The Helmholtz Network for Bioinformatics: an integrative web portal for bioinformatics resources.

SUMMARY: The Helmholtz Network for Bioinformatics (HNB) is a joint venture of eleven German bioinformatics research groups that offers convenient access to numerous bioinformatics resources through a single web portal. The 'Guided Solution Finder' which is available through the HNB portal helps users to locate the appropriate resources to answer their queries by employing a detailed, tree-like questionnaire. Furthermore, automated complex tool cascades ('tasks'), involving resources located on different servers, have been implemented, allowing users to perform comprehensive data analyses without the requirement of further manual intervention for data transfer and re-formatting. Currently, automated cascades for the analysis of regulatory DNA segments as well as for the prediction of protein functional properties are provided. AVAILABILITY: The HNB portal is available at http://www.hnbioinfo.de

Associative database of protein sequences.

MOTIVATION: We present a new concept that combines data storage and data analysis in genome research, based on an associative network memory. As an illustration, 115 000 conserved regions from over 73 000 published sequences (i.e. from the entire annotated part of the SWISSPROT sequence database) were identified and clustered by a self-organizing network. Similarity and kinship, as well as degree of distance between the conserved protein segments, are visualized as neighborhood relationship on a two-dimensional topographical map. RESULTS: Such a display overcomes the restrictions of linear list processing and allows local and global sequence relationships to be studied visually. Families are memorized as prototype vectors of conserved regions. On a massive parallel machine, clustering and updating of the database take only a few seconds; a rapid analysis of incoming data such as protein sequences or ESTs is carried out on present-day workstations. AVAILABILITY: Access to the database is available at http://www.bioinf.mdc-berlin.de/unter2.html++ + CONTACT: (hanke,lehmann,reich)@mdc-berlin.de; bork@embl-heidelberg.de

Merging extracellular domains: fold prediction for laminin G-like and amino-terminal thrombospondin-like modules based on homology to pentraxins.

Using a new method for construction and database searches of sequence consensus strings, we have identified a new superfamily of protein modules comprising laminin G, thrombospondin N and the pentraxin families. The conserved patterns correspond mainly to hydrophobic core residues located in central beta strands of the known three-dimensional structures of two pentraxins, the human C-reactive protein and the serum amyloid P-component. Thus, we predict a similar jellyroll fold for all members of this superfamily. In addition, the conservation of two exposed aspartate residues in the majority of superfamily members suggests hitherto unrecognised functional sites.

Use of a modified binding model for the investigation of affinity dependence on antibody concentration in immunoassay systems.

Affinity parameters obtained from standard calibration curves of radioimmunoassays (RIA) have been shown to decrease with increasing antibody concentration. A modified binding model was introduced in which not only binding capacity but also affinity was influenced by the antibody concentration. This allows a consistent overall description of RIA standards and titration curves for the characterization of binding assays.

Kohonen map as a visualization tool for the analysis of protein sequences: multiple alignments, domains and segments of secondary structures.

The method of Kohonen maps, a special form of neural networks, was applied as a visualization tool for the analysis of protein sequence similarity. The procedure converts sequence (domains, aligned sequences, segments of secondary structure) into a characteristic signal matrix. This conversion depends on the property or replacement score vector selected by the user. Similar sequences have small distance in the signal space. The trained Kohonen network is functionally equivalent to an unsupervised non-linear cluster analyzer. Protein families, or aligned sequences, or segments of similar secondary structure, aggregate as clusters, and their proximity may be inspected on a color screen or on paper. Pull-down menus permit access to background information in the established text-oriented way.

Self-organizing hierarchic networks for pattern recognition in protein sequence.

We present a method based on hierarchical self-organizing maps (SOMs) for recognizing patterns in protein sequences. The method is fully automatic, does not require prealigned sequences, is insensitive to redundancy in the training set, and works surprisingly well even with small learning sets. Because it uses unsupervised neural networks, it is able to extract patterns that are not present in all of the unaligned sequences of the learning set. The identification of these patterns in sequence databases is sensitive and efficient. The procedure comprises three main training stages. In the first stage, one SOM is trained to extract common features from the set of unaligned learning sequences. A feature is a number of ungapped sequence segments (usually 4-16 residues long) that are similar to segments in most of the sequences of the learning set according to an initial similarity matrix. In the second training stage, the recognition of each individual feature is refined by selecting an optimal weighting matrix out of a variety of existing amino acid similarity matrices. In a third stage of the SOM procedure, the position of the features in the individual sequences is learned. This allows for variants with feature repeats and feature shuffling. The procedure has been successfully applied to a number of notoriously difficult cases with distinct recognition problems: helix-turn-helix motifs in DNA-binding proteins, the CUB domain of developmentally regulated proteins, and the superfamily of ribokinases. A comparison with the established database search procedure PROFILE (and with several others) led to the conclusion that the new automatic method performs satisfactorily.

A simple statistical significance test of window scores in large dot matrices obtained from protein or nucleic acid sequences.

A test of statistical significance of dot constellations as detected by window search in large dot matrices is described. The procedure takes the correlation between overlapping windows on the diagonals of dot matrices into account. It is based on a confidence limit of the exact distribution of dot scores.

Unusual occurrence of EcoP1 and EcoP15 recognition sites and counterselection of type II methylation and restriction sequences in bacteriophage T7 DNA.

Selected and counterselected oligodeoxynucleotide sequences were identified in the total sequence of bacteriophage T7 DNA using a statistical criterion derived for a probability model of the Markov chain type. All extremely rare tetra- and pentadeoxynucleotides are (or contain) recognition sequences for the Escherichia coli DNA methylases dam or dcm. Most of the 37 hexadeoxynucleotides absent from T7 DNA are recognition sequences for type II modification/restriction enzymes of E. coli or related species. In contrast to most restriction sites counterselected during evolution, the EcoP1 site GGTCT occurs 126 times in the T7 genome, and phage T7 replication is severely repressed in P1-lysogenic host cells. We demonstrate that the frequency of the EcoP1 site is determined by that of the overlapping recognition sites for T7 primase, an essential phage enzyme. The recognition site of a type III enzyme, EcoP15, is also not counterselected. In T7 DNA all 36 EcoP15 sites are arranged in such a manner that the sequence CAGCAG is confined to the H strand, the complementary sequence CTGCTG to the L strand. This "strand bias" is highly significant and, therefore, very probably selected. A functional relation between this strand bias and the refractive behaviour of phage T7 to EcoP15 restriction is suspected.

Dinucleotide frequencies in different reading frame positions of coding mammalian DNA sequences.

A statistical model for the assessment of suppressions or preferences of 16 dinucleotides in DNA sequences was developed. It is based on the description by a hypergeometric distribution of the doublet frequencies in randomly "scrambled" DNA sequences. The statistical test is sequential and extracts one after another dinucleotides that differ significantly from their expected values. It is shown that in mammalian DNA only TA and CG are consistently depressed in all three reading frame positions. The deviations of other dinucleotides are either restricted to one frame position or not significant. The possibility that the coding commitments of the DNA sequences may be the causes of the non-random distribution was studied. Only in position 1/2 of the reading frame is the frequency behavior of TA adequately explained by the amino acid sequence coded for. It is concluded that TA and CG are avoided wherever possible for reasons that do not reside in the coding function of mammalian DNA sequences.

Dinucleotide frequencies in different reading frame positions of coding bacterial DNA sequences.

Results of the analysis of the dinucleotide frequencies in different frame positions of coding bacterial sequences are presented. They are compared with those obtained in mammalian sequences. It is concluded that the dinucleotide frequencies in both types of sequences are caused by different influence factors.

On optimum properties in the design of metabolic and epigenetic systems.

The actual potential of a metabolic system is determined by its enzyme "spectrum", i.e. the amount of the various enzymes present. The paper addresses the problem whether this spectrum can be designed in accordance with a general principle. The long prehistory of selection of any real metabolic system suggests the optimal adaptation to a given set of selection criteria as a possible design principle. It is shown that not every (bio)chemical reactor automatically displays an optimal construction in this sense, but metabolic systems are structured such that they can do so. Structural features that lead to optimizable dynamics include: kinetic autocatalysis, stoichiometric autocatalysis, competition for the allocation of biosynthetic capacity, required economy in external resources of the system, and competition of systems for common nutrients or other resources. In these circumstances the enzyme spectrum has a distinct optimum that can be approached by stepwise adaptation and selection.

Adaptation games between microorganisms sharing a common substrate niche.

Microorganisms adapt their enzymic outfit to the ambient substrate supply. If several species compete for a limiting substrate, then reciprocal influence on the state of adaptation results, which has been envisaged in the paper as strategic game. The most important types of strategies, which are assessed by the governing selection rule, are called aggressive, neutral and cooperative. A cooperative strategy brings the highest advantage, but triggers the temptation to increase the gain by defection. The neutral strategy, i.e. acting as if the competitor were alone in the medium, is promising only when the selection rule favours product maximization, whereas as aggressive strategy, e.g. maximizing the difference between own and foreign profit, is most effective on growth rate maximization as selection criterion. Gruelling competition reduces the metabolic output and weakens the community as a whole against other systems.

On the statistical assessment of similarities in DNA sequences.

The statistical behavior of the similarity score for unrelated DNA sequences calculated as letter-by-letter comparison or from various forms of optimal alignment was studied. It was found that natural DNA-sequences from a data base and true random sequences show the same statistical behavior in terms of such scores. This makes it possible to adopt a simple criterion for the rejection of fortuitous similarity. It is based on the mean and standard deviation of chance scores whose expected values, depending on chain length, gap penalty and probability of letter coincidence, may be calculated from formulae given in the paper.

Theoretical study of oscillatory and resonance phenomena in an open system with induction of enzyme by substrate.

A simple mathematical model of an open metabolic system is constructed in which the metabolism of a substrate is enhanced by induction of the catalyzing enzyme. It is demonstrated that the steady-state in such a system may be approached by damped oscillations of substrate and enzyme levels. If the input rate of substrate into the system has periodic fluctuations, the phenomenon of resonance is observed. Numerical analysis reveals the non-linear character of the resonance.

[Selection criteria in metabolic systems]. / Uber Selektionskriterien in metabolischen Systemen.

In a given environment the physico-chemical parameters of a metabolic system are externally prescribed constant quantities. The living cell, however, is able to regulate the catalytic activities through its programme of gene expression. The paper investigates to what extent this programme is amenable to optimization by evolutionary selection. Abstract, but typical mathematical models of the cellular metabolic system together with the epigenetic metabolism of biomacromolecules (enzyme protein, structural protein) were formulated and evaluated. Selection needs a criterion and the existence of an optimum of parameter values with respect to that criterion. As selection criterion served the growth rate of the cell. Parameters were the rate constants of metabolic reactions whose values may be chosen by regulation of gene expression. Two main types of parameters were distinguished: those which influence the growth rate in a monotonous manner, and those whose positive influence has a maximum. It was found that a maximum is defined if the parameter is part of an autocatalytic cycle limited by competition. The most common such situation is that of a biomacromolecule contributing directly or indirectly, but in an essential manner, to biosynthesis and growth, while competing at the same time for the share in biosynthesis. As the rate of biosynthesis has a natural upper bound, and as most proteins as end products of biosynthesis play an essential role in other parts of metabolism, the conclusion was reached that most of the controllable parameters of gene expression are amenable to evolutionary optimization.

On the relationship between Z delta pH and delta psi as components of the protonmotive potential in Mitchell's chemiosmotic system.

A basic system of oxidative phosphorylation in steady-state taking into account secondary ion translocations has been considered. It could be shown that over a relatively wide range of parameters the relation between the osmotic and the electric component of the proton-motive potential turned out to be constant. Under conditions of a small leakage a simple approximated formula for the ratio of both components could be given. The results allow in all kinetic equations of the oxidative phosphorylation to express the protonmotive potential and its electric component simply by its other component, the pH-dependent potential.

Theoretical analysis of the glutamate dehydrogenase kinetics under physiological conditions.

A kinetic model of the glutamate dehydrogenase reaction has been formulated for the reversible reaction including all seven reactants (substrates and cofactors NAD(H) and NADP(H)). The model parameters have been evaluated from published initial-rate data. Analysis of the model at cofactor concentration near to that in the intact mitochondrion has shown that the competition for active sites between cofactors and substrates simultaneously present in mitochondria diminishes the steady-state rate of the reaction by a factor of 10 to 100 as compared to the maximal reaction rate. The model predicts near-equilibrium of the reaction substrates with NAD+/NADH cofactor pair and off-equilibrium with NADP+/NADPH. Substrate cycling with futile transfer of hydrogen from NADP+-system to NAD+-system has been found to account under in vivo conditions for no more than 2% of the maximal glutamate dehydrogenase activity in the mitochondria.

[The economy of protein maintenance in the living cell]. / Zur Okonomie im Proteinhaushalt der lebenden Zelle.

On the economy of proteins in living cells Each synthesized protein of a cell is of certain metabolic advantage, but at the same time also a load on the energy and material metabolism. "Costs" as well as "profit" vary with the amount of protein formed, and the optimal amount would maximize the profitability, i.e. profit minus cost. The paper deals with the special case of an enzyme necessary for substrate uptake, which is therefore of advantage for the energy budget of the cell, whose biosynthesis, however, drains this budget at the same time. This example allows a mathematical formulation of the concept of profitability and shows, with the help of a basic kinetic model that indeed an optimum exists. Its existence follows from the rather general fact that the advantage of an enzyme reaches a saturation value, if its amount increases, while the "cost factor" continues to increase in a linear manner.

Theoretical study of an energy metabolizing system satisfying Mitchell's postulates.

An energy metabolizing system was modelled, which carries as characteristic features substrate uptake and product extrusion, respiration and ATP-synthesis, coupled by the protonmotive potential corresponding to Mitchell's chemiosmotic hypothesis of oxidative phosphorylation. The influence of the electric part of the protonmotive potential on the rate laws of the different processes has been taken into account by a phenomenological exponent of the proton concentration, thus fortifying the osmotic concentration part of the protonmotive potential. The steady-state behaviour of the model was investigated. It could be shown that a model of such kind reveals multistationarity, homeostasis and trigger behaviour. Curves of respiration control (respiration velocity over ADP/(ATP + ADP) or ATP/ADP) are in qualitative accordance with experimental facts (state 3 and state 4 of mitochondria) as well as curves describing the effect of uncouplers on the respiratory rate.