Abstract：
:The MexA-MexB-OprM efflux pump exports structurally and functionally diverse xenobiotics and confers multi-drug resistance on Pseudomonas aeruginosa cells. The MexB transporter traverses the inner membrane twelve times, bears two large periplasmic domains and has two homologous tandem repeats. To test whether two homologous halves of MexB function independently or interdependently, the protein was divided medially into two halves, each consisting of six amino- and carboxyl-proximal transmembrane segments. When two halves of MexB were coexpressed from independent open reading frames, the cells lacking chromosomal mexB exhibited restored antibiotic resistance at a level close to that in the cells producing a full-length MexB. In contrast, MexB protein containing either an amino- or carboxyl-half fragment failed to transport antibiotics. To test whether the amino- and carboxyl-proximal halves were present in a complex, we purified the histidine-tagged carboxyl-proximal half molecule using nickel-chelate chromatography from the cells that coexpressed two halves. The results showed that the nonhistidine-tagged amino-proximal half was co-purified with the carboxyl-proximal half, thereby indicating that the amino-proximal half fragment was tightly associated with the carboxyl-proximal half molecule. These findings suggest that the presence of both amino- and carboxyl-halves of MexB in a complex is essential for transport activity.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Eda S,Yoneyama H,Nakae Tdoi
10.1021/bi0300074subject
Has Abstractpub_date
2003-06-17 00:00:00pages
7238-44issue
23eissn
0006-2960issn
1520-4995journal_volume
42pub_type
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