化膿性鏈球菌為人類常見的致病菌，臨床上引起的疾病從輕微的咽喉炎、皮膚感染到嚴重的壞死性肌膜炎、鏈球菌毒性休克症候群等。寡胜肽膜透酶由五個蛋白組成，附著於細胞膜上，屬於 ABC-transporter 群。其中 OppA 為細胞外膜上可與受質結合的脂蛋白；OppB 及 OppC 為形成通道的穿膜蛋白；OppD 及 OppF 為內膜上提供能量的 ATPase。先前其他細菌的研究發現，OPP可能具有下列幾項功能：細菌養份的來源、傳送外部的訊息分子、影響細菌的附著能力及 OppA 具有ecto-ATPase 的功能。然而，化膿性鏈球菌 OPP 的生物功能目前並未十分清楚，本篇論文的研究在於了解 OppA 在化膿性鏈球菌的功能。首先進行 PCR分析篩選 41 株菌共 19 種 emm types，我們發現 opp 基因廣泛的存在於每一種 emm types。當我們利用從 TSBY broth 所純化出的 peptides mixture 與重組蛋白 r-OppA 作用後，經由 native cationic gel 電泳分析，發現可在膠體上造成位移改變。利用 KKKKK 及 VYIHPE 二種人工合成的寡胜肽，分別在 4 mM 及 2.5 mM 的濃度時可使得 r-OppA 在膠體上的 band 消失不見。以及將此二種人工合成的寡胜肽標定上螢光，比較 A20（wild-type）及 SW569（oppA mutant）二者運送寡胜肽的能力，發現 SW569 螢光強度明顯比 A20 來得低，因此推測 OppA 可能具有與寡胜肽結合的能力。為了了解寡胜肽與 OppA 結合的區域，我們也建構並純化 6 種不同片段的 r-OppA，但是發現這些片段均無法與寡胜肽結合，表示 OppA 可能必需在具備完整的結構下才會與寡胜肽結合。此外，我們也發現 r-OppA 並不具有 ecto-ATPase 的功能。利用 Western Blot、ligand overlay assay、CELISA、掃瞄式電子顯微鏡及 competitive adhesion assay 五種不同的實驗方式，發現 r-OppA 無法緊密的與細胞結合。但根據 invasion assay 以及antibody inhibited invasion assay 的結果發現 OppA 似乎與促進化膿性鏈球菌侵入宿主細胞有關。

Streptococcus pyogenes (group A Streptococcus, GAS) is known to be the causation of diseases in broad range symptoms that include streptococcal toxic shock syndrome, sepsis, and necrotising fasciitis. Oligopeptide permease (OPP) is a membrane-associated complex of five proteins (OppABCDF) belonging to the ABC-transporter family. OppA is an outer membrane-associated lipoprotein. OppB and OppC are transmembrane proteins. OppD and OppF are predicted as inner membrane-associated ATPases. The oligopeptide transport system has been described that involved in nutrient acquisition, sensing of extracellular signaling molecules, influence of adherence, and ecto-ATPase of OppA. However, the biological functions of OPP in S. pyogenes are unclear. The aim of this study was to demonstrate the functions of OppA in S. pyogenes. The opp genes were analyzed by PCR and presented in every strain so far we tested it (including 19 emm types of 41 strains). The full length of recombinant OppA (r-OppA) protein incubated with peptides mixture purified from TSBY broth can cause mobility shift on the native cationic gel. The high concentration of 4 mM KKKKK peptide and 2.5 mM VYIHPE peptide could cause the band of r-OppA disappeared on the native cationic gel. The FITC-labeled peptides of KKKKK and VYIHPE incubated with A20 (wild type) and SW569 (oppA mutant). The fluorescence intensity could be decreased in SW569. These results suggested that OppA could bind to oligopeptide. In order to find the peptide-binding domain of OppA, the six different truncated r-OppA proteins were constructed and purified. We found that the truncated r-OppA proteins could not bind to oligopeptide. This result suggested that the intact structure of OppA might be required for the peptide binding ability. In addition, we found that the r-OppA of S. pyogenes had no ATPase activity. The adhesion ability of OppA was determined by the five independent assays including Western Blot, ligand overlay assay, CELISA, scanning electron microscopy and competitive adhesion assay. None of assay could demonstrate that r-OppA can bind to host cells. However, OppA could enhance the invasion activity of S. pyogenes in the invasion assay and antibody inhibited invasion assay.

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