Download Lutz Heide, Pharmaceutical Institute, Tübingen University

L1: Julian Davies – Small is beautiful
 Antibiotic history
 Modes of action
 Production and resistance
 The age of streptomycetes
 The parvome: chemical ecology
 Cooperation vs. competition
 Thoughts on evolution

L2: Flavia Marinelli - Isolation and screening of secondary metabolite-producing
microorganisms
How many microbial secondary metabolites (potential drugs) are known?
Who are the microbial producers (ABL database)?
Overview of products on the market from ascomycota, actinomycetes, myxobacteria
and cyanobacteria
Novel antiinfectives, antitumor, statins and immunosuppressive agents
Isolation methods focusing on our experience in rare actinomycetes: challenges of
rare actinomycetes
Comparison of different methods of screening
A screening case history: lantibiotics
L3: Lutz Heide - Secondary metabolites: structure, biosynthesis, enzymology and
genetics
1) Most frequently used classes of antibiotics
2) How antibiotics kill bacteria: Antibiotic targets
3) Chemical classes of secondary metabolites:
 β-Lactam antibiotics (Penicillins, Aminopenicillins, Cephalosporins)
 Peptide antibiotics
 Glycopeptide antibiotics
 Lipopeptide antibiotics
 Aminocoumarins
 Tetracyclines
 Macrolides
 Polyene antibiotics
 Aminoglycosides
 Lincosamides
 Synthetic antibacterials: Fluorquinolones, Oxazolidinones, Sulfonamides,
Nitroimidazoles
4) Principles of biosynthesis and enzymology: polyketide synthases, non-ribosomal
peptide synthases and beyond
L4: Greg Challis and Govind Chandra: - Introduction to the computer
workshops
L5: Mervyn Bibb - Regulation of secondary metabolism and its analysis
1. Overview of regulation of secondary metabolism – pathway-specific and
pleiotropic regulatory genes
2. Specific examples – A-factor (refer to Kenji), tylosin, Ser-Thr protein kinases
(refer to Duska), DasR, ppGpp etc
3. Genomic approaches to regulation of secondary metabolism
a. Transcriptomics & defining regulons (arrays, RNA-seq, ChIP-on-Chip,
ChIP-seq)
b. Proteomics (2D gels and MALDI-ToF; mass spec-based proteomics)
4. Targeted approaches (Northern’s, SI nuclease protection analysis, primer extension,
RT –PCR & qRT-PCR)
5. Identifying genes involved in secondary metabolism (& making defined mutants)
L6: Roberto Kolter - Microbial chemical ecology: Secondary metabolites as
signals and killing agents
- quorum sensing: history and examples
- rethinking quorum sensing - the case of Bacillus subtilis and paracrine signaling
- secondary metabolites in symbioses
vibrio squid
attine ants
L7: Kenji Ueda - Microbial chemical ecology: Bacterial hormone-sensing
rediscovered in applied microbiology
- brief introduction to the unique development of applied microbiology in Japan
- overviewing the A-factor story
- gamma-butyrolactones in actinomycete communities
L8: Duška Vujaklija - Old concepts/new insights in bacterial phosphorylation
Introduction to phosphorylation: Reversible phosphorylation of proteins occurs in
all organisms and possesses crucial regulatory roles in a broad spectrum of biological
processes.
Retrospective: It was discovered in the mid 1950s and for many years it was tought
to exist only in eukaryotes.
Period of controversy: Serine, threonine and tyrosine phosphorylation is the most
common type of phosphorylation in eukaryotes, on contrary, in bacteria
phosphorylation occurs predominantly on histidine and aspartate (two-component
system). Until the early 1990s it was largely considered that these two
phosphorylation systems are mutually exclusive.
Two-component systems: Crucial bacterial regulatory mechanism for sensing and
responding to internal and extranal signals. It also regulates different functions related
to bacterial pathogenicity: including toxin production, cell adhesion, quorum sensing,
capsule synthesis, motility, and drug resistance.
More recent data: Genome sequencing confirmed the widespread presence of genes
encoding eukaryotic like Ser/Thr kinases and phosphatases.
Tyrosine phosphorylation in bacteria: The first studies only suggested tyrosine
kinase activities in bacteria, but the first conclusive evidence of bacterial tyrosine
phosphorylation came only a decade ago.
Bacterial tyrosine kinases exibit unexpected features and have been identified in a
variety of bacteria.
The list of substrates of BY-kinases is increasing and will be discussed with emphasis
on tyrosine phosphorylation of bacterial single stranded DNA binding proteins,
particularly SSB proteins from Streptomyces sp.
Powerful new methods: the number of serine-threonine- and tyrosinephosphorylated proteins have been discovered recently by mass spectrometry-based
gel-free phosphoproteomics.
L9: Flavia Marinelli - Industrial fermentation and strain improvement of
secondary metabolite-producing microorganisms
Strain maintenance
Fermentation process
Optimization of medium components and physico-chemical parameters
Examples from our recent works: modulation of teicoplanin complex, influence of
process parameters on sirolimus production
Strain improvement
Mutagenesis (recent examples on sirolimus and cyclosporin)
Ribosome engineering (Ochi work and one application to rare actinos)
Protoplast transformation and fusion (genome shuffling and our example of
Nonomuraea manipulation)
Link between production and resistance: examples in glycopeptide producers
Heterologous expression
L10: Greg Challis and Mervyn Bibb - Activation of cryptic pathways for drug
discovery
Greg Challis:
1. Brief overview to different approaches for identifying the metabolic products of
cryptic biosynthetic gene clusters.
2. Example of gene knockout/comparative metabolic profiling - coelichelin (and the
"answer" to the computer workshop).
3. Example of heterologous expression comparative metabolic profiling methylenomycin furans.
4. Example of induction of gene expression/comparative metabolic profiling.
5. Potential for new metabolite discovery in the genomics age.
Mervyn Bibb:
1. Identification and analysis of secondary metabolic gene clusters by “genome
scanning” – microbisporicin, cypemycin and tunicamycin
2. In vitro reconstitution of a cryptic secondary metabolic pathway - venezuelin
L11: Roberto Kolter - Microbial chemical ecology: Interspecies interactions as a
biological discovery tool
- on the need for thinking outside the box for understanding the role of secondary
metabolites
- a couple of examples:
cholesterol lowering drugs and new targets in bacteria
global warming and a novel algicide
L12: Kenji Ueda - Microbial chemical ecology: Interspecies interactions as a
fundamental of community structuring
- rethinking the role of secondary metabolism: the case of three Streptomyces
metabolites (polyethers, ferrioxamines and cobalamin)
- the pivotal role of carbon dioxide
L13: Lutz Heide - Combinatorial biosynthesis of novel natural products
1) Development of new antibiotics:
 Natural product screening
 Chemical synthesis/Combichem
 New target identification using pathogenomics
 Natural product sceening + genetic/genomic techniques
2) Combinatorial biosynthesis in vivo by gene inactivation/gene expression
a) in original producer strain
b) after heterologous expression of the clusters, including in “super-hosts”
3) Mutasynthesis: combining microbial genetics and synthetic chemistry
4) Chemoenzymatic synthesis: combining microbial genetics, synthetic chemistry and
enzymology
Examples will be used from aminocoumarins and other classes of antibiotics.
GS1: Hildgund Schrempf - Tools of streptomycetes to target, degrade and to live
on macromolecules
-Features of macromolecules within their natural habitat
-Compounds inferring with the biosynthesis of macromolecules
-Proteins targeting macromolecules
-Enzymatic repertoire
-Transport systems for the degradation products of macromolecules
-The advantage to build hyhae-aggregates
-The role of selected proteins and compounds to interact with macromolecules within
living organisms
-Future directions
GS2: Maria Mercedes Zambrano
GS3: Alison Foster – Plant natural products
Plant Natural Products – Our interactions with them.

The talk will comprise of a brief introduction to plants followed by examples
of plant natural products that are responsible for
o Taste in foods
o Fragrances and odours of flowers and leaves
o Colours of flowers and fruit
o Medicinal properties of plants
Themes running throughout will be the diversity of metabolite structures, biosynthesis
of the metabolites (not in any great detail), similarities and differences in metabolite
production within and between plant families.