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Analyzing the Role of Periplasmic Folding Factors in the Biogenesis of OMPs and Members of the Type V Secretion System

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The BAM Complex

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1329))

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Abstract

The outer membrane (OM) of gram-negative bacteria is highly packed with OM proteins (OMPs) and the trafficking and assembly of OMPs in gram-negative bacteria is a subject of intense research. Structurally, OMPs vary in the number of β-strands and in the size and complexity of extra-membrane domains, with extreme examples being the members of the type V protein secretion system (T5SS), such as the autotransporter (AT) and intimin/invasin families of secreted proteins, in which a large extracellular “passenger” domain is linked to a β-barrel that inserts in the OM. Despite their structural and functional diversity, OMPs interact in the periplasm with a relatively small set of protein chaperones that facilitate their transport from the inner membrane (IM) to the β-barrel assembly machinery (BAM complex), preventing aggregation and assisting their folding in various aspects including disulfide bond formation. This chapter is focused on the periplasmic folding factors involved in the biogenesis of integral OMPs and members of T5SS in E. coli, which are used as a model system in this field. Background information on these periplasmic folding factors is provided along with genetic methods to generate conditional mutants that deplete these factors from E. coli and biochemical methods to analyze the folding, surface display, disulfide formation and oligomerization state of OMPs/T5SS in these mutants.

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Acknowledgements

G.B. and E.M. contribute equally to this work. Work in the laboratory of LAF is supported by research grants from the Spanish “Ministerio de Economía y Competitividad” (MINECO) (BIO2011-26689), “Comunidad Autónoma de Madrid” (S2010-BMD-2312), “La Caixa” Foundation, and the European Research Council (ERC-2012-ADG_20120314).

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    Authors and Affiliations

    1. Department of Microbial Biotechnology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, 28049, Madrid, Spain

      Gustavo Bodelón, Elvira Marín & Luis Ángel Fernández

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    1. Gustavo Bodelón
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    2. Elvira Marín
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    3. Luis Ángel Fernández
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    Correspondence to Luis Ángel Fernández .

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    1. National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Laboratory of Molecular Biology, , Bethesda, Maryland, USA

      Susan Buchanan

    2. Department of Biological Sciences,, Markey Center for Structural Biology, Purdue University, West Lafayette, Indiana, USA

      Nicholas Noinaj

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    Bodelón, G., Marín, E., Fernández, L.Á. (2015). Analyzing the Role of Periplasmic Folding Factors in the Biogenesis of OMPs and Members of the Type V Secretion System. In: Buchanan, S., Noinaj, N. (eds) The BAM Complex. Methods in Molecular Biology, vol 1329. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2871-2_7

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    • DOI: https://doi.org/10.1007/978-1-4939-2871-2_7

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