Functional expression and purification of recombinant Tx1, a sodium channel blocker neurotoxin from the venom of the Brazilian "armed" spider, Phoneutria nigriventer.
Tx1 from the venom of the Brazilian spider, Phoneutria nigriventer, is a lethal neurotoxic polypeptide of M(r) 8600 Da with 14 cysteine residues. It is a novel sodium channel blocker which reversibly inhibits sodium currents in CHO cells expressing recombinant sodium (Nav1.2) channels. We cloned and expressed the Tx1 toxin as a thioredoxin fusion product in the cytoplasm of Escherichia coli. After semipurification by immobilized Ni-ion affinity chromatography, the recombinant Tx1 was purified by reverse phase chromatography and characterized. It displayed similar biochemical and pharmacological properties to the native toxin, and it should be useful for further investigation of structure-function relationship of Na channels.
Diniz
Marcelo R V
MR
Centro de Pesquisa e Desenvolvimento, Fundação Ezequiel Dias, Rua Conde Pereira Carneiro, 80, 30 510-010 Belo Horizonte (MG), Brazil. mdiniz@funed.mg.gov.br
Theakston
R David G
RD
Crampton
Julian M
JM
Nascimento Cordeiro
Marta do
Md
Pimenta
Adriano M C
AM
De Lima
Maria Elena
ME
Diniz
Carlos R
CR
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D001938:Brazil; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D007276:Injections, Intraventricular; D051379:Mice; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000096:biosynthesis / Q000302:isolation & purification* / Q000633:toxicity; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000633:toxicity; D012964:Sodium / Q000378:metabolism; D026941:Sodium Channel Blockers / Q000302:isolation & purification* / Q000378:metabolism / Q000633:toxicity; D015222:Sodium Channels / Q000187:drug effects* / Q000378:metabolism; D013045:Species Specificity; D013111:Spider Venoms / Q000737:chemistry / Q000235:genetics*; D013112:Spiders / Q000235:genetics*; D013329:Structure-Activity Relationship
D009479:Neuropeptides; D011994:Recombinant Proteins; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D013111:Spider Venoms; C064022:Tx1 neurotoxin; D012964:Sodium