Oral Health Cooperative Research Centre, Melbourne Dental School, The University of Melbourne, 720 Swanston Street, Victoria 3010, Australia.
Antimicrob Agents Chemother. 2011 Mar;55(3):1155-61. doi: 10.1128/AAC.00466-10. Epub 2010 Dec 20.
Porphyromonas gingivalis is a major pathogen associated with chronic periodontitis, an inflammatory disease of the supporting tissues of the teeth. The Arg-specific (RgpA/B) and Lys-specific (Kgp) cysteine proteinases of P. gingivalis are major virulence factors for the bacterium. In this study κ-casein(109-137) was identified in a chymosin digest of casein as an inhibiting peptide of the P. gingivalis proteinases. The peptide was synthesized and shown to inhibit proteolytic activity associated with P. gingivalis whole cells, purified RgpA-Kgp proteinase-adhesin complexes, and purified RgpB proteinase. The peptide κ-casein(109-137) exhibited synergism with Zn(II) against both Arg- and Lys-specific proteinases. The active region for inhibition was identified as κ-casein(117-137) using synthetic peptides. Kinetic studies revealed that κ-casein(109-137) inhibits in an uncompetitive manner. A molecular model based on the uncompetitive action and its synergistic ability with Zn(II) was developed to explain the mechanism of inhibition. Preincubation of P. gingivalis with κ-casein(109-137) significantly reduced lesion development in a murine model of infection.
牙龈卟啉单胞菌是一种与慢性牙周炎相关的主要病原体，慢性牙周炎是一种牙齿支持组织的炎症性疾病。牙龈卟啉单胞菌的 Arg 特异性（RgpA/B）和 Lys 特异性（Kgp）半胱氨酸蛋白酶是该细菌的主要毒力因子。在这项研究中，κ-酪蛋白（109-137）在凝乳酶消化酪蛋白中被鉴定为牙龈卟啉单胞菌蛋白酶的抑制肽。该肽被合成并显示出抑制与牙龈卟啉单胞菌全细胞、纯化的 RgpA-Kgp 蛋白酶-黏附复合物和纯化的 RgpB 蛋白酶相关的蛋白水解活性。肽 κ-酪蛋白（109-137）与 Zn(II) 对 Arg-和 Lys-特异性蛋白酶表现出协同作用。使用合成肽鉴定出抑制的活性区域为 κ-酪蛋白（117-137）。动力学研究表明，κ-酪蛋白（109-137）以非竞争性方式抑制。基于非竞争性作用及其与 Zn(II) 的协同能力，建立了一个分子模型来解释抑制机制。在感染的小鼠模型中，将 κ-酪蛋白（109-137）预孵育牙龈卟啉单胞菌可显著减少病变的发展。
