Shmunis School of Biomedicine and Cancer Research, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
Department of Chemistry Materials, Bar-Ilan University, Ramat-Gan 5290002, Israel.
Proc Natl Acad Sci U S A. 2023 Oct 31;120(44):e2310569120. doi: 10.1073/pnas.2310569120. Epub 2023 Oct 23.
Most biocatalytic processes in eukaryotic cells are regulated by subcellular microenvironments such as membrane-bound or membraneless organelles. These natural compartmentalization systems have inspired the design of synthetic compartments composed of a variety of building blocks. Recently, the emerging field of liquid-liquid phase separation has facilitated the design of biomolecular condensates composed of proteins and nucleic acids, with controllable properties including polarity, diffusivity, surface tension, and encapsulation efficiency. However, utilizing phase-separated condensates as optical sensors has not yet been attempted. Here, we were inspired by the biosynthesis of melanin pigments, a key biocatalytic process that is regulated by compartmentalization in organelles, to design minimalistic biomolecular condensates with emergent optical properties. Melanins are ubiquitous pigment materials with a range of functionalities including photoprotection, coloration, and free radical scavenging activity. Their biosynthesis in the confined melanosomes involves oxidation-polymerization of tyrosine (Tyr), catalyzed by the enzyme tyrosinase. We have now developed condensates that are formed by an interaction between a Tyr-containing peptide and RNA and can serve as both microreactors and substrates for tyrosinase. Importantly, partitioning of Tyr into the condensates and subsequent oxidation-polymerization gives rise to unique optical properties including far-red fluorescence. We now demonstrate that individual condensates can serve as sensors to detect tyrosinase activity, with a limit of detection similar to that of synthetic fluorescent probes. This approach opens opportunities to utilize designer biomolecular condensates as diagnostic tools for various disorders involving abnormal enzymatic activity.
真核细胞中的大多数生物催化过程都受到亚细胞微环境的调节，如膜结合或无膜细胞器。这些天然的分隔系统启发了合成隔间的设计，这些隔间由各种构建块组成。最近，新兴的液-液相分离领域促进了由蛋白质和核酸组成的生物分子凝聚物的设计，其具有可控的性质，包括极性、扩散性、表面张力和封装效率。然而，利用相分离凝聚物作为光学传感器尚未尝试过。在这里，我们受到黑色素生物合成的启发，黑色素生物合成是一种受细胞器分隔调节的关键生物催化过程，设计了具有新兴光学特性的极简生物分子凝聚物。黑色素是一种普遍存在的色素材料，具有多种功能，包括光保护、着色和清除自由基活性。它们在受限的黑素体中的生物合成涉及酪氨酸（Tyr）的氧化聚合，由酶酪氨酸酶催化。我们现在已经开发出由含有 Tyr 的肽和 RNA 相互作用形成的凝聚物，它们可以作为微反应器和酪氨酸酶的底物。重要的是，Tyr 分配到凝聚物中并随后进行氧化聚合会产生独特的光学性质，包括远红荧光。我们现在证明，单个凝聚物可以作为传感器来检测酪氨酸酶的活性，其检测限与合成荧光探针相似。这种方法为利用设计的生物分子凝聚物作为涉及异常酶活性的各种疾病的诊断工具开辟了机会。
