity, and hemolytic assays.

    • Main Results:

    • Isolation and complete sequencing of a 2961-nucleotide full-length human C2 cDNA clone.
    • Identification of two significant differences compared to previous C2 cDNA sequences, including a longer 5' untranslated region and a single nucleotide change affecting an amino acid.
    • Successful expression of recombinant C2 (rC2) in COS cells, exhibiting antigenicity and hemolytic activity comparable to native C2.
    • Western blot confirmed rC2 molecular weight, and Northern blot showed longer C2 mRNA transcripts from the construct.

    Conclusions:

    • The isolated full-length C2 cDNA clone represents a functional gene for human complement component C2.
    • The characterized cDNA sequence provides a more complete understanding of C2 gene structure and potential regulatory elements.
    • The successful expression and functional validation of rC2 pave the way for further studies on complement biology and C2-related disorders.