Crystal structure of the membrane fusion protein CusB from Escherichia coli.
Gram-negative bacteria, such as Escherichia coli, frequently utilize tripartite efflux complexes belonging to the resistance-nodulation-division family to expel diverse toxic compounds from the cell. These systems contain a periplasmic membrane fusion protein (MFP) that is critical for substrate transport. We here present the x-ray structures of the CusB MFP from the copper/silver efflux system of E. coli. This is the first structure of any MFPs associated with heavy-metal efflux transporters. CusB bridges the inner-membrane efflux pump CusA and outer-membrane channel CusC to mediate resistance to Cu(+) and Ag(+) ions. Two distinct structures of the elongated molecules of CusB were found in the asymmetric unit of a single crystal, which suggests the flexible nature of this protein. Each protomer of CusB can be divided into four different domains, whereby the first three domains are mostly beta-strands and the last domain adopts an entirely helical architecture. Unlike other known structures of MFPs, the alpha-helical domain of CusB is folded into a three-helix bundle. This three-helix bundle presumably interacts with the periplasmic domain of CusC. The N- and C-termini of CusB form the first beta-strand domain, which is found to interact with the periplasmic domain of the CusA efflux pump. Atomic details of how this efflux protein binds Cu(+) and Ag(+) were revealed by the crystals of the CusB-Cu(I) and CusB-Ag(I) complexes. The structures indicate that CusB consists of multiple binding sites for these metal ions. These findings reveal novel structural features of an MFP in the resistance-nodulation-division efflux system and provide direct evidence that this protein specifically interacts with transported substrates.
Su
Chih-Chia
CC
Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, 50011, USA.
Yang
Feng
F
Long
Feng
F
Reyon
Deepak
D
Routh
Mathew D
MD
Kuo
Dennis W
DW
Mokhtari
Adam K
AK
Van Ornam
Jonathan D
JD
Rabe
Katherine L
KL
Hoy
Julie A
JA
Lee
Young Jin
YJ
Rajashankar
Kanagalaghatta R
KR
Yu
Edward W
EW
D001665:Binding Sites; D003300:Copper / Q000737:chemistry / Q000378:metabolism; D018360:Crystallography, X-Ray / Q000379:methods*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013058:Mass Spectrometry; D050576:Membrane Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012834:Silver / Q000737:chemistry / Q000378:metabolism
D029968:Escherichia coli Proteins; D050576:Membrane Fusion Proteins; D012834:Silver; D003300:Copper
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