Evidence that L1AD3, an apoptosis-inducing cyclic peptide, binds a leukemic T-cell membrane protein receptor.
Human leukemic T-lymphocytes undergo extensive and rapid apoptosis in the presence of L1AD3, a small cyclic peptide derivative of cobra cardiotoxin. The first step in this process involves its binding to membranes of susceptible cells. By the use of a biotin "handle" synthetically incorporated at the N-terminus of L1AD3, we show that binding is saturable and selective: normal human peripheral blood lymphocytes do not bind this peptide. Fluorescence resonance energy transfer experiments indicate that the binding sites are separated by at least 55 A. Loss of binding occurs if membrane proteins are enzymatically degraded, suggesting that L1AD3's target is a cell-membrane surface protein receptor. Finally, crosslinking of cyclic BTNL1AD3 peptide to a leukemic T-cell membrane surface receptor, as examined using a biotin-avidin blot, indicated a molecular weight of approximately 34,400.
Smith
Charles A
CA
Department of Medicinal and Biological Chemistry, College of Pharmacy, The University of Toledo, 2801 W. Bancroft, Wolfe Hall, Toledo, OH 43606, USA. charlesasmithphd@buckeye-express.com
Hinman
Channing L
CL
D000818:Animals; D017209:Apoptosis; D001360:Avidin / Q000737:chemistry; D001402:B-Lymphocytes / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D001710:Biotin / Q000737:chemistry; D019921:Biotinylation; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D002851:Chromatography, High Pressure Liquid; D004179:Cobra Cardiotoxin Proteins / Q000737:chemistry* / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004220:Disulfides; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D015458:Leukemia, T-Cell / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism; D013058:Mass Spectrometry; D008956:Models, Chemical; D008958:Models, Molecular; D009000:Monocytes / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D013601:T-Lymphocytes / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology
D004179:Cobra Cardiotoxin Proteins; D003432:Cross-Linking Reagents; D004220:Disulfides; C494064:L1AD3 peptide, Naja naja atra; D010455:Peptides; D001360:Avidin; D001710:Biotin; D014357:Trypsin; D010100:Oxygen