The Bacterial Cell Surface: Structure, Biochemistry, Pathogenicity

Group Leader:  PD Dr. Dirk Linke
	from left to right: Iwan Grin, Thomas Arnold, Silvia Deiss, Dirk Linke, Dorothea Röhrich (picture taken in PhotoShop country, 2008)
	from left to right: Iwan Grin, Nagarajan Paramasivam, Thomas Arnold, Dorothea Röhrich, Marcus Thein  (picture taken in Gothenburg, FEMS Conference 2009)
	Back: Marcus Thein, Thomas Arnold, Iwan Grin Front: Dirk Linke, Nagarajan Paramasivam, Shakeel Shahid (picture taken on PhD day, Tübingen 2009)

Contact:

PD Dr. Dirk Linke
Max Planck Institute for Developmental Biology
Department I, Protein Evolution
Spemannstr. 35
D-72076 Tübingen
Germany
Tel   +49 (0)7071 - 601 357
Fax  +49 (0)7071 - 601 349
email    dirk.linke [at] tuebingen.mpg.de

Summary:

In Gram-negative bacteria, the cell surface is a complex mixture of carbohydrates, lipopolysaccharides, lipids and proteins. The bacterial cell surface serves two central purposes: protection from and interaction with the environment.

Protection

In pathogens, protection is achieved e.g. by capsule and biofilm formation, by binding and inactivation of components of the host immune system, or by variable expression of surface molecules to evade detection. We are interested in the regulation of surface molecule expression: at what time point are surface molecules made? What environmental signals trigger the production or shut it down? We are also interested in the process of biofilm formation. What determines the formation of biofilms? What surface molecules are involved and how are they synthesized?

Interaction

Bacterial infections can manifest themselves in very different forms, from mild skin irritation to diarrhea, from fever to the formation of tumors. Nevertheless, different infections by different microbes follow similar patterns: adhesion to host cells is followed by complex interactions that involve secretion of effector proteins (e.g. toxins) into host cells and induction of signaling processes by the host cells. We are interested in the onset of infection, which is mostly determined by the ability of the pathogen to adhere to host cells; we study protein and carbohydrate adhesins. How are these adhesins synthesized and exported to the cell surface? How is the synthesis regulated? What are the host cell binding partners of different adhesins?

Applications

We try to exploit our knowledge to create technical applications, typically in joint projects with other groups or industrial partners. Such applications include the production of artificial outer membrane proteins for sensory applications, and the evaluation of surface proteins for their use as vaccine targets, biomarkers, or as antigens for the production of therapeutic antibodies.

Transmission (A,C) and scanning (B,D) electron micrographs  of Bartonella henselae. The upper panel (A and B) shows a mutant strain deficient in the major Bartonella adhesin BadA. The lower panel (C and D) shows the wildtype bacteria. BadA has a length of 240 nm and is involved in host cell adhesion and – as can be seen in panel D – in biofilm formation.

Current Projects:

Current projects involve

• protein export across the outer membrane in Enterobacteria
• bacterial adhesion to host cells and abiotic surfaces
• outer membrane protein folding in Enterobacteria and in vitro
• development of artificial pore proteins for sensory applications
• carbohydrate synthesis in the periplasm and export across the outer membrane in Enterobacteria
• development of vaccines against Gram-negative bacteria
• a cooperation project on solid-state NMR of an outer membrane protein, with the FMP Berlin

Model Organisms:

We do most experiments in an S1 environment in Escherichia coli. Where necessary, we work in close cooperation with the Institutes for Medical Microbiology of the University of Tübingen and of the University of Frankfurt, where we can access S2 laboratories. This is relevant e.g. for work with Salmonella, Bartonella and Yersinia pathogens. 

Approach:

Our approach is a comparative one: instead of focusing on a single species, we compare proteins, protein modules, and mechanisms between related species. Evolutionary conservation is indicative of functionally relevant residues or domains (on the protein level). On the genome level, we are interested in the structure of operons and in the regulation of their components.

Methodology:

We use a broad range of methods, from bioinformatics, NMR and protein crystallography to classical microbiology, biochemistry, biophysics, and genetics. We also make use of in-house facilities for electron microscopy, mass spectrometry, fluorescence microscopy and DNA sequencing.

Cooperations:

We closely cooperate with groups from the University of Tübingen in the framework of the Collaborative Research Centre SFB766

Funding:

Our work is funded by

the Max Planck Society,

the Bill & Melinda Gates Foundation, Grand Challenges Explorations Program,

the Baden-Württemberg Stiftung, Nanotechnology Network and Methods for the Life Sciences Program

and by the German Science Foundation (DFG), SFB 766.

Jobs

Current job offers of the Max Planck Institute can be found here

We have no vacant PhD or PostDoc positions at the moment, but feel free to contact us for informal inquiries.

If you want to work with us

you should have a strong background in one or more of the following fields:

• Microbiology
• Biochemistry
• Structural Biology (x-ray crystallography or NMR)
• Biophysics

We regularly employ

• Summer students (Praktikum)
• Diploma / Master thesis students
• PhD students (PhD students should apply through the institute's central PhD Program)

Publication List:

(January 2012)

2012

Paulmann M, Arnold T, Linke D, Özdirekcan S, Kopp A, Gutsmann T, Kalbacher H, Wanke I, Schünemann V, Habeck M, Buerck J, Ulrich A, Schittek B (2012)
Structure-activity analysis of the dermcidin-derived peptide DCD-1L, an anionic antimicrobial peptide present in human sweat.
Journal of Biological Chemistry, in press

Leo J, Grin I, Linke D (2012)
Mechanisms of Autotransport in Gram-negative bacteria.
Philosophical Transactions of the Royal Society B, in press

Kaiser P, Linke D, Leo J, Schwarz H, Kempf V (2012)
Analysis of the BadA stalk from B. henselae reveals domain-specific and domain-overlapping functions in the host cell infection process.
Cellular Microbiology 14: 198-209.

2011

Paramasivam N, Linke D (2011)
ClubSub-P: Cluster-based subcellular localization prediction for Gram-negative bacteria and Archaea.
Frontiers in Microbiology 2: 218.

Gessmann D, Mager F, Naveed H, Arnold T, Weirich S, Linke D, Liang J, Nussberger S (2011)
Improving the resistance of a eukaryotic beta-barrel protein to thermal and chemical perturbation.
Journal of Molecular Biology 413: 150-161.

Wagner C, Polke M, Gerlach R, Linke D, Schwarz H, Hensel M (2011)
Functional and molecular dissection of SiiE, a giant non-fimbrial adhesin of Salmonella enterica.
Cellular Microbiology 13: 1286-1301.

Grin I, Linke D (2011)
GCView: the Genomic Context Viewer for protein homology searches.
Nucleic Acids Research 39: W353-W356.

Müller N, Kaiser P, Riess T, Schwarz H, Linke D, Schäfer A, Sinzger C, Kempf V (2011)
Trimeric autotransporter adhesin-dependent adherence of B. henselae, B. quintana and Y. enterocolitica to matrix components and endothelial cells under static and dynamic flow conditions.
Infection and Immunity 79: 2544-2553.

Linke D (2011)
Detergents in the purification, stabilization, and refolding of membrane proteins.
Chemistry Today 29(2), Focus on Oligos & Peptides: 32-35.

Björnsson H, Marteinsson V, Friðjónsson O, Linke D, Benediktsdóttir E (2011)
Isolation and characterisation of an antigen from the fish pathogen Moritella viscosa.
Journal of Applied Microbiology 111: 17-25.

Leo J, Lyskowski A, Hattula K, Hartmann M, Schwarz H, Butcher S, Linke D, Lupas A, Goldman A (2011)
The structure of E. coli IgG-binding protein D suggests a general model for bending and binding in trimeric autotransporter adhesins.
Structure 19: 1021-1030.

Vogelmann J, Ammelburg M, Finger C, Guezguez J, Linke D, Flötenmeyer M, Stierhof Y, Wohlleben W, Muth G (2011)
Conjugal plasmid transfer in Streptomyces resembles bacterial chromosome segregation by FtsK/SpoIIIE.
EMBO Journal 30: 2246-2254.

Müller J, Papic D, Ulrich T, Grin I, Schütz M, Oberhettinger P, Tommassen J, Linke D, Dimmer K, Autenrieth I, Rapaport D (2011)
Mitochondria can recognize and assemble fragments of a beta-barrel structure.
Molecular and Cellular Biology 22: 1638-1647.

Grin I, Schwarz H, Linke D (2011)
Electron microscopy techniques to study bacterial adhesion.
In: Bacterial Adhesion - Chemistry, Biology and Physics. Linke D and Goldman A, editors,
Advances in Experimental Medicine and Biology 715: 257-269.

O'Rourke F, Schmidgen T, Kaiser PO, Linke D, Kempf VA (2011)
Adhesins of Bartonella spp.
In: Bacterial Adhesion - Chemistry, Biology and Physics. Linke D and Goldman A, editors,
Advances in Experimental Medicine and Biology 715: 51-70.

Linke D (2011)
Choosing the right detergent.
Methods Navigator (Elsevier Online Resource)

Kaiser PO, Riess T, O'Rourke F, Linke D, Kempf VA (2011)
Bartonella spp.: throwing light on uncommon human infections.
International Journal of Medical Microbiology 301: 7-15.

Oberhettinger P, Schütz M, Raddatz G, Keller H, Autenrieth I, Linke D (2011)
The sequence of the pYV virulence plasmid from Yersinia enterocolitica strain WA-314 biogroup 1B serotype O:8.
Plasmid 65:20-24.

2010

Thein M, Sauer G, Paramasivam N, Grin I, Linke D (2010)
Efficient subfractionation of Gram-negative bacteria for proteomics studies.
Journal of Proteome Research 9: 6135-6147.

Arnold T, Linke D (2010)
Phase separation in the isolation and purification of membrane proteins.
Chemical Technology (South Africa), Dec 2010: 20-26, reprinted from BioTechniques 43: 427-434.

Lehr U, Schütz M, Oberhettinger P, Ruiz-Perez F, Donald JW, Palmer T, Linke D, Henderson IR, Autenrieth IB (2010)
C-terminal amino acid residues of the trimeric autotransporter adhesin YadA of Yersinia enterocolitica are decisive for its recognition and assembly by BamA.
Molecular Microbiology 78: 932-946.

Arnold T, Zeth K, Linke D (2010)
Omp85 from the thermophilic cyanobacterium Thermosynechococcus elongatus differs from proteobacterial Omp85 in structure and domain composition.
Journal of Biological Chemistry 285: 18003-18015.

Norum M, Tång E, Chavoshi T, Schwarz H, Linke D, Uv A, Moussian B (2010)
Trafficking through COPII stabilises cell polarity and drives secretion during Drosophila epidermal differentiation.
PLoS One 5(5): e10802.

Schütz M, Weiss E, Schindler M, Hallström T, Zipfel P, Linke D, Autenrieth I (2010)
Trimer stability of YadA is critical for virulence of Yersinia enterocolitica.
Infection and Immunity 78: 2677-2690.

Remmert M, Biegert A, Linke D, Lupas A, Söding J (2010)
Evolution of outer membrane beta-barrels from an ancestral beta-beta hairpin.
Molecular Biology and Evolution 27: 1348-1358. 

2009

Linke D (2009)
Commentary: never trust your word processor.
Biochemistry and Molecular Biology Education 37: 377.

Linke D (2009)
Detergents: an overview.
In: Guide to Protein Purification (2nd edition), Chapter 34, Burgess R, Deutscher M (editors),
Methods in Enzymology 463: 603-617.

Rohner N, Bercsény M, Orbán L, Kolanczyk M, Linke D, Brand M, Nüsslein-Vollhard C, Harris M (2009)
Duplication of fgfr1 permits FGF signalling to serve as a target for selection during domestication.
Current Biology 19: 1642-1647.

Remmert M, Linke D, Lupas A, Söding J (2009)
HHomp - Prediction and classification of outer membrane proteins.
Nucleic Acids Research 37: W446-W451.

Gröbner S, Linke D, Schütz W, Fladerer C, Madlung J, Autenrieth I, Witte W, Pfeifer Y (2009)
Emergence of carbapenem-non-susceptible extended-spectrum-beta-lactamase (ESBL)-producing Klebsiella pneumoniae isolates at the university hospital of Tübingen, Germany.
Journal of Medical Microbiology 58: 912-922.

Arnold T, Zeth K, Linke D (2009)
Structure and function of colicin S4, a colicin with a duplicated receptor binding domain.
Journal of Biological Chemistry 284: 6403-6413.

2008

Kaiser P, Riess T, Wagner C, Linke D, Lupas A, Schwarz H, Raddatz G, Schäfer A, Kempf V (2008)
The head of Bartonella adhesin A is crucial for host cell interaction of Bartonella henselae.
Cellular Microbiology 10: 2223-2234.

Szczesny P, Linke D, Ursinus A, Bär K, Schwarz H, Riess T, Kempf V, Lupas A, Martin J, Zeth K (2008)
Structure of the head of the Bartonella adhesin BadA.
PLoS Pathogens 4(8): e1000119.

Wagner C, Riess T, Linke D, Eberhardt C, Schäfer A, Reutter S, Maggi R, Kempf V (2008)
Use of Bartonella adhesin A (BadA) in serodiagnosis of Bartonella henselae infections.
International Journal of Medical Microbiology 298: 579-590.

Arnold T, Linke D (2008)
The use of detergents to purify membrane proteins.
In: Coligan J, Dunn B, Speicher D, Wingfield P (editors),
Current Protocols in Protein Science 53: 4.8.1.-4.8.30. or follow this link to the Current Protocols page with commenting functions

Hernandez Alvarez B, Hartmann M, Albrecht R, Lupas A, Zeth K, Linke D (2008)
A new expression system for protein crystallization using trimeric coiled-coil adaptors.
Protein Engineering Design and Selection 21: 11-18.

2007

Riess T, Raddatz G, Linke D, Schäfer A, Kempf V (2007)
Analysis of Bartonella adhesin A expression reveals differences between various B. henselae strains.
Infection and Immunity 75: 35-43.

Hsiao N, Söding J, Linke D, Lange C, Hertweck C, Wohlleben W, Takano E (2007)
ScbA from Streptomyces coelicolor A3(2) has homology to fatty acid synthases and is able to synthesize gamma-butyrolactones.
Microbiology-SGM 153: 1394-1404.

Grosskinsky U, Schütz M, Fritz M, Schmid Y, Lamparter M, Szczesny P, Lupas A, Autenrieth I, Linke D (2007)
A conserved glycine residue of trimeric autotransporter domains plays a key role in Yersinia adhesin A autotransport.
Journal of Bacteriology 189: 9011-9019.

Arnold T, Poynor M, Nussberger S, Lupas A, Linke D (2007)
Gene duplication of the eight-stranded beta-barrel OmpX produces a functional pore: A scenario for the evolution of transmembrane beta-barrels.
Journal of Molecular Biology 366: 1174-1184.

Arnold T, Linke D (2007)
Phase separation in the isolation and purification of membrane proteins.
BioTechniques 43: 427-434.

2006

Schulte B, Linke D, Klumpp S, Schaller M, Riess T, Autenrieth I, Kempf V (2006)
Bartonella quintana variably expressed outer membrane proteins mediate vascular endothelial growth factor secretion but not host cell adherence.
Infection and Immunity 74: 5003-5013.

Linke D, Riess T, Autenrieth I, Lupas A, Kempf V (2006)
Trimeric autotransporter adhesins: variable structure, common function.
Trends in Microbiology 14: 264-270.

Wollmann P, Zeth K, Lupas A, Linke D (2006)
Purification of the YadA membrane anchor for secondary structure analysis and crystallization.
International Journal of Biological Macromolecules 39: 3-9.

Albrecht R, Zeth K, Söding J, Lupas A, Linke D (2006)
Expression, crystallization and preliminary X-ray crystallographic studies of the outer membrane protein OmpW from Escherichia coli.
Acta Crystallographica Section F 62: 415-418.

2005 and earlier

Linke D, Frank J, Pope M, Soll J, Ilkavets I, Fromme P, Burstein E, Reshetnyak Y, Emelyanenko V (2004)
Folding kinetics and structure of OEP16.
Biophysical Journal 86: 1479-1487.

Linke D (2002)
Investigations on structure and function of a pore protein of the outer chloroplast membrane.
Thesis, Technical University Berlin (in german)

Linke D, Frank J, Holzwarth J, Soll J, Boettcher C, Fromme P (2000)
In vitro reconstitution and biophysical characterization of OEP16, an outer envelope pore protein of pea chloroplasts.
Biochemistry 39: 11050-11056.

Strauss T, Botha A, Kock J, Paul I, Smith D, Linke D, Schewe T, Nigam S (2000)
Mapping the distribution of 3-hydroxylipins in the Mucorales using immunofluorescence microscopy.
Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology 78: 39-42.

Smith D, Kock J, van Wyk P, Venter P, Coetzee D, van Heerden E, Linke D, Nigam S (2000)
The occurrence of 3-hydroxy oxylipins in the ascomycetous yeast family Lipomycetaceae.
South African Journal of Science 96: 247-249.

Kock J, Venter P, Linke D, Schewe T, Nigam S (1998)
Biological dynamics and distribution of 3-hydroxy fatty acids in the yeast Dipodascopsis uninucleata as investigated by immunofluorescence microscopy. Evidence for a putative regulatory role in the sexual reproductive cycle.
FEBS Letters 427: 345-348.

Essigmann B, Güler S, Narang R, Linke D, Benning C (1998)
Phosphate availability affects the thylakoid lipid composition and the expression of SQD1, a gene required for sulfolipid biosynthesis in Arabidopsis thaliana.
Proceedings of the National Academy of Sciences U.S.A. 95: 1950-1955.