Covalent binding of [35S]cysteine to the labile binding site of the third component of complement: A physiological approach

Cynthia J. Rutherford, David R. Jenkins

Research output: Contribution to journalArticle

Abstract

The alternative pathway of complement was activated physiologically by agarose beads to which [35S]cysteine had been bound by a disulfide link. The activated form of the third component of complement, C3b, which had bound to the radioactive cysteine was then released from tharose bead with dithiothreitol. The [35S]cysteine was shown to be covalently bound to the α′ chain of C3b, and to its known major breakdown product, the 66,000-Da polypeptide. This is highly suggestive that complement activation has led to formation of a peptide bond between the radioactive cysteine and the labile binding site on the α′ chain of C3b. This radioactive marker will enable the amino acid sequence of the labile binding site to be determined, with the knowledge that the labeling of the amino acid(s) has occurred during physiological activation.

Original languageEnglish (US)
Pages (from-to)77-82
Number of pages6
JournalClinical Immunology and Immunopathology
Volume37
Issue number1
DOIs
StatePublished - 1985

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Cysteine
Binding Sites
Complement C3b
Alternative Complement Pathway
Peptides
Complement Activation
Dithiothreitol
Disulfides
Sepharose
Amino Acid Sequence
Amino Acids

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy
  • Pathology and Forensic Medicine

Cite this

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abstract = "The alternative pathway of complement was activated physiologically by agarose beads to which [35S]cysteine had been bound by a disulfide link. The activated form of the third component of complement, C3b, which had bound to the radioactive cysteine was then released from tharose bead with dithiothreitol. The [35S]cysteine was shown to be covalently bound to the α′ chain of C3b, and to its known major breakdown product, the 66,000-Da polypeptide. This is highly suggestive that complement activation has led to formation of a peptide bond between the radioactive cysteine and the labile binding site on the α′ chain of C3b. This radioactive marker will enable the amino acid sequence of the labile binding site to be determined, with the knowledge that the labeling of the amino acid(s) has occurred during physiological activation.",
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AB - The alternative pathway of complement was activated physiologically by agarose beads to which [35S]cysteine had been bound by a disulfide link. The activated form of the third component of complement, C3b, which had bound to the radioactive cysteine was then released from tharose bead with dithiothreitol. The [35S]cysteine was shown to be covalently bound to the α′ chain of C3b, and to its known major breakdown product, the 66,000-Da polypeptide. This is highly suggestive that complement activation has led to formation of a peptide bond between the radioactive cysteine and the labile binding site on the α′ chain of C3b. This radioactive marker will enable the amino acid sequence of the labile binding site to be determined, with the knowledge that the labeling of the amino acid(s) has occurred during physiological activation.

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