Margarete Bauer

TETRA: a web-service and a stand-alone program for the analysis and comparison of tetranucleotide usage patterns in DNA sequences

AbstractBackgroundIn the emerging field of environmental genomics, direct cloning and sequencing of genomic fragments from complex microbial communities has proven to be a valuable source of new enzymes, expanding the knowledge of basic biological processes. The central problem of this so called metagenome-approach is that the cloned fragments often lack suitable phylogenetic marker genes, rendering the identification of clones that are likely to originate from the same genome difficult or impossible. In such cases, the analysis of intrinsic DNA-signatures like tetranucleotide frequencies can provide valuable hints on fragment affiliation. With this application in mind, the TETRA web-service and the TETRA stand-alone program have been developed, both of which automate the task of comparative tetranucleotide frequency analysis. Availability:http://www.megx.net/tetraResultsTETRA provides a statistical analysis of tetranucleotide usage patterns in genomic fragments, either via a web-service or a stand-alone program. With respect to discriminatory power, such an analysis outperforms the assignment of genomic fragments based on the (G+C)-content, which is a widely-used sequence-based measure for assessing fragment relatedness. While the web-service is restricted to the calculation of correlation coefficients between tetranucleotide usage patterns of submitted DNA sequences, the stand-alone program generates a much more detailed output, comprising all raw data and graphical plots. The stand-alone program is controlled via a graphical user interface and can batch-process a multitude of sequences. Furthermore, it comes with pre-computed tetranucleotide usage patterns for 166 prokaryote chromosomes, providing a useful reference dataset and source for data-mining.ConclusionsUp to now, the analysis of skewed oligonucleotide distributions within DNA sequences is not a commonly used tool within metagenomics. With the TETRA web-service and stand-alone program, the method is now accessible in an easy to use manner for a broad audience. This will hopefully facilitate the interrelation of genomic fragments from metagenome libraries, ultimately leading to new insights into the genetic potentials of yet uncultured microorganisms.

Archaea-Like Genes for C1-Transfer Enzymes in Planctomycetes: Phylogenetic Implications of Their Unexpected Presence in This Phylum

The unexpected presence of archaea-like genes for tetrahydromethanopterin (H4MPT)-dependent enzymes in the completely sequenced genome of the aerobic marine planctomycete Pirellula sp. strain 1 (“Rhodopirellula baltica”) and in the currently sequenced genome of the aerobic freshwater planctomycete Gemmata obscuriglobus strain UQM2246 revives the discussion on the origin of these genes in the bacterial domain. We compared the genomic arrangement of these genes in Planctomycetes and methylotrophic proteobacteria and performed a phylogenetic analysis of the encoded protein sequences to address the question whether the genes have been present in the common ancestor of Bacteria and Archaea or were transferred laterally from the archaeal to the bacterial domain and therein. Although this question could not be solved using the data presented here, some constraints on the evolution of the genes involved in archaeal and bacterial H4MPT-dependent C1-transfer may be proposed: (i) lateral gene transfer (LGT) from Archaea to a common ancestor of Proteobacteria and Planctomycetes seems more likely than the presence of the genes in the common ancestor of Bacteria and Archaea; (ii) a single event of interdomain LGT can be favored over two independent events; and (iii) the archaeal donor of the genes might have been a representative of the Methanosarcinales. In the bacterial domain, the acquired genes evolved according to distinct environmental and metabolic constraints, reflected by specific rearrangements of gene order, gene recruitment, and gene duplication, with subsequent functional specialization. During the course of evolution, genes were lost from some planctomycete genomes or replaced by orthologous genes from proteobacterial lineages.

Analysis of N-acetylglucosamine metabolism in the marine bacterium Pirellula sp. strain 1 by a proteomic approach

Pirellula sp. strain 1 is a marine bacterium that can grow with the chitin monomer N‐acetylglucosamine as sole source of carbon and nitrogen under aerobic conditions, and that is a member of the bacterial phylum Planctomycetes. As a basis for the proteomic studies we quantified growth of strain 1 with N‐acetylglucosamine and glucose, revealing doubling times of 14 and 10 h, respectively. Studies with dense cell suspensions indicated that the capacity to degrade N‐acetylglucosamine and glucose may not be tightly regulated. Proteins from soluble extracts prepared from exponential cultures grown either with N‐acetylglucosamine or glucose were separated by two‐dimensional gel electrophoresis and visualized by fluorescence staining (Sypro® Ruby). Analysis of the protein patterns revealed the presence of several protein spots only detectable in soluble extracts of N‐acetylglucosamine grown cells. Determination of amino acid sequences and peptide mass fingerprints from tryptic fragments of the most abundant one of these spots allowed the identification of the coding gene on the genomic sequence of Pirellula sp. strain 1. This gene showed similarities to a dehydrogenase from Bacillus subtilis, and is closely located to a gene similar to glucosamine‐6‐phosphate isomerase from B. subtilis. Genes of two other proteins expressed during growth on N‐acetylglucosamine as well as on glucose were also identified and found to be similar to a glyceraldehyde‐3‐phosphate‐dehydrogenase and a NADH‐dehydrogenase, respectively. Thus the coding genes of three proteins expressed during growth of Pirellula sp. strain 1 on carbohydrates were identified and related by sequence similarity to carbohydrate metabolism.