Salivary histatin 5 is an inhibitor of both host and bacterial enzymes implicated in periodontal disease.

ABSTRACT

One of the salient features of periodontitis and gingivitis is the increase in the levels of bacterial and host-derived proteolytic enzymes in oral inflammatory exudates. This study evaluated the potential of histatin 5, a 24-residue histidine-rich salivary antimicrobial protein, to inhibit these enzymes. Using biotinylated gelatin as a substrate, histatin 5 was found to inhibit the activity of the host matrix metalloproteinases MMP-2 and MMP-9 with 50% inhibitory concentrations (IC50s) of 0.57 and 0.25 microM, respectively. To localize the domain responsible for this inhibition, three peptides containing different regions of histatin 5 were synthesized and tested as inhibitors of MMP-9. Peptides comprising residues 1 to 14 and residues 4 to 15 of histatin 5 showed much lower inhibitory activities (IC50, 21.4 and 20.5 microM, respectively), while a peptide comprising residues 9 to 22 showed identical activity to histatin 5 against MMP-9. These results point to a functional domain localized in the C-terminal part of histatin 5. To evaluate the effect of histatin 5 on bacterial proteases, a detailed characterization of histatin 5 inhibition of gingipains from Porphyromonas gingivalis was carried out using purified Arg- and Lys-specific enzymes. Kinetic analysis of the inhibition of the Arg-gingipain revealed that histatin 5 is a competitive inhibitor, affecting only the Km with a K(i) of 15 microM. In contrast, inhibition of Lys-gingipain affected both the Km and Vmax, suggesting that both competitive and noncompetitive competitive processes underlie this inhibition. The inhibitory activity of histatin 5 against host and bacterial proteases at physiological concentrations points to a new potential biological function of histatin in the oral cavity.