Biology subjects

Kagami, Y.

Publications and source records attributed to Kagami, Y..

3 recordsLinked to original sources

bioRxiv2026

Solid-Phase Synthesis of ProTide Fluorogenic Probes Enables Systematic Profiling of Carboxypeptidase Activity

Carboxypeptidases play diverse roles in physiological and pathological processes, yet comprehensive analysis of their activities in complex biological samples remains challenging. Here we report a solid-phase synthesis strategy for fluorogenic ProTide-based probes that enables systematic profiling of carboxypeptidase activities based on defined C-terminal amino acid motifs. By modular synthesis of dipeptide-fluorophore conjugates, we generated a focused probe set that revealed distinct substrate preferences among carboxypeptidases, including carboxypeptidase A and B family enzymes. Integration of these probes with a single-molecule enzyme activity assay allowed ultrasensitive detection of circulating carboxypeptidase activities in human blood samples. Application of this platform to clinical specimens demonstrated that specific carboxypeptidase activities are elevated in patients with pancreatic cancer compared with healthy controls, whereas closely related enzymes showed limited diagnostic value. These results establish a scalable chemical strategy for activity-based profiling of exopeptidases and highlight circulating carboxypeptidase activity as a functional enzymatic signature associated with pancreatic cancer.

biochemistry↗

bioRxiv2026

Synthesis based on covalent capture and release (SCCR): a programmable strategy for automated preparation of protease-activatable molecules

Enzyme-activatable chemical tools, including fluorogenic probes and prodrugs, are essential in chemical biology and targeted therapeutics but remain challenging to access in structurally diverse forms because their synthesis is often bespoke and difficult to standardize. Here, we introduce synthesis based on covalent capture and release (SCCR) as a programmable chemical strategy that enables the modular assembly of protease-activatable molecules through specifically designed protecting-group logic. The SCCR framework establishes a standardized capture-elongation- release workflow that decouples molecular diversification from individual synthetic optimization, thereby enabling automated preparation of complex libraries. Using this chemistry, we generated a diverse set of fluorogenic probes and applied them to single-molecule enzyme activity analyses to identify candidate activity-based biomarkers of liver diseases. The generality of the SCCR strategy was further demonstrated by extending the same chemical logic to the preparation of antibody-drug conjugate (ADC) linkers, allowing systematic evaluation of plasma stability and cytotoxic potential. By establishing a programmable capture-release chemistry for the synthesis of enzyme-activatable molecules, this work provides a generalizable chemical foundation for the scalable and automated construction of functional small-molecule tools across biological and translational research.

biochemistry↗

bioRxiv2025

Platform of single-molecule enzyme activity-based liquid biopsy for detection of pancreatic adenocarcinoma at early stages

Liquid biopsy analyzes biomolecules in biofluids to provide critical information for the early detection of diseases and the development of more effective treatment strategies. Among these biomolecules, the functional states of proteins offer particularly valuable insights due to their direct correlation with phenotypic changes. However, current approaches for assessing protein function in liquid biopsies are often limited by low detection sensitivity. Here, we present a liquid biopsy platform that analyzes single-molecule protease/peptidase activity to detect pathological alterations in enzyme function in blood samples. The platform demonstrates potential for identifying patients with early-stage (stage I-II) pancreatic ductal adenocarcinoma (PDAC).

biochemistry↗