Differential scanning fluorimetry measurement of protein stability changes upon binding to glycosaminoglycans: a screening test for binding specificity.
Anal Chem
; 82(9): 3796-802, 2010 May 01.
Article
en En
| MEDLINE
| ID: mdl-20353159
ABSTRACT
The interaction between glycosaminoglycans (GAGs) and proteins is important for the regulation of protein transport and activity. Here we present a novel method for the measurement of protein-GAG interactions suitable for high-throughput screening, able to discriminate between the interactions of a protein with GAGs of different structures. Binding of proteins to the GAG heparin, a proxy for sulfated regions of extracellular heparan sulfate, was found to enhance the stability of three test proteins, fibroblast growth factors (FGFs)-1, -2, and -18. Chemically modified heparins and heparin oligosaccharides of different lengths stabilized the three FGFs to different extents, depending on the pattern of sugar binding specificity. The method is based on a differential scanning fluorescence approach. It uses a Sypro Orange dye, which binds to exposed core residues of a denatured protein and results in an increased fluorescence signal. It is convenient, requiring low micromolar amounts of protein and ligand compared to other interaction assays, employing only a real-time polymerase chain reaction (PCR) instrument.
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Fluorometría
/
Glicosaminoglicanos
Tipo de estudio:
Diagnostic_studies
/
Screening_studies
Idioma:
En
Revista:
Anal Chem
Año:
2010
Tipo del documento:
Article
País de afiliación:
Reino Unido