Functional studies of streptococcal pyrogenic exotoxin B and its associated proteins S-adenosylhomocysteine hydrolase and keratin 14

• Main Authors: Ju-Fei Hsu, 許茹斐
• Other Authors: Wenya Huang
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/84293544967515897552

碩士 === 國立成功大學 === 醫學檢驗生物技術學系 === 94 === Streptococcus pyogenes is a common pathogen for pharyngitis, impetigo and myositis. It also causes severe streptococcal toxic shock syndrome (STSS), which is lethal in humans. GAS secrets a number of exotoxins, among which the streptococcal pyrogenic exotoxin B (SPE B) is highly associated with the disease severities. SPE B is a cysteine protease and matures itself by autocalalysis. To study the in vivo function of SPE B, we screened for the proteins that interact with it by yeast two-hybrid assays. We found that S-adenosylhomocysteine hydrolase (AdoHcyase), an essential factor for nucleotide metabolism, is associated with SPE B. Protein level of AdoHcyase was reduced in the human kidney fibroblast 293T cells infected with S. pyogenes NZ131 strain (wild-type); whereas they were not reduced in the cells infected with SW510 strain (spe B mutant). AdoHcyase has been shown to cleave the AdoHcy into the products of adenosine and homocysteine. By enzyme immunoassay, we found that the homocysteine level was significantly reduced in plasma of mice infected with NZ131, but not with SW510. By tandem mass spectrometry, we found that methionine was accumulated in blood of mice infected with NZ131, but not with SW510. We also found that AdoHcyase was a substrate for SPE B cysteine protease and SPE B could not cleave at specific sites. Based on our results, we indicated that SPE B induced host hyper-methioninemia through interaction with AdoHcyase. The SPE B has a novel pathogenic mechanism by group A streptococcal infection through suppressing AdoHcyase activity. We also found that SPE B was associated with keratin 14, a tissue-specific marker for keratinocytes. Protein level of keratin 14 was reduced in the keratinocyte HaCaT cells infected with S. pyogenes A20 strain (wild-type); whereas they were not reduced in the cells infected with SW507 strain (spe B mutant). By in vitro cleavage assay, we also found that keratin 14 appears to be a substrate of SPE B enzyme. Based on our results, we suggest that skin pathogenesis caused by S. pyogenes infection was mediated through keratin 14. Future work will focus on studies of functional relationships of SPE B with keratin 14 in mouse models. Thus, SPE B increases the ability of S. pyogenes to cause severe invasive disease and therefore is an important virulence factor for this organism.