Work

Tracking protein transitions through fluorescence spectral phasor analysis with ACDAN

Year: 2025

Type: article

Abstract: This study investigates the use of spectral phasor analysis, hyperspectral imaging, and 6-acetyl-2-dimethylaminonaphthalene (ACDAN) fluorescence to explore key protein transitions: unfolding, amyloid aggregation, and liquid-liquid phase separation. We show that ACDAN fluorescence can sensitively detect subtle conformational changes before the complete protein unfolds, revealing early microenvironmental shifts. During amyloid formation, ACDAN identifies solvent dipolar relaxation events undetectable by conventional thioflavin T, providing critical insight into early aggregation events. Additionally, we map the physicochemical properties of protein biocondensates and highlight distinct microenvironments within these condensates, emphasizing the significance of dipolar relaxation in phase-separated systems. The approach provides a flexible and user-friendly toolkit for studying protein transitions, which can be easily implemented in commercial spectrofluorometers and microscopes.

Source: Biophysical Reports

DOI: 10.1016/j.bpr.2025.100209

Authors: Leandro Cruz Rodríguez, Nahuel Naum Foressi, María Soledad Celej

Institutions: Consejo Nacional de Investigaciones Científicas y Técnicas, Universidad Nacional de Córdoba, Research Centre in Biological Chemistry of Córdoba

Language: English

Topic: Protein Structure and Dynamics

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: gold

Estimated APC paid: $2,400

Funders: Consejo Nacional de Investigaciones Científicas y Técnicas, Secretaria de Ciencia y Tecnología - Universidad Nacional de Córdoba, Agencia Nacional de Promoción Científica y Tecnológica, Fondo para la Investigación Científica y Tecnológica