pH-Dependent Solution Structure and Activity of a Reduced Form of the Host-Defense Peptide Myticin C (Myt C) from the Mussel Mytilus galloprovincialis

Myticin C (Myt C) is a highly variable host-defense peptide (HDP) associated to the immune response in the mediterranean mussel (Mytilus galloprovincialis), which has shown to be active across species due to its strong antiviral activity against a fish rhabdovirus found in fish cells overexpressing this HDP. However, the potential antimicrobial properties of any synthetic analogue of Myt C has not yet been analysed. Thus, in this work we have synthesised the sequence of the mature peptide of Myt C variant c and analysed the structure activity relationships of its reduced (non-oxidized) form (red-MytCc). In contrast to results previously reported for oxidized isoforms of mussel myticins, red-MytCc was not active against bacteria at physiological pH and showed a moderate antiviral activity against the viral haemorrhagic septicaemia (VHS) rhabdovirus. However, its chemotactic properties remained active. Structure/function studies in neutral and acid environments by means of infrared spectroscopy indicated that the structure of red-MytCc is pH dependent, with acid media increasing its alpha-helical content. Furthermore, red-MytCc was able to efficiently aggregate artificial phospholipid membranes at low pH, as well as to inhibit the Escherichia coli growth, suggesting that this activity is attributable to its more structured form in an acidic environment. All together, these results highlight the dynamic and environmentally sensitive behavior of red-Myt C in solution, and provide important insights into Myt C structure/activity relationships and the requirements to exert its antimicrobial/immunomodulatory activities. On the other hand, the pH-dependent direct antimicrobial activity of Myt C suggests that this HDP may be a suitable template for the development of antimicrobial agents that would function selectively in specific pH environments, which are sorely needed in this “antibiotic-resistance era”.     

厚壳贻贝抗菌肽mytilin和myticin的cDNA基因的克隆与序列分析

抗菌肽是贻贝免疫系统中的一个重要组成部分,也是目前海洋生物抗菌肽研究的重要内容。在贻贝抗菌肽家族中,mytilin和myticin是目前贻贝抗菌肽研究中发现成员最多,丰度最高的两个家族,在贻贝免疫系统中占有重要的地位。为了解厚壳贻贝抗菌肽mytilin和myticin的基因组成及其多样性机制,通过设计特异性引物,以厚壳贻贝血细胞cDNA文库为模板进行PCR扩增以筛选文库,共克隆到8条mytilin的同源cDNA序列和9条myticin的同源cDNA序列,并对其基因序列及推导的氨基酸序列进行了序列比对和变异位点初步分析。上述研究为将来深入研究厚壳贻贝的免疫机制及筛选具有开发价值的抗菌肽分子奠定了理论基础。