Recombinant protein production and purification of SiiD, SiiE and SiiF - Components of the SPI4-encoded type I secretion system from Salmonella Typhimurium

Klingl S, Kordes S, Schmid B, Gerlach R, Hensel M, Muller Y (2020)

Publication Type: Journal article

Publication year: 2020

Journal

Protein Expression and Purification Elsevier

Book Volume: 172

Article Number: 105632

DOI: 10.1016/j.pep.2020.105632

Abstract

In humans, Salmonella enterica infections are responsible for a plethora of medical conditions. These include intestinal inflammation and typhoid fever. The initial contact between Salmonella and polarized epithelial cells is established by the SPI4-encoded type I secretion system (T1SS), which secretes SiiE, a giant non-fimbrial adhesin. We have recombinantly produced various domains of this T1SS from Salmonella enterica serovar Typhimurium in Escherichia coli for further experimental characterization. We purified three variants of SiiD, the periplasmic adapter protein spanning the space between the inner and outer membrane, two variants of the SiiE N-terminal region and the N-terminal domain of the SiiF ATP-binding cassette (ABC) transporter. In all three proteins, at least one variant yielded high amounts of pure soluble protein. Secondary structure content and cooperative unfolding were investigated by circular dichroism (CD) spectroscopy. Secondary structure contents were in good agreement with estimates derived from SiiD and SiiF homology models or, in case of the SiiE N-terminal region, a secondary structure prediction. For one SiiD variant, protein crystals could be obtained that diffracted X-rays to approximately 4 Å resolution.

Authors with CRIS profile

Stefan Klingl Lehrstuhl für Biotechnik (Proteinstruktur und -design) Benedikt Schmid Lehrstuhl für Biotechnik (Proteinstruktur und -design) Roman Gerlach Institute of Microbiology – Clinical Microbiology, Immunology and Hygiene Yves Muller Lehrstuhl für Biotechnik (Proteinstruktur und -design)

Involved external institutions

Universität Osnabrück DE Germany (DE)

How to cite

APA:

Klingl, S., Kordes, S., Schmid, B., Gerlach, R., Hensel, M., & Muller, Y. (2020). Recombinant protein production and purification of SiiD, SiiE and SiiF - Components of the SPI4-encoded type I secretion system from Salmonella Typhimurium. Protein Expression and Purification, 172. https://doi.org/10.1016/j.pep.2020.105632

MLA:

Klingl, Stefan, et al. "Recombinant protein production and purification of SiiD, SiiE and SiiF - Components of the SPI4-encoded type I secretion system from Salmonella Typhimurium." Protein Expression and Purification 172 (2020).

BibTeX: Download