title: Covalent binding of human alpha 2-macroglobulin to deglycosylated ricin A chain and its immunotoxins.
abstract: In this report we demonstrated that human alpha 2-macroglobulin (alpha 2M) 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 degrees C of 5 h and reaches completion at 24 h. Complexes of alpha 2M-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 alpha 2M by disulfide bonds. The dgA-binding site on alpha 2M and the mechanism underlying its interaction with dgA are different from those described for proteases or methylamine. alpha 2M 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 alpha 2M. The interaction of alpha 2M 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 alpha 2M because of the slow kinetics of the interaction versus the more rapid t1/2 of the immunotoxin in the circulation.
authors.affiliation: Department of Microbiology, University of Texas Southwestern Medical Center, Dallas 75235.
authors.forename: M A
authors.initials: MA
authors.lastname: Ghetie
authors.forename: J W
authors.initials: JW
authors.lastname: Uhr
authors.forename: E S
authors.initials: ES
authors.lastname: Vitetta
mesh_terms: D000818:Animals; D006031:Glycosylation; D006207:Half-Life; D006801:Humans; D000922:Immunotoxins / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D011817:Rabbits; D012276:Ricin / Q000378:metabolism* / Q000633:toxicity; D000511:alpha-Macroglobulins / Q000378:metabolism*
chemical_list: D000922:Immunotoxins; D000511:alpha-Macroglobulins; D012276:Ricin