A Protease-Responsive Polymer/Peptide Conjugate and Reversible Assembly of Silver Clusters for the Detection of Porphyromonas gingivalis Enzymatic Activity.
A Protease-Responsive Polymer/Peptide Conjugate and Reversible Assembly of Silver Clusters for the Detection of Porphyromonas gingivalis Enzymatic Activity.
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DOI：
10.1021/acsnano.3c05268
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发表时间：
2023-08
期刊：
影响因子：
17.1
通讯作者：
Maurice Retout;Lubna Amer;Wonjun Yim;Matthew N Creyer;Benjamin Lam;Diego F. Trujillo;J. Potempa;A. O’Donoghue;Casey Chen;J. Jokerst
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作者：
Maurice Retout;Lubna Amer;Wonjun Yim;Matthew N Creyer;Benjamin Lam;Diego F. Trujillo;J. Potempa;A. O’Donoghue;Casey Chen;J. Jokerst
我们报告的可逆聚集的银纳米粒子（AgNP）组件使用的组合的阳离子甘氨酸为基础的肽和硫封端的聚乙二醇（PEG）。通过光学方法和电子显微镜研究了聚集体的形成和解离。银簇的解离取决于肽序列和PEG大小。对于PEG，1 kDa的分子量对于解离是最佳的。这种解离方法最重要的特点是，它可以在复杂的生物流体中操作，如血浆，唾液，胆汁，尿液，细胞培养基，甚至海水，而不会显着降低性能。此外，肽-颗粒组装体是高度稳定的，并且当干燥和再溶解、冷冻和解冻或在日光下放置一个月时不降解（或在解离时表现出信号损失）。重要的是，PEG的解离能力可以通过肽可切割底物的缀合来降低。解离能力在酶的存在下恢复。基于这些发现，我们设计了一个PEG-肽杂合分子的牙龈卟啉单胞菌蛋白酶RgpB特异性。我们的动机是这种细菌是牙周炎的关键病原体，并且RgpB活性与包括阿尔茨海默病在内的慢性疾病相关。RgpB的体外检测限为100 pM RgpB。该系统用于测量龈沟液（GCF）样品中的RgpB，与牙龈卟啉单胞菌的PCR相比，检测率为40%，假阴性率为0%（n = 37）。PEG-肽和纳米颗粒解离方法的组合允许开发可以独立于介质组成操作的方便的蛋白酶感测。
We report the reversible aggregation of silver nanoparticle (AgNP) assemblies using the combination of a cationic arginine-based peptide and sulfur-capped polyethylene glycol (PEG). The formation and dissociation of the aggregates were studied by optical methods and electron microscopy. The dissociation of silver clusters depends on the peptide sequence and PEG size. A molecular weight of 1 kDa for PEG was optimal for the dissociation. The most important feature of this dissociation method is that it can operate in complex biofluids such as plasma, saliva, bile, urine, cell media, or even seawater without a significant decrease in performance. Moreover, the peptide-particle assemblies are highly stable and do not degrade (or express of loss of signal upon dissociation) when dried and resolubilized, frozen and thawed, or left in daylight for a month. Importantly, the dissociation capacity of PEG can be reduced via the conjugation of a peptide-cleavable substrate. The dissociation capacity is restored in the presence of an enzyme. Based on these findings, we designed a PEG-peptide hybrid molecule specific to the Porphyromonas gingivalis protease RgpB. Our motivation was that this bacterium is a key pathogen in periodontitis, and RgpB activity has been correlated with chronic diseases including Alzheimer's disease. The RgpB limit of detection was 100 pM RgpB in vitro. This system was used to measure RgpB in gingival crevicular fluid (GCF) samples with a detection rate of 40% with 0% false negatives versus PCR for P. gingivalis (n = 37). The combination of PEG-peptide and nanoparticles dissociation method allows the development of convenient protease sensing that can operate independently of the media composition.