Structural and functional characterization of a novel ShK peptide and its analogue from zoanthids (corals)

摘要

OBJECTIVE To identify and characterize a novel neuroprotective ShK peptide and its analogue originated from coral P.caribaeoru.METHODS P.caribaeoru was collected and subjected to transcriptome sequencing at which further bioinformatics analysis had identified a unigene encoding a putative ShK protein candidate,named as PcShK1.PcShK1 and its rhodamine derivative(PcShK1-RhoB) were synthesized and tested for the neuroprotective effect in 6-OHDA induced Parkinson disease models in vitro and in vivo.Briefly,zebrafish larvae were co-exposed to 6-OHDA and various doses of the peptides;then,dopaminergic(DA) neurons immunoreactivity and locomotion behavior of the zebrafish larvae were examined.Similarly,PC12 cells were cultured with 6-OHDA in the absence or presence of different concentrations of peptides.Cell viability was determined by MTT assay while calcium flow in the PC12 cells was monitored by Fluo-4 fluorescent dye.RESULTS Compared with control group,6-OHDA treatment could lead to DA neurons loss and locomotion deficits in PD model of zebrafish larvae(P<0.01).Both PcS hK 1(2.5,5.0 and 7.5 μmol·L^(-1)) and PcS hK 1-RhoB(0.50 and 0.75 μmol·L^(-1)) were found to protect and restore dopaminergic neurons from6-OHDA mediated injury and locomotion deficiency in the PD zebrafish respectively(P<0.01).In addition,PcShK1(2.5 to 20.0 μmol·L^(-1)) and PcShK1-RhoB(0.6 to 2.5 μmol·L^(-1)) effectively prevented against 6-OHDA toxicity in PC12 cells(P<0.01).Further study revealed that they might exert their neuroprotective effects through regulating the calcium homeostasis.CONCLUSION PcS hK 1 and PcS hK 1-RhoB show neuroprotective effects on 6-OHDA induced PD models,and the underlying protective mechanisms of these peptides probably involve calcium homeostasis regulation.