Characterization of a leucokinin binding protein in Aedes aegypti (Diptera: Culicidae) Malpighian tubule
Autor(es): Pietrantonio, P. V.; Gibsona, G. E.; Streya, A. A.; Petzel, D.; Hayesa, T. K.
Resumo: The insect myokinin (leucokinin-like) neuropeptide family includes peptides that have different physiological effects such as the induction of hindgut myotropic activity and stimulation of urine production. The C-terminal pentamer of myokinins, Phe-X-(Ser/Pro/Ala)-Trp-Gly-amide [X=Phe, His, Asn, Ser or Tyr], had been previously determined as the minimum fragment able to elicit a functional response. The receptor(s) for these insect neuropeptides has not yet been identified. In order to characterize the Malpighian tubule leucokinin-like peptide receptor(s) from the yellow fever mosquito (Aedes aegypti), a leucokinin photoaffinity analogue (LPA) of sequence dAla-dTyr-Bpa-dLys-Phe-Phe-Ser-Trp-Gly-amide was designed based on structure/activity relationships for leucokinins. LPA caused depolarization of the transepithelial voltage (TEV) in female Malpighian tubule, confirming the activity of the peptide. The effective concentration to give half the maximum depolarization (EC sub(50)) was 17 nM. The super(125)I-LPA was then used to characterize leucokinin binding proteins in female Malpighian tubule membranes. It specifically labeled and saturated a protein(s) of about 54 kDa as shown by SDS-PAGE/autoradiography and by competition experiments with excess unlabeled leucokinin analogues. super(125)I-LPA bound to the 54 kDa protein(s) with a K sub(d) value of 13 plus or minus 3 nM in agreement with the EC sub(50) for the TEV bioassay. Altogether these data suggest that the 54 kDa protein is an Aedes-leucokinin receptor. This is the first characterization of an insect leucokinin receptor and reveals that LPA is a powerful tool to label insect myokinin receptors.
Palavras-Chave: Neuropeptides; Leucokinin; Chemoreceptors; Proteins; Peptides; Hindgut; Urine; Malpighian tubules; Enzymatic Activity; Aedes Aegypti; Culicidae
Imprenta: Insect Biochemistry and Molecular Biology, v. 30, n. 12, p. 1147-1159, 2000.
Descritores: Aedes aegypti - Molecular Structure ; Aedes aegypti - Proteins
Data de publicação: 2000
