Lee Richard V, Zareie Hadi M, Sarikaya Mehmet
GEMSEC, Genetically Engineered Materials Science and Engineering Center, Department of Materials Science and Engineering, University of Washington, Seattle, Washington 98195, United States.
ACS Appl Mater Interfaces. 2022 May 25;14(20):23164-23181. doi: 10.1021/acsami.2c03341. Epub 2022 May 11.
Nucleic acid biomarkers hold great potential as key indicators for the diagnosis and monitoring of diseases. Herein we design and implement bifunctional chimeric biomolecules composed of a solid-binding peptide (SBP) domain that specifically adsorbs onto solid sensor surfaces and a peptide nucleic acid (PNA) moiety that facilitates anchoring of antisense oligonucleotide (ASO) probes for the detection of nucleic acid targets. A gold-binding peptide, AuBP1, previously selected by directed evolution to specifically bind to gold, served as the basis for immobilizing nucleic acid probes onto gold substrates. Using surface plasmon resonance (SPR) spectroscopy and quartz crystal microbalance (QCM) analyses, we demonstrate the sequential biomolecular assembly of the heterofunctional solid-binding peptide-antisense oligomer (SBP-ASO) construct onto a sensor surface and the subsequent detection of DNA in an aqueous environment. The effect of steric hindrance on optimal probe assembly is observed, establishing that less packing density results in greater target capture efficacy. In addition, an adsorbed layer of chimeric solid-binding peptide-peptide nucleic acid (SBP-PNA) undergoes viscoelastic changes at the solid-liquid interface upon probe immobilization and DNA target capture, whereby the rigid biofunctional layer becomes more flexible. The dual nature of the chimeric construct is highly amenable to a variety of platforms allowing for both specific recognition and probe immobilization on the sensor surface, while the modular design of the solid-binding peptide-antisense oligonucleotide provides facile functionalization of a wide diversity of solid substrates.
核酸生物标志物作为疾病诊断和监测的关键指标具有巨大潜力。在此，我们设计并实现了双功能嵌合生物分子，其由特异性吸附到固体传感器表面的固体结合肽（SBP）结构域和促进反义寡核苷酸（ASO）探针锚定以检测核酸靶标的肽核酸（PNA）部分组成。一种先前通过定向进化筛选出的特异性结合金的金结合肽AuBP1，作为将核酸探针固定到金基底上的基础。使用表面等离子体共振（SPR）光谱和石英晶体微天平（QCM）分析，我们展示了异功能固体结合肽 - 反义寡聚物（SBP - ASO）构建体在传感器表面上的顺序生物分子组装以及随后在水性环境中对DNA的检测。观察到空间位阻对最佳探针组装的影响，确定较低的堆积密度导致更高的靶标捕获效率。此外，嵌合固体结合肽 - 肽核酸（SBP - PNA）的吸附层在探针固定和DNA靶标捕获时在固 - 液界面处发生粘弹性变化，由此刚性生物功能层变得更加灵活。嵌合构建体的双重性质非常适合各种平台，既允许在传感器表面进行特异性识别又允许探针固定，而固体结合肽 - 反义寡核苷酸的模块化设计为多种固体底物提供了简便的功能化。
