Site directed mutants of Noxiustoxin reveal specific interactions with potassium channels.
Several site directed mutations were introduced into a synthetic Noxiustoxin (NTX) gene. Alanine scanning of the nonapeptide at the N-terminal segment of NTX (threonine 1 (T1) to serine 9 (S9)) was constructed and the recombinant products were obtained in pure form. Additionally, lysine 28 (K28) was changed to arginine (R) or glutamic acid (E), cysteine 29 was changed to alanine, and residues 37-39 (Tyr-Asn-Asn) of the carboxyl end were deleted. The recombinant mutants were tested for their ability to displace 125I-NTX from rat brain synaptosome membranes, as well as for their efficiency in blocking the activity of Kv1.1 K+ channels expressed in Xenopus laevis oocytes. The main results indicate that residues K6, T8 at the amino end, and K28 and the tripeptide YNN at the carboxyl end are involved in specific interactions of NTX with rat brain and/or Kv1.1 K+ channels.
Martínez
F
F
Department of Molecular Recognition and Structural Biology, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca.
Muñoz-Garay
C
C
Gurrola
G
G
Darszon
A
A
Possani
L D
LD
Becerril
B
B
D000818:Animals; D001667:Binding, Competitive; D001921:Brain; D004252:DNA Mutational Analysis; D004305:Dose-Response Relationship, Drug; D051662:Kv1.1 Potassium Channel; D016297:Mutagenesis, Site-Directed; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000187:drug effects*; D024642:Potassium Channels, Voltage-Gated; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D012604:Scorpion Venoms / Q000235:genetics / Q000494:pharmacology*; D013574:Synaptosomes
D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011994:Recombinant Proteins; D012604:Scorpion Venoms; D051662:Kv1.1 Potassium Channel; C052567:noxiustoxin