On the Folding and Conformation of Peptides and the Development of Novel Methods for Their Study

Proteins are polymers, whose complex chemistry and physics, in the course of evo- lution, permitted their recruitment for immensely diverse functions in the biological machinery. Therefore, the study of protein conformational behavior and interactions of proteins with other entities of the intra- and extra-cellular milieu is of great inter- est. In this Thesis, we do not only quantitatively study the folding of peptide and protein systems using well-established spectroscopic techniques, but also, in recognizing the limits of existing experimental techniques, develop novel methods for studying protein folding, protein-protein and protein-membrane interactions.In the first part of this Thesis, we are concerned with studying the folding process under various conditions using small peptides or proteins as model systems that al- low a tractable, quantitative description of the order-disorder transition. Employing a combined site-directed mutagenesis/temperature-jump kinetics approach, we eluci- dated the folding mechanism of a helical hairpin, which is an important intermediate in protein folding reactions. We further considered how the folding kinetics of such model peptides and proteins are altered in the presence of high concentrations of macromolecular cosolutes, which are thought to provide a simplified model system for the interior of the cellular environment. In another study, we addressed current questions concerning the structural properties of the unfolded state ensemble using a polypeptide that is unstructured by design as a model system. The second part of this Thesis is focused on the development of novel methods to study both protein folding and interactions of proteins with other molecular species. In recent years, the nitrile stretching bands of nitrile-derivatized amino acid side- chains have emerged as attractive probes for monitoring biomolecular processes. In this Thesis, we theoretically and experimentally studied the molecular determinants of the bandwidth of the nitrile stretching band of 5-cyanotryptophan. We further evaluated the feasibility of using light-induced reductive cleavage of disulfide bonds as a means to phototrigger protein folding reactions

Haemogregarina bigemina (Protozoa : Apicomplexa : Adeleorina) – Past, present and future

This paper reviews past, current and likely future research on the fish haemogregarine, Haemogregarina bigemina Laveran et Mesnil, 1901. Recorded from 96 species of fishes, across 70 genera and 34 families, this broad distribution for H. bigemina is questioned. In its type hosts and other fishes, the parasite undergoes intraerythrocytic binary fission, finally forming mature paired gamonts. An intraleukocytic phase is also reported, but not from the type hosts. This paper asks whether stages from the white cell series are truly H. bigemina. A future aim should be to compare the molecular constitution of so-called H. bigemina from a number of locations to determine whether all represent the same species. The transmission of H. bigemina between fishes is also considered. Past studies show that young fish acquire the haemogregarine when close to metamorphosis, but vertical and faecal-oral transmission seem unlikely. Some fish haemogregarines are leech-transmitted, but where fish populations with H. bigemina have been studied, these annelids are largely absent. However, haematophagous larval gnathiid isopods occur on such fishes and may be readily eaten by them. Sequential squashes of gnathiids from fishes with H. bigemina have demonstrated development of the haemogregarine in these isopods. Examination of histological sections through gnathiids is now underway to determine the precise development sites of the haemogregarine, particularly whether merozoites finally invade the salivary glands. To assist in this procedure and to clarify the internal anatomy of gnathiids, 3D visualisation of stacked, serial histological sections is being undertaken. Biological transmission experiments should follow these processes

The phylogeny of Nudibranchia (Opisthobranchia, Gastropoda, Mollusca) reconstructed by three molecular markers

The phylogeny of the Nudibranchia and its major constituent taxa is investigated by comparing the complete sequences of the 18S rDNA of 54 species, a part of the 16S rDNA of 38 species and part of cytochrome c oxidase I (cox1) of 45 species. These datasets are analyzed individually and in combination for the subset of taxa where information on all three markers is available. The results are compared to published cladistic analyses based on morphological data. The monophyly of the Nudibranchia and the monophyly of its two major groups, the Anthobranchia/Doridoidea and Cladobranchia, is confirmed. Incongruencies between the molecular and morphological data is discussed, as well as incongruencies between the three molecular markers

CRONOS: the cross-reference navigation server

Summary: Cross-mapping of gene and protein identifiers between different databases is a tedious and time-consuming task. To overcome this, we developed CRONOS, a cross-reference server that contains entries from five mammalian organisms presented by major gene and protein information resources. Sequence similarity analysis of the mapped entries shows that the cross-references are highly accurate. In total, up to 18 different identifier types can be used for identification of cross-references. The quality of the mapping could be improved substantially by exclusion of ambiguous gene and protein names which were manually validated. Organism-specific lists of ambiguous terms, which are valuable for a variety of bioinformatics applications like text mining are available for download

CORUM: the comprehensive resource of mammalian protein complexes—2009

CORUM is a database that provides a manually curated repository of experimentally characterized protein complexes from mammalian organisms, mainly human (64%), mouse (16%) and rat (12%). Protein complexes are key molecular entities that integrate multiple gene products to perform cellular functions. The new CORUM 2.0 release encompasses 2837 protein complexes offering the largest and most comprehensive publicly available dataset of mammalian protein complexes. The CORUM dataset is built from 3198 different genes, representing ∼16% of the protein coding genes in humans. Each protein complex is described by a protein complex name, subunit composition, function as well as the literature reference that characterizes the respective protein complex. Recent developments include mapping of functional annotation to Gene Ontology terms as well as cross-references to Entrez Gene identifiers. In addition, a ‘Phylogenetic Conservation’ analysis tool was implemented that analyses the potential occurrence of orthologous protein complex subunits in mammals and other selected groups of organisms. This allows one to predict the occurrence of protein complexes in different phylogenetic groups. CORUM is freely accessible at (http://mips.helmholtz-muenchen.de/genre/proj/corum/index.html)

OREST: the online resource for EST analysis

The generation of expressed sequence tag (EST) libraries offers an affordable approach to investigate organisms, if no genome sequence is available. OREST (http://mips.gsf.de/genre/proj/orest/index.html) is a server-based EST analysis pipeline, which allows the rapid analysis of large amounts of ESTs or cDNAs from mammalia and fungi. In order to assign the ESTs to genes or proteins OREST maps DNA sequences to reference datasets of gene products and in a second step to complete genome sequences. Mapping against genome sequences recovers additional 13% of EST data, which otherwise would escape further analysis. To enable functional analysis of the datasets, ESTs are functionally annotated using the hierarchical FunCat annotation scheme as well as GO annotation terms. OREST also allows to predict the association of gene products and diseases by Morbid Map (OMIM) classification. A statistical analysis of the results of the dataset is possible with the included PROMPT software, which provides information about enrichment and depletion of functional and disease annotation terms. OREST was successfully applied for the identification and functional characterization of more than 3000 EST sequences of the common marmoset monkey (Callithrix jacchus) as part of an international collaboration

Development of the first marmoset-specific DNA microarray (EUMAMA): a new genetic tool for large-scale expression profiling in a non-human primate

Contains fulltext : 34911.pdf (publisher's version ) (Open Access)BACKGROUND: The common marmoset monkey (Callithrix jacchus), a small non-endangered New World primate native to eastern Brazil, is becoming increasingly used as a non-human primate model in biomedical research, drug development and safety assessment. In contrast to the growing interest for the marmoset as an animal model, the molecular tools for genetic analysis are extremely limited. RESULTS: Here we report the development of the first marmoset-specific oligonucleotide microarray (EUMAMA) containing probe sets targeting 1541 different marmoset transcripts expressed in hippocampus. These 1541 transcripts represent a wide variety of different functional gene classes. Hybridisation of the marmoset microarray with labelled RNA from hippocampus, cortex and a panel of 7 different peripheral tissues resulted in high detection rates of 85% in the neuronal tissues and on average 70% in the non-neuronal tissues. The expression profiles of the 2 neuronal tissues, hippocampus and cortex, were highly similar, as indicated by a correlation coefficient of 0.96. Several transcripts with a tissue-specific pattern of expression were identified. Besides the marmoset microarray we have generated 3215 ESTs derived from marmoset hippocampus, which have been annotated and submitted to GenBank [GenBank: EF214838-EF215447, EH380242-EH382846]. CONCLUSION: We have generated the first marmoset-specific DNA microarray and demonstrated its use to characterise large-scale gene expression profiles of hippocampus but also of other neuronal and non-neuronal tissues. In addition, we have generated a large collection of ESTs of marmoset origin, which are now available in the public domain. These new tools will facilitate molecular genetic research into this non-human primate animal model

CORUM: the comprehensive resource of mammalian protein complexes

Protein complexes are key molecular entities that integrate multiple gene products to perform cellular functions. The CORUM (http://mips.gsf.de/genre/proj/corum/index.html) database is a collection of experimentally verified mammalian protein complexes. Information is manually derived by critical reading of the scientific literature from expert annotators. Information about protein complexes includes protein complex names, subunits, literature references as well as the function of the complexes. For functional annotation, we use the FunCat catalogue that enables to organize the protein complex space into biologically meaningful subsets. The database contains more than 1750 protein complexes that are built from 2400 different genes, thus representing 12% of the protein-coding genes in human. A web-based system is available to query, view and download the data. CORUM provides a comprehensive dataset of protein complexes for discoveries in systems biology, analyses of protein networks and protein complex-associated diseases. Comparable to the MIPS reference dataset of protein complexes from yeast, CORUM intends to serve as a reference for mammalian protein complexes

Quasiparticle interfacial level alignment of highly hybridized frontier levels: H2_2O on TiO2_2(110)

Knowledge of the frontier levels' alignment prior to photo-irradiation is necessary to achieve a complete quantitative description of H$_2$O photocatalysis on TiO$_2$(110). Although H$_2$O on rutile TiO$_2$(110) has been thoroughly studied both experimentally and theoretically, a quantitative value for the energy of the highest H$_2$O occupied levels is still lacking. For experiment, this is due to the H$_2$O levels being obscured by hybridization with TiO$_2$(110) levels in the difference spectra obtained via ultraviolet photoemission spectroscopy (UPS). For theory, this is due to inherent difficulties in properly describing many-body effects at the H$_2$O-TiO$_2$(110) interface. Using the projected density of states (DOS) from state-of-the-art quasiparticle (QP) $G_0W_0$, we disentangle the adsorbate and surface contributions to the complex UPS spectra of H$_2$O on TiO$_2$(110). We perform this separation as a function of H$_2$O coverage and dissociation on stoichiometric and reduced surfaces. Due to hybridization with the TiO$_2$(110) surface, the H$_2$O 3a$_1$ and 1b$_1$ levels are broadened into several peaks between 5 and 1 eV below the TiO$_2$(110) valence band maximum (VBM). These peaks have both intermolecular and interfacial bonding and antibonding character. We find the highest occupied levels of H$_2$O adsorbed intact and dissociated on stoichiometric TiO$_2$(110) are 1.1 and 0.9 eV below the VBM. We also find a similar energy of 1.1 eV for the highest occupied levels of H$_2$O when adsorbed dissociatively on a bridging O vacancy of the reduced surface. In both cases, these energies are significantly higher (by 0.6 to 2.6 eV) than those estimated from UPS difference spectra, which are inconclusive in this energy region. Finally, we apply self-consistent QP$GW$ (scQP$GW$1) to obtain the ionization potential of the H$_2$O-TiO$_2$(110) interface.Comment: 12 pages, 12 figures, 1 tabl

The BridgeDb framework: standardized access to gene, protein and metabolite identifier mapping services

BACKGROUND: Many complementary solutions are available for the identifier mapping problem. This creates an opportunity for bioinformatics tool developers. Tools can be made to flexibly support multiple mapping services or mapping services could be combined to get broader coverage. This approach requires an interface layer between tools and mapping services. RESULTS: Here we present BridgeDb, a software framework for gene, protein and metabolite identifier mapping. This framework provides a standardized interface layer through which bioinformatics tools can be connected to different identifier mapping services. This approach makes it easier for tool developers to support identifier mapping. Mapping services can be combined or merged to support multi-omics experiments or to integrate custom microarray annotations. BridgeDb provides its own ready-to-go mapping services, both in webservice and local database forms. However, the framework is intended for customization and adaptation to any identifier mapping service. BridgeDb has already been integrated into several bioinformatics applications. CONCLUSION: By uncoupling bioinformatics tools from mapping services, BridgeDb improves capability and flexibility of those tools. All described software is open source and available at http://www.bridgedb.org