Journal of Biomolecular NMR

, Volume 61, Issue 3–4, pp 333–345 | Cite as

Characterization of the insertase BamA in three different membrane mimetics by solution NMR spectroscopy

  • Leonor Morgado
  • Kornelius Zeth
  • Björn M. Burmann
  • Timm Maier
  • Sebastian Hiller


The insertase BamA is the central protein of the Bam complex responsible for outer membrane protein biogenesis in Gram-negative bacteria. BamA features a 16-stranded transmembrane β-barrel and five periplasmic POTRA domains, with a total molecular weight of 88 kDa. Whereas the structure of BamA has recently been determined by X-ray crystallography, its functional mechanism is not well understood. This mechanism comprises the insertion of substrates from a dynamic, chaperone-bound state into the bacterial outer membrane, and NMR spectroscopy is thus a method of choice for its elucidation. Here, we report solution NMR studies of different BamA constructs in three different membrane mimetic systems: LDAO micelles, DMPC:DiC7PC bicelles and MSP1D1:DMPC nanodiscs. The impact of biochemical parameters on the spectral quality was investigated, including the total protein concentration and the detergent:protein ratio. The barrel of BamA is folded in micelles, bicelles and nanodiscs, but the N-terminal POTRA5 domain is flexibly unfolded in the absence of POTRA4. Measurements of backbone dynamics show that the variable insertion region of BamA, located in the extracellular lid loop L6, features high local flexibility. Our work establishes biochemical preparation schemes for BamA, which will serve as a platform for structural and functional studies of BamA and its role within the Bam complex by solution NMR spectroscopy.


Membrane protein Omp85 insertase NMR spectroscopy Backbone dynamics Bicelles Nanodiscs 









Polypeptide transport-associated


β-barrel assembly machinery


Outer membrane protein



We thank Sina Reckel and Thomas Raschle for technical advice and discussions. This work was supported by grants from the Swiss National Science Foundation (Grant PP00P3_128419) and the European Research Council (FP7 contract MOMP 281764) to S.H. The sequence-specific resonance assignments of BamA in lipid bilayer nanodiscs (residues D678–Q698) were deposited in the BMRB (accession code 25359).

Supplementary material

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Supplementary material 1 (PDF 1818 kb)


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Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Leonor Morgado
    • 1
  • Kornelius Zeth
    • 1
    • 2
    • 3
  • Björn M. Burmann
    • 1
  • Timm Maier
    • 1
  • Sebastian Hiller
    • 1
  1. 1.BiozentrumUniversity of BaselBaselSwitzerland
  2. 2.Department of BiochemistryUniversity of the Basque CountryLeioaSpain
  3. 3.IKERBASQUE Basque Foundation for ResearchBilbaoSpain

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