RESUMEN
Limited proteolysis of the Escherichia coli cell division inhibitor MinC reveals that its dimerization function resides in a structurally autonomous C-terminal domain. We show that cytoplasmic MinC is poised near the monomer-dimer equilibrium and propose that it only becomes entirely dimeric once recruited to the membrane by MinD.
Asunto(s)
Proteínas Bacterianas/metabolismo , Escherichia coli/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/química , División Celular , Dimerización , Escherichia coli/citología , Espectrometría de Masas , Datos de Secuencia Molecular , Fragmentos de Péptidos/análisis , Fragmentos de Péptidos/química , Prolina , Análisis de Secuencia de ProteínaRESUMEN
delta-Atracotoxins (delta-ACTXs) are peptide toxins isolated from the venom of Australian funnel-web spiders that slow sodium current inactivation in a similar manner to scorpion alpha-toxins. We have isolated and determined the amino acid sequence of a novel delta-ACTX, designated delta-ACTX-Hv1b, from the venom of the funnel-web spider Hadronyche versuta. This 42 residue toxin shows 67% sequence identity with delta-ACTX-Hv1a previously isolated from the same spider. Under whole-cell voltage-clamp conditions, the toxin had no effect on tetrodotoxin (TTX)-resistant sodium currents in rat dorsal root ganglion neurones but exerted a concentration-dependent reduction in peak TTX-sensitive sodium current amplitude accompanied by a slowing of sodium current inactivation similar to other delta-ACTXs. However, delta-ACTX-Hv1b is approximately 15-30-fold less potent than other delta-ACTXs and is remarkable for its complete lack of insecticidal activity. Thus, the sequence differences between delta-ACTX-Hv1a and -Hv1b provide key insights into the residues that are critical for targeting of these toxins to vertebrate and invertebrate sodium channels.
Asunto(s)
Bloqueadores de los Canales de Sodio , Venenos de Araña/química , Venenos de Araña/aislamiento & purificación , Venenos de Araña/farmacología , Arañas/química , Toxinas Biológicas/aislamiento & purificación , Toxinas Biológicas/farmacología , Secuencia de Aminoácidos , Animales , Cromatografía Líquida de Alta Presión , Relación Dosis-Respuesta a Droga , Conductividad Eléctrica , Femenino , Ganglios Espinales/citología , Ganglios Espinales/efectos de los fármacos , Gryllidae/efectos de los fármacos , Insecticidas/química , Activación del Canal Iónico/efectos de los fármacos , Masculino , Datos de Secuencia Molecular , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Ratas , Alineación de Secuencia , Análisis de Secuencia de Proteína , Sodio/metabolismo , Canales de Sodio/metabolismo , Especificidad de la Especie , Especificidad por Sustrato , Tetrodotoxina/farmacología , Toxinas Biológicas/químicaRESUMEN
We have isolated and determined the amino acid sequence of a novel peptide component from the venom of the Australian funnel-web spider Hadronyche versuta. This 68-residue toxin, ACTX-Hvf17, does not function like classical neurotoxins in modulating ion channel function as evidenced by its lack of insecticidal activity and its inability to affect vertebrate smooth or skeletal muscle contractility. The peptide shows significant sequence homology with mamba intestinal toxin 1 (MIT1) and to a lesser extent with a variety of colipases. The strong structural homology between MIT1 and porcine colipase leads us to propose that ACTX-Hvf17 also adopts the MIT1/colipase three-dimensional fold. However, we show that ACTX-Hvf17 has no colipase activity and does not stimulate muscle contractility like MITI. We also show that MIT1 and ACTX-Hvf17 display significant sequence homology with the C-terminal cysteine-rich domain of the Dickkopf-1 family of proteins that induce head formation in developing embryos, which leads us to propose that this domain of Dickkopf-1 also adopts the MIT1 colipase fold.