Fletcher H M, Schenkein H A, Macrina F L
Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond 23298-0678.
Infect Immun. 1994 Oct;62(10):4279-86. doi: 10.1128/iai.62.10.4279-4286.1994.
Porphyromonas gingivalis has been implicated as a contributing etiological agent of adult periodontitis and generalized forms of early-onset periodontitis. Proteases of P. gingivalis may contribute to its pathogenicity by destroying connective tissue as well as inactivating key plasma proteins that might mediate protective host functions. In order to explore this problem, antiserum raised against membrane vesicles of P. gingivalis W83 was used to screen a genomic library of strain W83 constructed by using the lambda DASH vector system. A recombinant phage (lambda 34) expressing a P. gingivalis protease from the library was identified and characterized. Casein substrate zymography of lambda 34 lysates revealed a protease with an apparent molecular mass of 97 kDa. The gene encoding this protease was designated prtH. It was localized to a 3.7-kb HindIII-BamHI fragment and specified an enzyme which hydrolyzed the human C3 complement protein under defined conditions. The nucleotide sequence of this 3.7-kb fragment was determined, and one 2.9-kb open reading frame (992 amino acids) corresponding to a 110-kDa protein was detected, suggesting it might be a precursor of the 97-kDa active protease. prtH is not similar to any previously cloned protease gene from P. gingivalis.
牙龈卟啉单胞菌被认为是成人牙周炎和早发性牙周炎的一种致病因素。牙龈卟啉单胞菌的蛋白酶可能通过破坏结缔组织以及使可能介导宿主保护功能的关键血浆蛋白失活来促进其致病性。为了探究这个问题，用针对牙龈卟啉单胞菌W83膜泡产生的抗血清来筛选使用λDASH载体系统构建的菌株W83的基因组文库。从文库中鉴定并表征了一个表达牙龈卟啉单胞菌蛋白酶的重组噬菌体（λ34）。λ34裂解物的酪蛋白底物酶谱显示出一种表观分子量为97 kDa的蛋白酶。编码该蛋白酶的基因被命名为prtH。它定位于一个3.7 kb的HindIII - BamHI片段，并且确定了一种在特定条件下能水解人C3补体蛋白的酶。测定了这个3.7 kb片段的核苷酸序列，检测到一个对应于110 kDa蛋白质的2.9 kb开放阅读框（992个氨基酸），表明它可能是97 kDa活性蛋白酶的前体。prtH与先前从牙龈卟啉单胞菌克隆的任何蛋白酶基因都不相似。
