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Development and Characterization of Polymeric Peptides for Antibody Tagging of Bacterial Targets.
Protein & Peptide Letters ( IF 1.0 ) Pub Date : 2020-09-30 , DOI: 10.2174/0929866527666200427212940
Fresthel Monica M Climacosa 1, 2 , Ruby Anne N King 2 , Bobbie Marie M Santos 2, 3 , Salvador Eugenio C Caoili 2
Affiliation  

Background: Microbe-Binding Peptides (MBPs) are currently being investigated to address the problem of antimicrobial resistance. Strategies enhancing their antimicrobial activity have been developed, including peptide dimerization. Here, we present an alternative approach based on peptide polymerization, yielding hapten-labelled polymeric MBPs that mediate tagging of bacteria with anti-hapten antibodies, for enhanced immune recognition by host phagocytes.

Methods: C-terminally amidated analogs of the bacterial-binding peptide IIGGR were synthesized, with or without addition of cysteine residues at both N- and C-termini. Peptides were subjected to oxidizing conditions in a dimethyl-sulfoxide/water solvent system, and polymerization was demonstrated using SDS-PAGE. Peptides were then N-terminally labelled with a trinitrophenyl (TNP) group using trinitrobenzene sulfonate (TNBS). Binding to representative bacteria was demonstrated by ELISA using anti-TNP antibodies and was quantified as half-maximal effective concentration (EC50). Minimum Inhibitory Concentration (MIC) and concentration yielding 50% hemolysis (H50) were estimated. Neutrophil phagocytic index was determined for TNP-labelled polymeric bacterial- binding peptide (Pbac) with anti-TNP antibodies and/or serum complement.

Results: Polydisperse Pbac was synthesized. EC50 was lower for Pbac than for the corresponding monomeric form (Mbac), for both Staphylococcus aureus ATCC 29213 and Escherichia coli ATCC 25922. MIC and H50 were >250μg/mL for both Pbac and Mbac. A complement-independent increase in neutrophil phagocytic index was observed for E. coli treated with TNP-labelled Pbac in conjunction with anti-TNP antibodies.

Conclusion: Our data suggest that hapten-labelled polymeric bacterial-binding peptides may easily be produced from even crude synthetic oligopeptide precursors, and that such bacterial-binding peptides in conjunction with cognate anti-hapten antibodies can enhance immune recognition of bacteria by host phagocytes.



中文翻译:

用于细菌靶标抗体的聚合物肽的开发和表征。

背景:目前正在研究微生物结合肽(MBP),以解决抗药性问题。已经开发出增强其抗菌活性的策略,包括肽二聚化。在这里,我们提出了一种基于肽聚合的替代方法,可产生半抗原标记的聚合物MBP,以抗半抗原抗体介导细菌的标签,以增强宿主吞噬细胞的免疫识别能力。

方法:合成细菌结合肽IIGGR的C末端酰胺化类似物,在N和C末端均添加或不添加半胱氨酸残基。使肽在二甲基亚砜/水溶剂体系中经受氧化条件,并使用SDS-PAGE证明聚合。然后使用三硝基苯磺酸盐(TNBS)在肽的N端用三硝基苯基(TNP)基团标记。使用抗TNP抗体的ELISA证实了与代表性细菌的结合,并量化为最大有效浓度的一半(EC50)。估计了最小抑制浓度(MIC)和产生50%溶血的浓度(H50)。测定具有抗TNP抗体和/或血清补体的TNP标记的聚合物细菌结合肽(Pbac)的中性粒细胞吞噬指数。

结果:合成了多分散Pbac。对于金黄色葡萄球菌ATCC 29213和大肠杆菌ATCC 25922,Pbac的EC50均低于相应的单体形式(Mbac)。Pbac和Mbac的MIC和H50均大于250μg/ mL。对于用TNP标记的Pbac结合抗TNP抗体处理的大肠杆菌,观察到嗜中性粒细胞吞噬指数的补体非依赖性增加。

结论:我们的数据表明,即使是粗合成的寡肽前体也很容易产生半抗原标记的聚合物细菌结合肽,并且这种细菌结合肽与相关的抗半抗原抗体结合可以增强宿主吞噬细胞对细菌的免疫识别。

更新日期:2020-11-04
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