Prasad Pragati K, Eizenshtadt Noa, Goliand Inna, Fellus-Alyagor Liat, Oren Roni, Golani Ofra, Motiei Leila, Margulies David
Department of Chemical and Structural Biology, Weizmann Institute of Science Rehovot, 7610001, Israel.
Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot, 7610001, Israel.
Mater Today Bio. 2023 May 23;20:100669. doi: 10.1016/j.mtbio.2023.100669. eCollection 2023 Jun.
Common methods to label cell surface proteins (CSPs) involve the use of fluorescently modified antibodies (Abs) or small-molecule-based ligands. However, optimizing the labeling efficiency of such systems, for example, by modifying them with additional fluorophores or recognition elements, is challenging. Herein we show that effective labeling of CSPs overexpressed in cancer cells and tissues can be obtained with fluorescent probes based on chemically modified bacteria. The bacterial probes (B-probes) are generated by non-covalently linking a bacterial membrane protein to DNA duplexes appended with fluorophores and small-molecule binders of CSPs overexpressed in cancer cells. We show that B-probes are exceptionally simple to prepare and modify because they are generated from self-assembled and easily synthesized components, such as self-replicating bacterial scaffolds and DNA constructs that can be readily appended, at well-defined positions, with various types of dyes and CSP binders. This structural programmability enabled us to create B-probes that can label different types of cancer cells with distinct colors, as well as generate very bright B-probes in which the multiple dyes are spatially separated along the DNA scaffold to avoid self-quenching. This enhancement in the emission signal enabled us to label the cancer cells with greater sensitivity and follow the internalization of the B-probes into these cells. The potential to apply the design principles underlying B-probes in therapy or inhibitor screening is also discussed here.
标记细胞表面蛋白（CSPs）的常用方法包括使用荧光修饰抗体（Abs）或基于小分子的配体。然而，优化此类系统的标记效率具有挑战性，例如通过用额外的荧光团或识别元件对其进行修饰。在此我们表明，基于化学修饰细菌的荧光探针能够有效标记在癌细胞和组织中过表达的CSPs。细菌探针（B-探针）是通过将细菌膜蛋白与连接有荧光团和癌细胞中过表达的CSPs小分子结合剂的DNA双链非共价连接而产生的。我们表明B-探针制备和修饰异常简单，因为它们由自组装且易于合成的组件生成，如自我复制的细菌支架和DNA构建体，这些构建体可在明确的位置轻松连接各种类型的染料和CSP结合剂。这种结构可编程性使我们能够创建可使用不同颜色标记不同类型癌细胞的B-探针，以及生成非常明亮的B-探针，其中多种染料沿DNA支架在空间上分离以避免自猝灭。发射信号的这种增强使我们能够以更高的灵敏度标记癌细胞，并追踪B-探针内化进入这些细胞的过程。本文还讨论了将B-探针的设计原理应用于治疗或抑制剂筛选的潜力。
