In vitro expression of the beta subunit of human complement component C8

Mol Immunol. 1996 Dec;33(17-18):1295-300. doi: 10.1016/s0161-5890(96)00108-3.

Abstract

Human C8 is one of five components of the cytolytic C5b-9 complex of complement. It is an oligomeric protein composed of three subunits (alpha, beta, gamma) encoded in separate genes. These are arranged as a disulfide-linked alpha-gamma dimer and a noncovalently associated beta chain. Biosynthesis studies and analyses of humans with hereditary C8 deficiencies suggest that C8 alpha-gamma synthesis and secretion can occur independently of C8 beta, but that serum levels of C8 beta are dependent on C8 alpha-gamma. One aim of the present study was to determine if functional human C8 beta could be synthesized in the absence of C8 alpha-gamma. Human C8 beta expression constructs were prepared and used to produce recombinant C8 beta (rC8 beta) in insect and COS-7 cells. Both cell types secreted rC8 beta that was similar in size to human C8 beta and exhibited similar ability to associate with human C8 alpha-gamma and form functional C8. A mutant form of C8 beta in which N-glycosylation sites were eliminated was also expressed and found to be functionally similar to rC8 beta and human C8 beta. These results indicate that C8 alpha-gamma is not required for intracellular processing and secretion of C8 beta. Furthermore, N-linked carbohydrate on C8 beta is not necessary for association with C8 alpha-gamma or for C8 activity.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Baculoviridae / genetics
  • Baculoviridae / metabolism
  • COS Cells
  • Complement C8 / biosynthesis*
  • Complement C8 / genetics*
  • Gene Expression Regulation
  • Genetic Vectors / biosynthesis
  • Glycosylation
  • Hemolysis
  • Humans
  • Insecta / cytology
  • Insecta / genetics
  • Peptide Fragments / biosynthesis
  • Peptide Fragments / blood
  • Peptide Fragments / genetics
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / metabolism

Substances

  • Complement C8
  • Peptide Fragments
  • Recombinant Proteins