Monitoring of stress responses

2 downloads 0 Views 1MB Size Report
Oct 10, 2006 - ... physiology

The organisers would like to thank Novozymes Delta Ltd who generously supported the meeting.

Microbial Cell Factories

BioMed Central

Open Access

Oral Presentation

Monitoring of stress responses Thomas Schweder*1, Britta Jürgen1, Birgit Voigt2, Daniel Pioch1, Michael Hecker2, Stefan Evers3 and Karl-Heinz Maurer3 Address: 1Pharmaceutical Biotechnology, Institute of Pharmacy, Ernst-Moritz-Arndt-University, D-17487 Greifswald, Germany, 2Institute of Microbiology, Ernst-Moritz-Arndt-University, D-17487 Greifswald, Germany and 3VTB Enzyme Technology, Henkel KGaA, D-40191 Düsseldorf, Germany * Corresponding author

from The 4th Recombinant Protein Production Meeting: a comparative view on host physiology Barcelona, Spain. 21–23 September 2006 Published: 10 October 2006 Microbial Cell Factories 2006, 5(Suppl 1):S23

doi:10.1186/1475-2859-5-S1-S23

The 4th Recombinant Protein Production Meeting: a comparative view on host physiology

The organisers would like to thank Novozymes Delta Ltd who generously supported the meeting. Meeting abstracts – A single PDF containing all abstracts in this supplement is available here http://www.biomedcentral.com/content/pdf/1475-2859-5-S1-info.pdf

© 2006 Schweder et al; licensee BioMed Central Ltd.

Background The consideration of bacterial stress and starvation responses is of great importance for the successful establishment of an industrial large scale fermentation process. Suitable analysis techniques for stress and starvation specific genes are therefore particularly interesting for the monitoring and control of such processes. The combined methods of transcriptome analysis, high resolution twodimensional polyacrylamide gel electrophoresis (2DPAGE) and mass spectrometry have been extensively applied for the description of general and specific stress and starvation responses of industrial microorganisms.

ess-relevant marker genes would be valuable diagnostic tools for the monitoring of cellular physiology. In this g

Results By means of proteomics and transcriptome analyses we identified marker genes of the gram-positive bacteria Bacillus subtilis and Bacillus licheniformis. The expression of such marker genes is specifically regulated by distinct stress and starvation conditions. For both bacteria, which represent important industrial hosts with a long history in industrial enzyme production, we have filtered a set of marker genes, which could be used as indicators for process-relevant stress situations during protein production fermentation processes. For example, in Figure 1 starvation specific marker proteins for nitrogen, phosphate and glucose limitation of B. licheniformis are summarised. Such process-critical genes/proteins can be used as biomarkers in order to control the fitness and productivity of these industrial bacterial hosts during fermentation processes. DNA- and protein-chips specific for such proc-

tions for Colour formis Figure coding 1 phosphate, of extracellular glucose and marker nitrogen proteins starvation of B. lichenicondiColour coding of extracellular marker proteins of B. licheniformis for phosphate, glucose and nitrogen starvation conditions. Colour coding was done with the Delta 2D software http://www.decodon.com. (P: phosphate starvation, C: glucose starvation, N: nitrogen starvation). [1]

Page 1 of 3 (page number not for citation purposes)

Microbial Cell Factories 2006, 5(Suppl 1):S23

A

B

A electric Figure Schematic 2 DNA-chip presentation during aofglucose-limited the electric chip fermentation principle and process (B) analysis of a B. subtilis glucose-starvation marker gene with an A Schematic presentation of the electric chip principle and B analysis of a B. subtilis glucose-starvation marker gene with an electric DNA-chip during a glucose-limited fermentation process. EBC = electric DNA-chip, RT-PCR = real time RT-PCR

respect fast mRNA and protein analytical techniques for an at-line monitoring of gene expression during bioprocesses are required. The electric chip technique fulfills these requirements [2]. This technique allows a fast and reproducible expression analysis of process-relevant marker genes (see Figure 2).

Conclusion It is demonstrated that electric chips loaded with mRNA specific DNA-probes or with marker protein specific antibodies represent a suitable alternative for gene expression analyses in competition with the real time RT-PCR during fermentation processes. The electric chip technique is easy

Page 2 of 3 (page number not for citation purposes)

Microbial Cell Factories 2006, 5(Suppl 1):S23

to automate and could be cheaper in the handling than the established gene expression analysis techniques. The electric biochip combined with an automated sample preparation establishes a basis for continuous at-line monitoring of host cell physiology during industrial bioprocesses.

Acknowledgements This study was financially supported by the Ministry of Education, Science and Culture of Mecklenburg-Vorpommern (grant number: 0202120) and the Federal Ministry of Education, and Science (BMBF) (grant number: 031U213A).

References 1.

2.

Voigt B, Schweder T, Sibbald MJ, Albrecht D, Ehrenreich A, Bernhardt J, Feesche J, Maurer KH, Gottschalk G, Van Dijl JM, Hecker M: The extracellular proteome of Bacillus licheniformis grown in different media and under different nutrient starvation conditions. Proteomics 2006, 6:268-281. Jürgen B, Barken KB, Tobisch S, Pioch D, Wümpelmann M, Hecker M, Schweder T: Application of an Electrical DNA-Chip for the Expression Analysis of Bioprocess-Relevant Genes of Bacillus subtilis. Biotechnol Bioeng 2005, 92:299-307.

Publish with Bio Med Central and every scientist can read your work free of charge "BioMed Central will be the most significant development for disseminating the results of biomedical researc h in our lifetime." Sir Paul Nurse, Cancer Research UK

Your research papers will be: available free of charge to the entire biomedical community peer reviewed and published immediately upon acceptance cited in PubMed and archived on PubMed Central yours — you keep the copyright

BioMedcentral

Submit your manuscript here: http://www.biomedcentral.com/info/publishing_adv.asp

Page 3 of 3 (page number not for citation purposes)