Mistletoe lectin A-chain unfolds during the intracellular transport.
Protein conformation during intracellular routing and translocation of the ribosome-inactivating proteins was investigated on hybridomas producing monoclonal antibodies (monAbs) against mistletoe lectin (ML). Decrease in the toxin activity towards these hybridomas is accounted for by the intracellular interaction of monAbs and the toxin resulting in the interruption of enzymatic subunit translocation into the cytosol. Obtained monAbs interacted with denatured ML A-chain (MLA) and a panel of MLA synthetic octapeptides linked to the surface of polyethylene pins. Enzyme-linked immunosorbent assay (ELISA) shows that monAbs recognize five epitopes in denatured MLA. Treatment of MLA by 3 M of guanidine hydrochloride leads to appearance of the epitopes. Hybridoma TA7 has been shown to be insensitive to cytotoxic action of ML. TA7 monAb as we have shown recognizes epitope 101-105, FTGTT, and inhibits the liposome aggregation induced by MLA. A study of the cytotoxicity of ML and ricin for the hybridomas revealed that the unfolding of A-chain is probably required for intracellular transport and cytotoxic activity of ML.
Agapov
I I
II
State Research Institute of Genetics and Selection of Industrial Microorganisms, 1st Dorozhnyi pr. 1, 113545, Moscow, Russia.
Tonevitsky
A G
AG
Maluchenko
N V
NV
Moisenovich
M M
MM
Bulah
Y S
YS
Kirpichnikov
M P
MP
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology; D019791:Guanidine / Q000494:pharmacology; D006825:Hybridomas / Q000276:immunology; D020128:Inhibitory Concentration 50; D037102:Lectins / Q000737:chemistry* / Q000276:immunology; D008081:Liposomes / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D014784:Mistletoe / Q000737:chemistry* / Q000276:immunology; D037121:Plant Lectins; D010946:Plants, Medicinal; D011487:Protein Conformation; D017510:Protein Folding; D012276:Ricin / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D037102:Lectins; D008081:Liposomes; D037121:Plant Lectins; D012276:Ricin; D019791:Guanidine