2002 Fiscal Year Final Research Report Summary
Structure and function of the xenobiotic-multidrug efflux pump
| Project/Area Number |
12470066
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Bacteriology (including Mycology)
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| Research Institution | Tokai University |
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Principal Investigator |
NAKAE Taiji Tokai University School of Medicine, Professor, 医学部, 教授 (50102851)
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| Co-Investigator(Kenkyū-buntansha) |
NARITA Shin-ichiro Tokai University School of Medicine, Assistant Researcher, 医学部, 助手 (30338751)
YONEYAMA Hiroshi Tokai University School of Medicine, Assistant Professor, 大学院・農学研究科, 講師 (10220774)
YOSHIHARA Eisaku Tokai University School of Medicine, Associate Professor, 医学部, 助教授 (70167063)
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| Project Period (FY) |
2000 – 2002
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| Keywords | efflux / pump / transporter / antibiotic / resistance / drug / bacteria / membrane proteins |
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| Research Abstract |
Psudomonas aeruginosa exhibits resistant to structurally and functionally dissimilar antibiotics , which is mainly due to the expression of the multidrug efflux proteins. To solve the problems, we studied structure and function of the efflux pump and obtained the following results. 1. Membrane topology of MexA and OprM: (l) MexA is a lipoprotein anchoring the inner membrane and exposed most of the polypeptide moiety to the periplasmic aqueous phase : (2) OprM anchored the outer membrane via fatty acids exposing protein moiety to the periplasmic space. 2. Role of charged amino acid residues of MexB TMS has been analyzed and obtained the results that Asp407 and Asp408 in the TMS-4 and a Lys939 in TMS-10 form a salt bridge and a free Asp residue may be bound with proton. These amino acid residues are likely to form the proton pathway, which supply energy to the transporter. 3. To access the regulation of the mexR-mexA-MexB-oprM operon, physical interaction of MexR with MexOP-DNA was tested and found that MexR binds to the mexR proximal site of MexOP-DNA and regulates transcription of both mexAB-oprM and mexR. 4. Assembly process of MexEF-OprN transporter has been studied. The transporter appeared to be assembled by the following sequence. In the presence of functional intact MexF, MexE have partial proteolytic processing that is essential to assemble OprN into the complex. Presence of the impaired MexF or deletion of MexF caused proteolytic degradation of OprN and only a partial processing of MexE. 5. The substrate-recognition domain was analyzed by the domain-swapping experiments between MexB and MexY. The results showed that the transporters recognized the substrate at large periplasmic domains and directly transport the substrate from periplasmic space. A new active transport model has been proposed.
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