Comparison of protein surface hydrophobicity measured at various pH valuesusing three different fluorescent probes

The influence of type of fluorescent probe on the surface hydrophobicity va lues determined for three native and heated proteins was assessed using unc harged 6-propionyl-2-(N,N-dimethylamino)naphthalene or PRODAN] versus anion ic aliphatic (cis-parinaric acid or CPA) and aromatic (1-anilinonaphthalene -8-sulfonic acid or ANS) probes. Surface hydrophobicities of whey protein i solate, beta-lactoglobulin, and bovine serum albumin under heated (80 degre es C for 30 min) and unheated conditions and at varying pH values (3.0, 5.0 , 7.0, and 9.0) were measured using ANS, CPA, and PRODAN. ANS and CPA yield ed opposing results for the effects of pH and heating on protein hydrophobi city. Hydrophobicity was lower at pH 3.0 than at other pH values for all pr oteins measured by PRODAN, whereas the values measured by ANS and CPA at pH 3.0 were quite high compared to those at other pH values, suggesting the i nfluence of electrostatic interactions on anionic probe-protein binding. Th ese results suggest that the presence or absence of a permanent charge as w ell as the aromatic and aliphatic nature of fluorescent probes can affect p rotein hydrophobicity values measured under various pH conditions.