ANS fluorescence: potential to augment the identification of the external binding sites of proteins.
8-anilino-1-naphthalenesulfonic acid (ANS) is believed to strongly bind cationic groups of proteins and polyamino acids through ion pair formation. A paucity of data exists on the fluorescent properties of ANS in these interactions. ANS binding to arginine and lysine derivatives was studied by fluorescence and circular dichroism spectroscopies to augment published information attained by isothermal titration calorimetry (ITC). Fluorescence enhancement with a hypsochromic shift results from the interaction of the charged group of lysine and arginine with the sulfonate group of ANS. Ion pairing between Arg (or Lys) and the sulfonate group of ANS reduce the intermolecular charge transfer (CT) rate constant that leads to enhancement of fluorescence. A positive charge near the -NH group of ANS changes the intramolecular CT process producing a blue shift of fluorescence. The Arg side chain compared to that of Lys more effectively interacts with both the -NH and sulfonate groups of ANS. ANS binding also induces a random coil-alpha helix transition in poly-Arg. Our data, in contrast to ITC results, indicate that electrostatic interactions between ANS derivatives and positively charged side chains do not account for binding affinity in the micromolar range. In addition to ion pairing complementary interactions, such as van der Waals, should be considered for high affinity (K(d)<1 mM) external binding sites of proteins.
Gasymov
Oktay K
OK
Department of Pathology, UCLA School of Medicine, Jules Stein Eye Institute, 100 Stein Plaza, Los Angeles, CA 90095, USA.
Glasgow
Ben J
BJ
D000817:Anilino Naphthalenesulfonates / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008968:Molecular Conformation; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D017186:Titrimetry
C515594:8-anilino-1-naphthalenesulfonic acid; D000817:Anilino Naphthalenesulfonates; D011506:Proteins