Home Therapy to Reduce Office Visits for Patients with Chronic Kidney Disease and Anemia.

Anemia is a common complication of chronic kidney disease (CKD) and a predictor of increased mortality. This project integrated erythropoietin-stimulating agent (ESA) with CKD care under one practice setting, co-managing anemia with CKD while reducing frequency of office visits in a rural setting. Patients self-administered their weekly dosage of erythropoietin with monthly follow-ups. As a result, office visits decreased by 56% for patients with CKD Stage 4 and by 54% for patients with CKD Stage 5.

Evolution by leaps: gene duplication in bacteria.

BACKGROUND: Sequence related families of genes and proteins are common in bacterial genomes. In Escherichia coli they constitute over half of the genome. The presence of families and superfamilies of proteins suggest a history of gene duplication and divergence during evolution. Genome encoded protein families, their size and functional composition, reflect metabolic potentials of the organisms they are found in. Comparing protein families of different organisms give insight into functional differences and similarities. RESULTS: Equivalent enzyme families with metabolic functions were selected from the genomes of four experimentally characterized bacteria belonging to separate genera. Both similarities and differences were detected in the protein family memberships, with more similarities being detected among the more closely related organisms. Protein family memberships reflected known metabolic characteristics of the organisms. Differences in divergence of functionally characterized enzyme family members accounted for characteristics of taxa known to differ in those biochemical properties and capabilities. While some members of the gene families will have been acquired by lateral exchange and other former family members will have been lost over time, duplication and divergence of genes and functions appear to have been a significant contributor to the functional diversity of today's microbes. CONCLUSIONS: Protein families seem likely to have arisen during evolution by gene duplication and divergence where the gene copies that have been retained are the variants that have led to distinct bacterial physiologies and taxa. Thus divergence of the duplicate enzymes has been a major process in the generation of different kinds of bacteria.

Genomics of an extreme psychrophile, Psychromonas ingrahamii.

BACKGROUND: The genome sequence of the sea-ice bacterium Psychromonas ingrahamii 37, which grows exponentially at -12C, may reveal features that help to explain how this extreme psychrophile is able to grow at such low temperatures. Determination of the whole genome sequence allows comparison with genes of other psychrophiles and mesophiles. RESULTS: Correspondence analysis of the composition of all P. ingrahamii proteins showed that (1) there are 6 classes of proteins, at least one more than other bacteria, (2) integral inner membrane proteins are not sharply separated from bulk proteins suggesting that, overall, they may have a lower hydrophobic character, and (3) there is strong opposition between asparagine and the oxygen-sensitive amino acids methionine, arginine, cysteine and histidine and (4) one of the previously unseen clusters of proteins has a high proportion of "orphan" hypothetical proteins, raising the possibility these are cold-specific proteins. Based on annotation of proteins by sequence similarity, (1) P. ingrahamii has a large number (61) of regulators of cyclic GDP, suggesting that this bacterium produces an extracellular polysaccharide that may help sequester water or lower the freezing point in the vicinity of the cell. (2) P. ingrahamii has genes for production of the osmolyte, betaine choline, which may balance the osmotic pressure as sea ice freezes. (3) P. ingrahamii has a large number (11) of three-subunit TRAP systems that may play an important role in the transport of nutrients into the cell at low temperatures. (4) Chaperones and stress proteins may play a critical role in transforming nascent polypeptides into 3-dimensional configurations that permit low temperature growth. (5) Metabolic properties of P. ingrahamii were deduced. Finally, a few small sets of proteins of unknown function which may play a role in psychrophily have been singled out as worthy of future study. CONCLUSION: The results of this genomic analysis provide a springboard for further investigations into mechanisms of psychrophily. Focus on the role of asparagine excess in proteins, targeted phenotypic characterizations and gene expression investigations are needed to ascertain if and how the organism regulates various proteins in response to growth at lower temperatures.

Searchlight on domains.

In this issue of Structure, examine in detail the functions of selected domains within proteins both when they are alone and when in combination with others. Domain function is relevant to molecular evolution and to annotation of proteins known only by sequence.

Genomic analysis of carbon source metabolism of Shewanella oneidensis MR-1: Predictions versus experiments.

Genomic sequences have been used to find the genetic foundation for carbon source metabolism in Shewanella oneidensis MR-1. Annotated S. oneidensis MR-1 gene products were examined for their sequence similarity to enzymes participating in pathways for utilization of carbon and energy as described in the BioCyc database (http://www.biocyc.org/) or in the primary literature. A picture emerges that relegates five- and six-carbon sugars to minor roles as carbon sources, whereas multiple pathways for utilization of up to three-carbon carbohydrates seem to be present. Capacity to utilize amino acids for carbon and energy is also present. A few contradictions emerged in which enzymes appear to be present by annotations but are not active in the cell according to physiological experiments. Annotations are based on close sequence similarity and will not reveal inactivity due to deleterious mutations or due to lack of coordination of regulation and transport. Genes for a few enzymes known by experiment to be active are not found in the genome. This may be due to extensive divergence after duplication or convergence of function in separate lines in evolution rendering activities undetectable by sequence similarity. To minimize false predictions from protein sequences, we have been conservative in predicting pathways. We did not predict any pathway when, although a partial pathway was seen it was composed largely of enzymes already accounted for in any other complete pathway. This is an example of how a biochemically oriented sequence analysis can generate questions and direct further experimental investigation.

Escherichia coli K-12: a cooperatively developed annotation snapshot--2005.

The goal of this group project has been to coordinate and bring up-to-date information on all genes of Escherichia coli K-12. Annotation of the genome of an organism entails identification of genes, the boundaries of genes in terms of precise start and end sites, and description of the gene products. Known and predicted functions were assigned to each gene product on the basis of experimental evidence or sequence analysis. Since both kinds of evidence are constantly expanding, no annotation is complete at any moment in time. This is a snapshot analysis based on the most recent genome sequences of two E.coli K-12 bacteria. An accurate and up-to-date description of E.coli K-12 genes is of particular importance to the scientific community because experimentally determined properties of its gene products provide fundamental information for annotation of innumerable genes of other organisms. Availability of the complete genome sequence of two K-12 strains allows comparison of their genotypes and mutant status of alleles.

Gene fusions and gene duplications: relevance to genomic annotation and functional analysis.

BACKGROUND: Escherichia coli a model organism provides information for annotation of other genomes. Our analysis of its genome has shown that proteins encoded by fused genes need special attention. Such composite (multimodular) proteins consist of two or more components (modules) encoding distinct functions. Multimodular proteins have been found to complicate both annotation and generation of sequence similar groups. Previous work overstated the number of multimodular proteins in E. coli. This work corrects the identification of modules by including sequence information from proteins in 50 sequenced microbial genomes. RESULTS: Multimodular E. coli K-12 proteins were identified from sequence similarities between their component modules and non-fused proteins in 50 genomes and from the literature. We found 109 multimodular proteins in E. coli containing either two or three modules. Most modules had standalone sequence relatives in other genomes. The separated modules together with all the single (un-fused) proteins constitute the sum of all unimodular proteins of E. coli. Pairwise sequence relationships among all E. coli unimodular proteins generated 490 sequence similar, paralogous groups. Groups ranged in size from 92 to 2 members and had varying degrees of relatedness among their members. Some E. coli enzyme groups were compared to homologs in other bacterial genomes. CONCLUSION: The deleterious effects of multimodular proteins on annotation and on the formation of groups of paralogs are emphasized. To improve annotation results, all multimodular proteins in an organism should be detected and when known each function should be connected with its location in the sequence of the protein. When transferring functions by sequence similarity, alignment locations must be noted, particularly when alignments cover only part of the sequences, in order to enable transfer of the correct function. Separating multimodular proteins into module units makes it possible to generate protein groups related by both sequence and function, avoiding mixing of unrelated sequences. Organisms differ in sizes of groups of sequence-related proteins. A sample comparison of orthologs to selected E. coli paralogous groups correlates with known physiological and taxonomic relationships between the organisms.

Domain evolution and functional diversification of sulfite reductases.

Sulfite reductases are key enzymes of assimilatory and dissimilatory sulfur metabolism, which occur in diverse bacterial and archaeal lineages. They share a highly conserved domain "C-X5-C-n-C-X3-C" for binding siroheme and iron-sulfur clusters that facilitate electron transfer to the substrate. For each sulfite reductase cluster, the siroheme-binding domain is positioned slightly differently at the N-terminus of dsrA and dsrB, while in the assimilatory proteins the siroheme domain is located at the C-terminus. Our sequence and phylogenetic analysis of the siroheme-binding domain shows that sulfite reductase sequences diverged from a common ancestor into four separate clusters (aSir, alSir, dsr, and asrC) that are biochemically distinct; each serves a different assimilatory or dissimilatory role in sulfur metabolism. The phylogenetic distribution and functional grouping in sulfite reductase clusters (dsrA and dsrB vs. aSiR, asrC, and alSir) suggest that their functional diversification during evolution may have preceded the bacterial/archaeal divergence.

Global profiling of Shewanella oneidensis MR-1: expression of hypothetical genes and improved functional annotations.

The gamma-proteobacterium Shewanella oneidensis strain MR-1 is a metabolically versatile organism that can reduce a wide range of organic compounds, metal ions, and radionuclides. Similar to most other sequenced organisms, approximately 40% of the predicted ORFs in the S. oneidensis genome were annotated as uncharacterized "hypothetical" genes. We implemented an integrative approach by using experimental and computational analyses to provide more detailed insight into gene function. Global expression profiles were determined for cells after UV irradiation and under aerobic and suboxic growth conditions. Transcriptomic and proteomic analyses confidently identified 538 hypothetical genes as expressed in S. oneidensis cells both as mRNAs and proteins (33% of all predicted hypothetical proteins). Publicly available analysis tools and databases and the expression data were applied to improve the annotation of these genes. The annotation results were scored by using a seven-category schema that ranked both confidence and precision of the functional assignment. We were able to identify homologs for nearly all of these hypothetical proteins (97%), but could confidently assign exact biochemical functions for only 16 proteins (category 1; 3%). Altogether, computational and experimental evidence provided functional assignments or insights for 240 more genes (categories 2-5; 45%). These functional annotations advance our understanding of genes involved in vital cellular processes, including energy conversion, ion transport, secondary metabolism, and signal transduction. We propose that this integrative approach offers a valuable means to undertake the enormous challenge of characterizing the rapidly growing number of hypothetical proteins with each newly sequenced genome.

GenProtEC: an updated and improved analysis of functions of Escherichia coli K-12 proteins.

Using more than one approach to characterizing functions of unknown proteins, we now present in GenProtEC (http://genprotec.mbl.edu/) some level of function information for 87% of Escherichia coli K-12 proteins. A new approach that has yielded new information entails assigning content of structural domains and their functions to E.coli proteins. In addition, some earlier methods have been further refined to provide more meaningful data. The process of identifying and separating multimodular or fused proteins into component modules has been improved. As a result, groups of sequence-similar (paralogous) proteins have been refined. Experimental information from recent literature on previously unknown genes has been incorporated. We now use a rich system of characterizing cell roles which accents the fact that many proteins play more than one cellular role and therefore carry more than one designation from our detailed catalog of roles, MultiFun.

Structural domains, protein modules, and sequence similarities enrich our understanding of the Shewanella oneidensis MR-1 proteome.

The protein coding sequences of S. oneidensis MR-1 were analyzed, and new annotations were given to 491 gene products, 306 of which were previously of unknown function. New information was mainly brought in from structural domain predictions for S. oneidensis proteins of the SUPERFAM database (http://supfam.mrc-lmb.cam.ac.uk/SUPERFAMILY/) and newly identified and experimentally verified functions of homologous proteins. Proteins encoded by fused genes were identified and separated into modules, protein units of at least 83 aa with independent functions and distinct evolutionary histories. A reannotation of the fused gene products was done to assign functions to the appropriate module within the protein. Groups of sequence-similar proteins of S. oneidensis were assembled. The fused gene products were represented by their modular entities for the grouping process. The protein groups were analyzed for their size and functions, and they were used to indicate activities that are of importance to the environmental adaptation of this organism. Making use of several approaches not commonly used in annotation, we have been able to enrich our understanding of the functions encoded by the S. oneidensis genome.

Extending knowledge of Escherichia coli metabolism by modeling and experiment.

One of the challenges for 'post-genomic' biology is the integration of data from many different sources. Two recent studies independently take steps towards this goal for Escherichia coli, using mathematical modeling and a combination of gene expression and protein levels to predict new gene functions and metabolic behaviors.

Physiological genomics of Escherichia coli protein families.

The well-researched Escherichia coli genome offers the opportunity to explore the value of using protein families within a single organism to enrich functional annotation procedures and to study mechanisms of protein evolution. Having identified multimodular proteins resulting from gene fusion, and treated each module as a separate protein, nonoverlapping sequence-similar families in E. coli could be assembled. Of 3,902 proteins of length 100 residues or more, 2,415 clustered into 609 protein families. The relatedness of function among members of each family was dissected in detail. Data on paralogous protein families provides valuable information in attributing putative function to unknown genes, supplementing existing function annotation. Enzymes, transporters, and regulators represent the three major types of proteins in E. coli. They are shown to have distinctive patterns in gene duplication and divergence and gene fusion, suggesting that details of protein evolution have been different for genes in these categories. Data for the complete list of paralogous protein families and updated functional annotation for E. coli K-12 are accessible in GenProtEC (http://genprotec.mbl.edu).

The EcoCyc Database.

EcoCyc is an organism-specific pathway/genome database that describes the metabolic and signal-transduction pathways of Escherichia coli, its enzymes, its transport proteins and its mechanisms of transcriptional control of gene expression. EcoCyc is queried using the Pathway Tools graphical user interface, which provides a wide variety of query operations and visualization tools. EcoCyc is available at http://ecocyc.org/.

The MetaCyc Database.

MetaCyc is a metabolic-pathway database that describes 445 pathways and 1115 enzymes occurring in 158 organisms. MetaCyc is a review-level database in that a given entry in MetaCyc often integrates information from multiple literature sources. The pathways in MetaCyc were determined experimentally, and are labeled with the species in which they are known to occur based on literature references examined to date. MetaCyc contains extensive commentary and literature citations. Applications of MetaCyc include pathway analysis of genomes, metabolic engineering and biochemistry education. MetaCyc is queried using the Pathway Tools graphical user interface, which provides a wide variety of query operations and visualization tools. MetaCyc is available via the World Wide Web at http://ecocyc.org/ecocyc/metacyc.html, and is available for local installation as a binary program for the PC and the Sun workstation, and as a set of flatfiles. Contact metacyc-info@ai.sri.com for information on obtaining a local copy of MetaCyc.