Migliolo Ludovico, Felício Mário R, Cardoso Marlon H, Silva Osmar N, Xavier Mary-Ann E, Nolasco Diego O, de Oliveira Adeliana Silva, Roca-Subira Ignasi, Vila Estape Jordi, Teixeira Leandro D, Freitas Sonia M, Otero-Gonzalez Anselmo J, Gonçalves Sónia, Santos Nuno C, Franco Octavio L
Centro de Análises Proteômicas e Bioquímicas, Programa de Pós-Graduação em Ciências Genômicas e Biotecnologia, UCB, Brasília, DF, Brazil; S-inova Biotech, Programa de Pós-Graduação em Biotecnologia, Universidade Católica Dom Bosco, Campo Grande, MS, Brazil.
Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
Biochim Biophys Acta. 2016 Jul;1858(7 Pt A):1488-98. doi: 10.1016/j.bbamem.2016.04.003. Epub 2016 Apr 8.
Recently, several peptides have been studied regarding the defence process against pathogenic microorganisms, which are able to act against different targets, with the purpose of developing novel bioactive compounds. The present work focuses on the structural and functional evaluation of the palindromic antimicrobial peptide Pa-MAP2, designed based on the peptide Pa-MAP from Pleuronectes americanus. For a better structural understanding, molecular modelling analyses were carried out, together with molecular dynamics and circular dichroism, in different media. Antibacterial activity against Gram-negative and positive bacteria was evaluated, as well as cytotoxicity against human erythrocytes, RAW 264.7, Vero and L6 cells. In silico docking experiments, lipid vesicle studies, and atomic force microscopy (AFM) imaging were carried out to explore the activity of the peptide. In vivo studies on infected mice were also done. The palindromic primary sequence favoured an α-helix structure that was pH dependent, only present on alkaline environment, with dynamic N- and C-terminals that are stabilized in anionic media. Pa-MAP2 only showed activity against Gram-negative bacteria, with a MIC of 3.2 μM, and without any cytotoxic effect. In silico, lipid vesicles and AFM studies confirm the preference for anionic lipids (POPG, POPS, DPPE, DPPG and LPS), with the positively charged lysine residues being essential for the initial electrostatic interaction. In vivo studies showed that Pa-MAP2 increases to 100% the survival rate of mice infected with Escherichia coli. Data here reported indicated that palindromic Pa-MAP2 could be an alternative candidate for use in therapeutics against Gram-negative bacterial infections.
最近，人们对几种肽在抵抗病原微生物的防御过程中的作用进行了研究，这些肽能够作用于不同靶点，目的是开发新型生物活性化合物。目前的工作重点是对基于美洲黄盖鲽的肽Pa-MAP设计的回文抗菌肽Pa-MAP2进行结构和功能评估。为了更好地理解其结构，在不同介质中进行了分子建模分析、分子动力学和圆二色性研究。评估了其对革兰氏阴性菌和阳性菌的抗菌活性，以及对人红细胞、RAW 264.7、Vero和L6细胞的细胞毒性。进行了计算机对接实验、脂质囊泡研究和原子力显微镜（AFM）成像，以探索该肽的活性。还对感染小鼠进行了体内研究。回文一级序列有利于形成pH依赖的α-螺旋结构，仅在碱性环境中存在，其动态的N端和C端在阴离子介质中稳定。Pa-MAP2仅对革兰氏阴性菌有活性，最低抑菌浓度为3.2 μM，且无任何细胞毒性作用。计算机模拟、脂质囊泡和AFM研究证实了其对阴离子脂质（POPG、POPS、DPPE、DPPG和LPS）的偏好，带正电荷的赖氨酸残基对于初始静电相互作用至关重要。体内研究表明，Pa-MAP2可使感染大肠杆菌的小鼠存活率提高到100%。此处报道的数据表明，回文Pa-MAP2可能是用于治疗革兰氏阴性菌感染的替代候选物。
