title: Affinity of native kappa-bungarotoxin and site-directed mutants for the muscle nicotinic acetylcholine receptor.
abstract: kappa-Bungarotoxin (kappa-bgt) is a 66-residue peptide originally purified from snake venom that acts as an antagonist at certain acetylcholine receptors. It is one of four homologous kappa-neurotoxins that are distinguished from the structurally related alpha-neurotoxins by their ability to block the alpha 3-subunit-containing neuronal nicotinic acetylcholine receptor (nAChR). It has been reported that venom-purified kappa-bgt also displays some affinity for the alpha 1-subunit-containing muscle nAChR to which the alpha-neurotoxins bind with high affinity. Here we report the effects of particular mutations on the ability of recombinant kappa-bgt to block the binding of 125I-alpha-bgt to nAChRs found in fetal mouse muscle and chick skeletal muscle. While the replacement of a proline residue found in all kappa-neurotoxins with an alanine (P-42-A) has relatively little effect, the introduction of a lysine, which is found in 90% of active alpha-neurotoxins at the same position (P-42-K), eliminates muscle receptor affinity at the concentrations tested. In contrast, the replacement of a glutamine in kappa-bgt with a tryptophan found in all active alpha-neurotoxins (Q-32-W) increases the affinity of kappa-bgt for the muscle receptor. When the arginine residue found in all active alpha- and kappa-neurotoxins is replaced by an alanine (R-40-A), the ability of kappa-bgt to block the muscle receptor is reduced to undetectable levels.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St. Louis, Missouri 63110.
authors.forename: J J
authors.initials: JJ
authors.lastname: Fiordalisi
authors.forename: R
authors.initials: R
authors.lastname: al-Rabiee
authors.forename: V A
authors.initials: VA
authors.lastname: Chiappinelli
authors.forename: G A
authors.initials: GA
authors.lastname: Grant
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002038:Bungarotoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002642:Chick Embryo; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005973:Glutamine / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000196:embryology / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011392:Proline / Q000737:chemistry; D011784:Quail; D011978:Receptors, Nicotinic / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D012910:Snake Venoms / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry
chemical_list: D002038:Bungarotoxins; D011978:Receptors, Nicotinic; D011994:Recombinant Proteins; D012910:Snake Venoms; D005973:Glutamine; D014364:Tryptophan; D011392:Proline; D008239:Lysine; D000409:Alanine