Covalent binding of human α2-macroglobulin to deglycosylated ricin A chain and its immunotoxins

Maria Ana Ghetie, Jonathan W. Uhr, Ellen S. Vitetta

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Abstract

In this report we demonstrate that human α2-macroglobulin (α2M) reacts with deglycosylated ricin A chain (dgA) and its immunotoxins to form high molecular weight complexes (molecular mass approximately 800 kDa). This interaction has a t1/2 at 37°C of 5 h and reaches completion at 24 h. Complexes of α2M-dgA cannot be dissociated by guanidine, sodium dodecyl sulfate, or low pH, but can be partially dissociated by reducing agents, such as 2-mercaptoethanol in the presence of sodium dodecyl sulfate. This indicates that dgA or dgA-containing immunotoxins are bound to α2M by disulfide bonds. The dgA-binding site on α2M and the mechanism underlying its interaction with dgA are different from those described for proteases or methylamine. α2M complexes do not bind to Blue-Sepharose 4B or anti-A chain-Sepharose, suggesting that the sites on dgA which bind Cibacron Blue or polyclonal anti-A chain antibodies are sterically blocked or modified by interaction with α2M. The interaction of α2M with dgA or its immunotoxins results in a 2- to 3-fold decrease in the activity of the dgA in both cell-free assays and cytotoxic assays. However 12 h after injection into mice, only 11% of immunotoxin was bound to α2M because of the slow kinetics of the interaction versus the more rapid t1/2 of the immunotoxin in the circulation.

Original languageEnglish (US)
Pages (from-to)1482-1487
Number of pages6
JournalCancer Research
Volume51
Issue number5
StatePublished - Mar 1 1991

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Ricin
Immunotoxins
Macroglobulins
Sodium Dodecyl Sulfate
Sepharose
Mercaptoethanol
Reducing Agents
Guanidine
Disulfides
Peptide Hydrolases
Molecular Weight
Binding Sites
Injections
Antibodies

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Covalent binding of human α2-macroglobulin to deglycosylated ricin A chain and its immunotoxins. / Ghetie, Maria Ana; Uhr, Jonathan W.; Vitetta, Ellen S.

In: Cancer Research, Vol. 51, No. 5, 01.03.1991, p. 1482-1487.

Research output: Contribution to journalArticle

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