The molecular interaction of human salivary histatins with polyphenolic compounds

Dietary tannins are polyphenols that are effectively precipitated by saliva ry histatins (Hsts), a novel family of tannin binding proteins. Epigallocat echin gallate (EGCG), a flavan-3-ol ester related to condensed tannins (pol ymerized products of flavan-3-ols), and pentagalloyl glucose (PGG), a hydro lyzable tannin, were used to evaluate the molecular nature of Hst-polypheno l interaction. NMR demonstrated that Hst5, a representative Hst, bound to E GCG in a hydrophobic manner via basic and aromatic residues. In contrast, p roline plays a dominant role in polyphenol binding to other tannin precipit ating proteins. The role of basic and aromatic amino acids in EGCG binding was investigated using a series of modified Hsts in each of which one type of amino acid was substituted by Ala. EGCG bound to all modified Hsts, but the binding was diminished. Optimal EGCG binding also depended on the prima ry structure, as a polypeptide with randomised Hst5 sequence showed signifi cantly diminished interaction with EGCG. Soluble EGCG/Hst5 complexes contai ning up to seven molecules of EGCG per mol of Hst5 had a 1-mm dissociation constant. In contrast to EGCG, PGG formed small soluble complexes with Hst5 consisting of only one molecule each of PGG and Hst5, as demonstrated by a nalytical ultracentrifugation. These complexes became insoluble upon bindin g of additional molecules of PGG. Diminished PGG binding was seen to a pept ide with a Hst5 randomized sequence showing the importance of the primary s tructure. Hsts may serve to form insoluble complexes with tannins thereby p reventing their absorption from the intestines and potentially harmful biol ogical effects. In contrast the much weaker interaction with EGCG may allow its uptake into the organism and exploitation of its antioxidant effect.