Expression and Association of the Yersinia pestis Translocon Proteins, YopB and YopD, Are Facilitated by Nanolipoprotein Particles

Coleman, Matthew A.; Cappuccio, Jenny A.; Blanchette, Craig D.; Gao, Tingjuan; Arroyo, Erin S.; Hinz, Angela K.; Bourguet, Feliza A.; Segelke, Brent; Hoeprich, Paul D.; Huser, Thomas; Laurence, Ted A.; Motin, Vladimir L.; Chromy, Brett A.

doi:10.1371/journal.pone.0150166

Title: Expression and Association of the Yersinia pestis Translocon Proteins, YopB and YopD, Are Facilitated by Nanolipoprotein Particles

Journal Article · 25 March 2016 · PLoS ONE

DOI: https://doi.org/10.1371/journal.pone.0150166 · OSTI ID:1259528

Coleman, Matthew A. ^[1]; Cappuccio, Jenny A. ^[2]; Blanchette, Craig D. ^[3]; Gao, Tingjuan ^[4]; Arroyo, Erin S. ^[3]; Hinz, Angela K. ^[3]; Bourguet, Feliza A. ^[3]; Segelke, Brent ^[3]; Hoeprich, Paul D. ^[3]; Huser, Thomas ^[4]; Laurence, Ted A. ^[3]; Motin, Vladimir L. ^[5]; Chromy, Brett A. ^[4]

Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Univ. of California, Davis, CA (United States)
Humboldt State Univ., Arcata, CA (United States)
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Univ. of California, Davis, CA (United States)
Univ. of Texas Medical Branch, Galveston, TX (United States)

Yersinia pestis enters host cells and evades host defenses, in part, through interactions between Yersinia pestis proteins and host membranes. One such interaction is through the type III secretion system, which uses a highly conserved and ordered complex for Yersinia pestis outer membrane effector protein translocation called the injectisome. The portion of the injectisome that interacts directly with host cell membranes is referred to as the translocon. The translocon is believed to form a pore allowing effector molecules to enter host cells. To facilitate mechanistic studies of the translocon, we have developed a cell-free approach for expressing translocon pore proteins as a complex supported in a bilayer membrane mimetic nano-scaffold known as a nanolipoprotein particle (NLP) Initial results show cell-free expression of Yersinia pestis outer membrane proteins YopB and YopD was enhanced in the presence of liposomes. However, these complexes tended to aggregate and precipitate. With the addition of co-expressed (NLP) forming components, the YopB and/or YopD complex was rendered soluble, increasing the yield of protein for biophysical studies. Biophysical methods such as Atomic Force Microscopy and Fluorescence Correlation Spectroscopy were used to confirm that the soluble YopB/D complex was associated with NLPs. An interaction between the YopB/D complex and NLP was validated by immunoprecipitation. The YopB/D translocon complex embedded in a NLP provides a platform for protein interaction studies between pathogen and host proteins. Ultimately, these studies will help elucidate the poorly understood mechanism which enables this pathogen to inject effector proteins into host cells, thus evading host defenses.

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Research Organization:: Lawrence Livermore National Laboratory (LLNL), Livermore, CA (United States)

Sponsoring Organization:: USDOE; National Science Foundation (NSF)

Grant/Contract Number:: AC52-07NA27344

OSTI ID:: 1259528

Journal Information:: PLoS ONE, Journal Name: PLoS ONE Journal Issue: 3 Vol. 11; ISSN 1932-6203

Publisher:: Public Library of ScienceCopyright Statement

Country of Publication:: United States

Language:: English

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Type III secretion gets an LcrV tip Mota, Luís Jaime Trends in Microbiology, Vol. 14, Issue 5 https://doi.org/10.1016/j.tim.2006.02.010	journal	May 2006
Different Apolipoproteins Impact Nanolipoprotein Particle Formation Chromy, Brett A.; Arroyo, Erin; Blanchette, Craig D. Journal of the American Chemical Society, Vol. 129, Issue 46, p. 14348-14354 https://doi.org/10.1021/ja074753y	journal	November 2007
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LcrV is a channel size-determining component of the Yop effector translocon of Yersinia Holmstrom, Anna; Olsson, Jan; Cherepanov, Peter Molecular Microbiology, Vol. 39, Issue 3 https://doi.org/10.1046/j.1365-2958.2001.02259.x	journal	February 2001
Type III export: new uses for an old pathway Plano, Gregory V.; Day, James B.; Ferracci, Franco Molecular Microbiology, Vol. 40, Issue 2 https://doi.org/10.1046/j.1365-2958.2001.02354.x	journal	April 2001
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Supramolecular Structure of the Salmonella typhimurium Type III Protein Secretion System Kubori, T. Science, Vol. 280, Issue 5363 https://doi.org/10.1126/science.280.5363.602	journal	April 1998
Vaccination of Mice with a Yop Translocon Complex Elicits Antibodies That Are Protective against Infection with F1- Yersinia pestis Ivanov, M. I.; Noel, B. L.; Rampersaud, R. Infection and Immunity, Vol. 76, Issue 11 https://doi.org/10.1128/IAI.00189-08	journal	September 2008
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Interleukin-10 and Inhibition of Innate Immunity to Yersiniae: Roles of Yops and LcrV (V Antigen) Brubaker, R. R. Infection and Immunity, Vol. 71, Issue 7 https://doi.org/10.1128/IAI.71.7.3673-3681.2003	journal	July 2003
Role of Predicted Transmembrane Domains for Type III Translocation, Pore Formation, and Signaling by the Yersinia pseudotuberculosis YopB Protein Ryndak, M. B.; Chung, H.; London, E. Infection and Immunity, Vol. 73, Issue 4 https://doi.org/10.1128/IAI.73.4.2433-2443.2005	journal	March 2005
The YopD Translocator of Yersinia pseudotuberculosis Is a Multifunctional Protein Comprised of Discrete Domains Olsson, J.; Edqvist, P. J.; Broms, J. E. Journal of Bacteriology, Vol. 186, Issue 13 https://doi.org/10.1128/JB.186.13.4110-4123.2004	journal	June 2004

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Cell-Free Co-Translational Approaches for Producing Mammalian Receptors: Expanding the Cell-Free Expression Toolbox Using Nanolipoproteins Shelby, Megan L.; He, Wei; Dang, Amanda T. Frontiers in Pharmacology, Vol. 10 https://doi.org/10.3389/fphar.2019.00744	journal	July 2019
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