Identification of activity-based biomarkers for early-stage pancreatic tumors in blood using single-molecule enzyme activity screening.
Cell Rep Methods
; 4(1): 100688, 2024 Jan 22.
Article
in En
| MEDLINE
| ID: mdl-38218189
ABSTRACT
Single-molecule enzyme activity-based enzyme profiling (SEAP) is a methodology to globally analyze protein functions in living samples at the single-molecule level. It has been previously applied to detect functional alterations in phosphatases and glycosidases. Here, we expand the potential for activity-based biomarker discovery by developing a semi-automated synthesis platform for fluorogenic probes that can detect various peptidases and protease activities at the single-molecule level. The peptidase/protease probes were prepared on the basis of a 7-amino-4-methylcoumarin fluorophore. The introduction of a phosphonic acid to the core scaffold made the probe suitable for use in a microdevice-based assay, while phosphonic acid served as the handle for the affinity separation of the probe using Phos-tag. Using this semi-automated scheme, 48 fluorogenic probes for the single-molecule peptidase/protease activity analysis were prepared. Activity-based screening using blood samples revealed altered single-molecule activity profiles of CD13 and DPP4 in blood samples of patients with early-stage pancreatic tumors. The study shows the power of single-molecule enzyme activity screening to discover biomarkers on the basis of the functional alterations of proteins.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Pancreatic Neoplasms
/
Peptide Hydrolases
/
Phosphorous Acids
Type of study:
Diagnostic_studies
/
Screening_studies
Limits:
Humans
Language:
En
Journal:
Cell Rep Methods
Year:
2024
Document type:
Article
Affiliation country:
Japan