Boackle R J, Dutton S L, Robinson W L, Vesely J, Lever J K, Su H R, Chang N S
Department of Stomatology, College of Dental Medicine, Medical University of South Carolina, Charleston 29425, USA.
Arch Oral Biol. 1999 Jul;44(7):575-85. doi: 10.1016/s0003-9969(99)00032-1.
Human leucocyte elastase from inflammatory gingival crevicular exudates (gingival crevicular fluid) contacts saliva and saliva-coated tooth surfaces coronal to the gingival margin. Major components of saliva are the salivary acidic proline-rich proteins (PRPs). These acidic PRPs, via the numerous negatively charged amino acid residues located predominantly within their amino-terminal region, bind to the hydroxyapatite mineral of the tooth surface and become part of the salivary pellicle. Thus the potential for human leucocyte elastase-mediated removal of the negatively charged amino-terminal region 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 susceptible to fragmentation by human leucocyte elastase, in which the 16 amino-terminal segment was removed, leaving the respective residual fragment named as the transitional product (tr). The transitional products were termed PRP-1tr, PRP-2tr (PIF-str), PRP-3tr and PRP-4tr (PIF-ftr). Each of the residual transitional products of acidic PRP had an amino-terminal beginning with serine residue no. 17, determined by amino acid sequencing. When samples of human leucocyte elastase-treated acidic PRPs were placed on native polyacrylamide gels and electrophoresed, the respective transitional products moved more slowly than the parental acidic PRP molecules, reflecting the loss of a portion of the negatively charged section. In comparison to the acidic PRPs, the acidic PRP transitional products had markedly reduced binding to hydroxyapatite. The transitional products were resistant to further enzymatic digestion as a function of increased incubation time and appeared to exert an antihuman leucocyte elastase effect. However, when increased concentrations of human leucocyte elastase were incubated with the acidic PRP, a more extensive digestion occurred, leaving a residual peptide with an amino-terminal beginning with alanine residue no. 44. Interestingly, intact acidic PRPs if prebound to hydroxyapatite particles, resisted digestion by human leucocyte elastase. In summary, human leucocyte elastase was capable of digesting fluid-phase (unbound) acidic PRP in a manner that eliminated part of their negatively charged region, which subsequently reduced their binding to hydroxyapatite. High concentrations of human leucocyte elastase, arriving from inflammatory gingival crevicular exudates, may interrupt the normal binding of fluid-phase acidic PRPs to hydroxyapatite.
来自炎症性牙龈沟渗出液（龈沟液）的人白细胞弹性蛋白酶会接触唾液以及位于龈缘冠方的唾液包被的牙面。唾液的主要成分是富含脯氨酸的唾液酸性蛋白（PRPs）。这些酸性PRPs通过主要位于其氨基末端区域的众多带负电荷的氨基酸残基，与牙面的羟基磷灰石矿物质结合，并成为唾液薄膜的一部分。因此，研究了人白细胞弹性蛋白酶介导去除酸性PRP变体（PRP-1、PRP-2、PRP-3、PRP-4、PIF-s和PIF-f）带负电荷的氨基末端区域的可能性。已确定每种酸性PRP变体都易受人白细胞弹性蛋白酶的裂解，其中16个氨基末端片段被去除，留下各自命名为过渡产物（tr）的残余片段。过渡产物被称为PRP-1tr、PRP-2tr（PIF-str）、PRP-3tr和PRP-4tr（PIF-ftr）。通过氨基酸测序确定，酸性PRP的每种残余过渡产物的氨基末端都以第17号丝氨酸残基开始。当将人白细胞弹性蛋白酶处理过的酸性PRPs样品置于天然聚丙烯酰胺凝胶上并进行电泳时，各自的过渡产物移动速度比亲本酸性PRP分子慢，这反映了一部分带负电荷部分的丢失。与酸性PRPs相比，酸性PRP过渡产物与羟基磷灰石的结合明显减少。随着孵育时间的增加，过渡产物对进一步的酶消化具有抗性，并且似乎发挥了抗人白细胞弹性蛋白酶的作用。然而，当将浓度增加的人白细胞弹性蛋白酶与酸性PRP一起孵育时，会发生更广泛的消化，留下一个氨基末端以第44号丙氨酸残基开始的残余肽段。有趣的是，如果完整的酸性PRPs预先与羟基磷灰石颗粒结合，则能抵抗人白细胞弹性蛋白酶的消化。总之，人白细胞弹性蛋白酶能够以消除其部分带负电荷区域的方式消化液相（未结合的）酸性PRP，这随后降低了它们与羟基磷灰石的结合。来自炎症性牙龈沟渗出液的高浓度人白细胞弹性蛋白酶可能会中断液相酸性PRPs与羟基磷灰石的正常结合。
