Covalent Binding of Human a2-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 a2-macroglobulin (a2M)reacts with deglycosylated ricin A chain (dgA) and its immunotoxins toform high molecular weight complexes (molecular mass approximately800 kDa). This interaction has a /i/2at 37°Cof 5 h and reaches completionat 24 h. Complexes of «2M-dgAcannot be dissociated by guanidine,sodium dodecyl sulfate, or low pi I, but can be partially dissociated byreducing agents, such as 2-mercaptoethanol in the presence of sodiumdodecyl sulfate. This indicates that dgA or dgA-containing immunotoxinsare bound to a2M by disulfide bonds. The dgA-binding site on a2M andthe mechanism underlying its interaction with dgA are different fromthose described for proteases or methylamine. 2Mcomplexes do notbind to Blue-Sepharose 4B or anti-A chain-Sepharose, suggesting thatthe sites on dgA which bind Cibacron Blue or polyclonal anti-A chainantibodies are sterically blocked or modified by interaction with,. 2M.The interaction of a2M with dgA or its immunotoxins results in a 2- to3-fold decrease in the activity of the dgA in both cell-free assays andcytotoxic assays. However 12 h after injection into mice, only 11% ofimmunotoxin was bound to «2Mbecause of the slow kinetics of theinteraction versus the more rapid r1/2 of the immunotoxin in thecirculation.

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

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ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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