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. 2011 Jun 10;409(3):348-57.
doi: 10.1016/j.jmb.2011.03.035. Epub 2011 Apr 2.

Crystal structure of BamD: an essential component of the β-Barrel assembly machinery of gram-negative bacteria

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Crystal structure of BamD: an essential component of the β-Barrel assembly machinery of gram-negative bacteria

Cristina M Sandoval et al. J Mol Biol. .

Abstract

Folding and insertion of integral β-barrel proteins in the outer membrane (OM) is an essential process for Gram-negative bacteria that requires the β-barrel assembly machinery (BAM). Efficient OM protein (OMP) folding and insertion appears to require a consensus C-terminal signal in OMPs characterized by terminal F or W residues. The BAM complex is embedded in the OM and, in Escherichia coli, consists of the β-barrel BamA and four lipoproteins BamBCDE. BamA and BamD are broadly distributed across all species of Gram-negative bacteria, whereas the other components are present in only a subset of species. BamA and BamD are also essential for viability, suggesting that these two proteins constitute the functional core of the bacterial BAM complex. Here, we present the crystal structure of BamD from the thermophilic bacteria Rhodothermus marinus refined to 2.15 Å resolution. The protein contains five tetratricopeptide repeats (TPRs) organized into two offset tandems, each capped by a terminal helix. The N-terminal domain contains three TPRs and displays remarkable structural similarity with proteins that recognize targeting signals in extended conformations. The C-terminal domain harbors the remaining two TPRs and previously described mutations that impair binding to other BAM components map to this domain. Therefore, the structure suggests a model where the C-terminal domain provides a scaffold for interaction with BAM components, while the N-terminal domain participates in interaction with the substrates, either recognizing the C-terminal consensus sequence or binding unfolded OMP intermediates.

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Figures

Figure 1
Figure 1. Crystal structure of rmBamD
Two views of rmBamD related by a 90º rotation. The N-terminal domain contains TPRs 1–3 (red, green and blue respectively) and is capped by a helix (light blue). The C-terminal domain contains TPRs 4 and 5 (yellow and orange) and the final capping helix (light orange). An extended loop in TPR3 is labeled magenta.
Figure 2
Figure 2. Electrostatic surface potential of rmBamD
The distribution of positive (blue) and negative (red) surface potential is mapped to a surface representation of rmBamD. For reference, the orientation of the molecule in the left panels is the same as in Figure 1. A negatively charged, semicircular groove between the N and C-terminal domains 24Å in diameter is labeled.
Figure 3
Figure 3. Sequence conservation in BamD
A sequence alignment of BamD homologs (see text) was analyzed with the Consurf sever. Conservation scores are normalized so that the average score is zero and the standard deviation is 1. Thus rapidly evolving positions are negative (colored cyan) and slowly evolving positions are positive (colored purple) while positions with an average rate of evolution are colored white. (A) Aligned sequences of BamD from Rhodothermus marinus (Rhodothermus), Escherichia coli (E. coli) and Neisseria meningitidis (Neisseria) colored by the Consurf scores. Structural elements are labeled and highlighted in boxes colored as in Figure 1. (B) Conservation scores mapped on a surface representation of the rmBamD structure showing a groove between the extended loop in TPR3 and the C-terminal domain lined with slowly evolving residues.
Figure 4
Figure 4. The N-terminal domain of rmBamD is similar to proteins binding extended polypeptides
The N-terminal domain of rmBamD (colored as in Figure 1) is superimposed to TPR domains of Hop (A, yellow), FKBP52 (B, salmon), Tom70(C, gray) and PEX5 (D, light orange); as well as the chaperones PcrH (E, light green) and IpgC (light blue). All these proteins bind extended polypeptides (shown as black C-alpha traces) in the concave face of the TPR tandem.
Figure 5
Figure 5. Working model of the BAM complex
BamA and BamD constitute the core of the complex. BamA is proposed to recognize and help fold nascent OMPs with its five POTRA repeats (show as orange ovals). The N-terminal domain of BamD may assist in this folding role, or may recognize the consensus sequence at the C-terminus of OMPs. BamC and E interact with the C-terminal domain of BamD and help stabilize the complex. BamB interacts with BamA independently of BamC, D and E.

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