Effects of removing the negatively charged N-terminal region of the salivary acidic proline-rich proteins by human leucocyte elastase

Human leucocyte elastase from inflammatory gingival crevicular exudates (gi ngival crevicular fluid) contacts saliva and saliva-coated tooth surfaces c oronal to the gingival margin, Major components of saliva are the salivary acidic proline-rich proteins (PRPs). These acidic PRPs, via the numerous ne gatively charged amino acid residues located predominantly within their ami no-terminal region. bind to the hydroxyapatite mineral of the tooth surface and become part of the salivary pellicle. Thus the potential for human leu cocyte elastase-mediated removal of the negatively charged amino-terminal r egion of acidic PRP variants (PRP-1, PRP-2, PRP-3, PRP-4, PIF-s and PIF-f) was examined. It was determined that each of the acidic PRP variants was su sceptible to fragmentation by human leucocyte elastase, in which the 16 ami no-terminal segment was removed, leaving the respective residual fragment n amed as the transitional product (tr). The transitional products were terme d PRP-ltr, PRP-2tr (PIF-str), PRP-3tr and PRP-ltr (PIF-ftr). Each of the re sidual transitional products of acidic PRP had an amino-terminal beginning with serine residue no. 17, determined by amino acid sequencing. When sampl es of human leucocyte elastase-treated acidic PRPs were placed on native po lyacrylamide gels and electrophoresed, the respective transitional products moved more slowly than the parental acidic PRP molecules, reflecting the l oss of a portion of the negatively charged section. In comparison to the ac idic PRPs, the acidic PRP transitional products had markedly reduced bindin g to hydroxyapatite. The transitional products were resistant to further en zymatic digestion as a function of increased incubation time and appeared t o exert an antihuman leucocyte elastase effect. However, when increased con centrations of human leucocyte elastase were incubated with the acidic PRP, a mure extensive digestion occurred, leaving a residual peptide with an am ino-terminal beginning with alanine residue no. 44. interestingly, intact a cidic PRPs if prebound to hydroxyapatite particles. resisted digestion by h uman leucocyte: elastase. In summary, human leucocyte elastase was capable of digesting fluid-phase (unbound) acidic PRP in a manner that eliminated p arr of their negatively charged region, which subsequently reduced their bi nding to hydroxyapatite. High concentrations of human leucocyte elastase, a rriving from inflammatory gingival crevicular exudates, may interrupt the n ormal binding of fluid-phase acidic PRPs to hydroxyapatite. (C) 1999 Elsevi er Science Ltd. All rights reserved.