I3S, Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Portugal; INEB - Instituto de Engenharia Biomédica, Universidade do Porto, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; Universidade do Porto, Faculdade de Engenharia, Porto, Portugal.
CIQ-UP - Centro de Investigação em Química da Universidade do Porto, Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto, Porto, Portugal.
Biomaterials. 2015 Jun;52:531-8. doi: 10.1016/j.biomaterials.2015.02.049. Epub 2015 Mar 18.
Bacterial colonization and subsequent biofilm formation is still one of the major problems associated with medical devices. Antimicrobial peptides (AMP) immobilization onto biomaterials surface is a promising strategy to avoid bacterial colonization. However, a correct peptide orientation and exposure from the surface is essential to maintain AMP antimicrobial activity. This work aims to evaluate the effect of the immobilization on antibacterial activity of Dhvar5 (LLLFLLKKRKKRKY), an AMP with a head-to-tail amphipathicity. Dhvar5 was linked to thin chitosan coatings in i) a controlled orientation and exposure, testing covalent immobilization of its N- or C-terminus and using spacers with different lengths and flexibilities or in ii) a random orientation by physical adsorption. Chitosan coating was chosen due to its antimicrobial properties and readiness to be functionalized. Surface characterization demonstrated the chemoselective immobilization of the peptide with different spacers in a similar concentration (∼2 ng/cm2). Efficacy assays demonstrated that covalent immobilization of Dhvar5 exposing its cationic end, improves the chitosan coating antimicrobial effect by decreasing Methicillin-resistant Staphylococcus aureus (MRSA) colonization. This effect was enhanced when longer spacers were used independently of their flexibility. In opposite, immobilized Dhvar5 exposing its hydrophobic end has no effect on bacterial adhesion to chitosan, and when adsorbed in a random orientation even induces bacterial adhesion to chitosan coating.
细菌定植和随后的生物膜形成仍然是与医疗器械相关的主要问题之一。将抗菌肽（AMP）固定在生物材料表面是避免细菌定植的一种有前途的策略。然而，从表面正确定向和暴露肽对于维持 AMP 的抗菌活性至关重要。本工作旨在评估 Dhvar5（LLLFLLKKRKKRKY）固定化对其抗菌活性的影响，Dhvar5 是一种具有头对头两亲性的 AMP。Dhvar5 通过以下两种方式与薄壳聚糖涂层结合：i）控制定向和暴露，测试其 N 端或 C 端的共价固定化，并使用不同长度和柔韧性的间隔物，或 ii）通过物理吸附实现随机定向。选择壳聚糖涂层是因为其具有抗菌性能且易于功能化。表面特性表明，在相似浓度（约 2ng/cm2）下，不同间隔物的肽可以进行化学选择性固定化。功效测定表明，Dhvar5 的共价固定化暴露其阳离子端，通过减少耐甲氧西林金黄色葡萄球菌（MRSA）定植来提高壳聚糖涂层的抗菌效果。当使用更长的间隔物时，无论其柔韧性如何，这种效果都会增强。相反，暴露其疏水性端的固定化 Dhvar5 对细菌在壳聚糖上的黏附没有影响，而当以随机取向吸附时，甚至会诱导细菌在壳聚糖涂层上黏附。
