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B G Barrell

Publications and source records attributed to B G Barrell.

At least 37 records · Page 2Linked to original sources

DNA sequencing and analysis of a 67.4 kb region from the right arm of Schizosaccharomyces pombe chromosome II reveals 28 open reading frames including the genes his5, pol5, ppa2, rip1, rpb8 and skb1.

67 393 bp of contiguous DNA located between markers cdc18 and cdc14 on the right arm of fission yeast chromosome II has been sequenced as part of the European Union Schizosaccharomyces pombe genome sequencing project. The complete sequence, contained in cosmid clones c15C4 and c21H7, has been determined on both strands. Sequence analysis shows that it contains 28 open reading frames capable of coding for proteins, 16 split by one or more introns, but no tRNA, rRNA or transposon sequences. The gene density is one per 2. 4 kb. Six genes have been previously described (his5, pol5, ppa2, rip1, rpb8 and skb1) and 22 are novel. Of the novel genes, 14 have significant similarity with proteins of known function, three have similarities with proteins of unknown function and five show no extensive similarities with known proteins. Sequence similarities suggest that three of the novel genes encode ATP-dependent RNA helicases, two encode transcription factor components and others encode a G-protein, a dehydrogenase, a Rab escort protein, an Abc1-like protein, a lipase, an ATP-binding transport protein, an amino acid permease, an acid phosphatase and a mannosyltransferase.

Chromosome Mapping↗

Analysis of the proteome of Mycobacterium tuberculosis in silico.

Novel bioinformatics routines have been used to provide a more detailed definition of the proteome of Mycobacterium tuberculosis H37Rv. Over half of the current proteins result from gene duplication or domain shuffling events while one-sixth show no similarity to polypeptides described in other organisms. Prominent among the genes that appear to have been duplicated on numerous occasions are those involved in fatty acid metabolism, regulation of gene expression, and the unusually glycine-rich PE and PPE proteins. Protein similarity analysis, coupled with inspection of the genetic neighbourhood, was used to explore possible functional relatedness. This uncovered four large mce operons whose proteins may mediate initial interactions between the tubercle bacillus and host cells, together with a cluster of genes that might encode components of a structure required for secretion of ESAT-6 like proteins. Close linkage of the mmpL genes, encoding large membrane proteins, with those required for fatty acid metabolism suggests involvement in lipid transport. Compared to free-living bacteria, M. tuberculosis has a significantly smaller transport protein repertoire and this may reflect its intracellular lifestyle.

Amino Acid Motifs↗

Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence.

Countless millions of people have died from tuberculosis, a chronic infectious disease caused by the tubercle bacillus. The complete genome sequence of the best-characterized strain of Mycobacterium tuberculosis, H37Rv, has been determined and analysed in order to improve our understanding of the biology of this slow-growing pathogen and to help the conception of new prophylactic and therapeutic interventions. The genome comprises 4,411,529 base pairs, contains around 4,000 genes, and has a very high guanine + cytosine content that is reflected in the biased amino-acid content of the proteins. M. tuberculosis differs radically from other bacteria in that a very large portion of its coding capacity is devoted to the production of enzymes involved in lipogenesis and lipolysis, and to two new families of glycine-rich proteins with a repetitive structure that may represent a source of antigenic variation.

Chromosome Mapping↗

Use of a Mycobacterium tuberculosis H37Rv bacterial artificial chromosome library for genome mapping, sequencing, and comparative genomics.

The bacterial artificial chromosome (BAC) cloning system is capable of stably propagating large, complex DNA inserts in Escherichia coli. As part of the Mycobacterium tuberculosis H37Rv genome sequencing project, a BAC library was constructed in the pBeloBAC11 vector and used for genome mapping, confirmation of sequence assembly, and sequencing. The library contains about 5,000 BAC clones, with inserts ranging in size from 25 to 104 kb, representing theoretically a 70-fold coverage of the M. tuberculosis genome (4.4 Mb). A total of 840 sequences from the T7 and SP6 termini of 420 BACs were determined and compared to those of a partial genomic database. These sequences showed excellent correlation between the estimated sizes and positions of the BAC clones and the sizes and positions of previously sequenced cosmids and the resulting contigs. Many BAC clones represent linking clones between sequenced cosmids, allowing full coverage of the H37Rv chromosome, and they are now being shotgun sequenced in the framework of the H37Rv sequencing project. Also, no chimeric, deleted, or rearranged BAC clones were detected, which was of major importance for the correct mapping and assembly of the H37Rv sequence. The minimal overlapping set contains 68 unique BAC clones and spans the whole H37Rv chromosome with the exception of a single gap of approximately 150 kb. As a postgenomic application, the canonical BAC set was used in a comparative study to reveal chromosomal polymorphisms between M. tuberculosis, M. bovis, and M. bovis BCG Pasteur, and a novel 12.7-kb segment present in M. tuberculosis but absent from M. bovis and M. bovis BCG was characterized. This region contains a set of genes whose products show low similarity to proteins involved in polysaccharide biosynthesis. The H37Rv BAC library therefore provides us with a powerful tool both for the generation and confirmation of sequence data as well as for comparative genomics and other postgenomic applications. It represents a major resource for present and future M. tuberculosis research projects.

Chromosome Mapping↗

Analysis of the genome of Mycobacterium tuberculosis H37Rv.

The powerful combination of genomics and bioinformatics is providing a wealth of information about Mycobacterium tuberculosis, the aetiological agent of human tuberculosis, that will facilitate the conception and development of new therapies. The starting point for genome sequencing was the integrated map of the 4.4 Mb circular chromosome of the widely used, virulent reference strain, M. tuberculosis H37Rv. Cosmids and bacterial artificial chromosomes were selected from ordered libraries and subjected to systematic shotgun sequence analysis. This approach simplified sequence assembly as the genome is rich in repetitive DNA. In common with most bacteria, > 90% of the potential coding capacity is used, and probable or tentative functions could be attributed to > 70% of the genes. The potential biological roles of two of the principal driving forces in genome dynamics, insertion sequence elements and polymorphic multigene families are discussed.

DNA Transposable Elements↗

Identification and characterization of a G protein-coupled receptor homolog encoded by murine cytomegalovirus.

This report describes the identification of a murine cytomegalovirus (MCMV) G protein-coupled receptor (GCR) homolog. This open reading frame (M33) is most closely related to, and collinear with, human cytomegalovirus UL33, and homologs are also present in human herpesvirus 6 and 7 (U12 for both viruses). Conserved counterparts in the sequenced alpha- or gammaherpesviruses have not been identified to date, suggesting that these genes encode proteins which are important for the biological characteristics of betaherpesviruses. We have detected transcripts for both UL33 and M33 as early as 3 or 4 h postinfection, and these reappear at late times. In addition, we have identified N-terminal splicing for both the UL33 and M33 RNA transcripts. For both open reading frames, splicing results in the introduction of amino acids which are highly conserved among known GCRs. To characterise the function of the M33 in the natural host, two independent MCMV recombinant viruses were prepared, each of which possesses an M33 open reading frame which has been disrupted with the beta-galactosidase gene. While the recombinant M33 null viruses showed no phenotypic differences in replication from wild-type MCMV in primary mouse embryo fibroblasts in vitro, they showed severely restricted growth in the salivary glands of infected mice. These data suggest that M33 plays an important role in vivo, in particular in the dissemination to or replication in the salivary gland, and provide the first evidence for the function of a viral GCR homolog in vivo.

Amino Acid Sequence↗

Life with 6000 genes.

The genome of the yeast Saccharomyces cerevisiae has been completely sequenced through a worldwide collaboration. The sequence of 12,068 kilobases defines 5885 potential protein-encoding genes, approximately 140 genes specifying ribosomal RNA, 40 genes for small nuclear RNA molecules, and 275 transfer RNA genes. In addition, the complete sequence provides information about the higher order organization of yeast's 16 chromosomes and allows some insight into their evolutionary history. The genome shows a considerable amount of apparent genetic redundancy, and one of the major problems to be tackled during the next stage of the yeast genome project is to elucidate the biological functions of all of these genes.

Amino Acid Sequence↗

Analysis of the complete DNA sequence of murine cytomegalovirus.

The complete DNA sequence of the Smith strain of murine cytomegalovirus (MCMV) was determined from virion DNA by using a whole-genome shotgun approach. The genome has an overall G+C content of 58.7%, consists of 230,278 bp, and is arranged as a single unique sequence with short (31-bp) terminal direct repeats and several short internal repeats. Significant similarity to the genome of the sequenced human cytomegalovirus (HCMV) strain AD169 is evident, particularly for 78 open reading frames encoded by the central part of the genome. There is a very similar distribution of G+C content across the two genomes. Sequences toward the ends of the MCMV genome encode tandem arrays of homologous glycoproteins (gps) arranged as two gene families. The left end encodes 15 gps that represent one family, and the right end encodes a different family of 11 gps. A homolog (m144) of cellular major histocompatibility complex (MHC) class I genes is located at the end of the genome opposite the HCMV MHC class I homolog (UL18). G protein-coupled receptor (GCR) homologs (M33 and M78) occur in positions congruent with two (UL33 and UL78) of the four putative HCMV GCR homologs. Counterparts of all of the known enzyme homologs in HCMV are present in the MCMV genome, including the phosphotransferase gene (M97), whose product phosphorylates ganciclovir in HCMV-infected cells, and the assembly protein (M80).

Animals↗

Tropomyosin is essential in yeast, yet the TPM1 and TPM2 products perform distinct functions.

Sequence analysis of chromosome IX of Saccharomyces cerevisiae revealed an open reading frame of 166 residues, designated TPM2, having 64.5% sequence identity to TPM1, that encodes the major form of tropomyosin in yeast. Purification and characterization of Tpm2p revealed a protein with the characteristics of a bona fide tropomyosin; it is present in vivo at about one sixth the abundance of Tpm1p. Biochemical and sequence analysis indicates that Tpm2p spans four actin monomers along a filament, whereas Tpmlp spans five. Despite its shorter length, Tpm2p can compete with Tpm1p for binding to F-actin. Over-expression of Tpm2p in vivo alters the axial budding of haploids to a bipolar pattern, and this can be partially suppressed by co-over-expression of Tpm1p. This suggests distinct functions for the two tropomyosins, and indicates that the ratio between them is important for correct morphogenesis. Loss of Tpm2p has no detectable phenotype in otherwise wild type cells, but is lethal in combination with tpm1 delta. Over-expression of Tpm2p does not suppress the growth or cell surface targeting defects associated with tpm1 delta, so the two tropomyosins must perform an essential function, yet are not functionally interchangeable. S. cerevisiae therefore provides a simple system for the study of two tropomyosins having distinct yet overlapping functions.

Actins↗

Human herpesvirus 6 (strain U1102) encodes homologues of the conserved herpesvirus glycoprotein gM and the alphaherpesvirus origin-binding protein.

The nucleotide sequence of 3,134 bp from the genome of human herpesvirus 6 (HHV-6) strain U1102 was determined. The sequence overlaps and is contiguous with the 21,858 bp nucleotide sequence published by us previously. The sequence reported here encodes two open reading frames, named 18L and 19R. The protein encoded by 18L shares amino acid sequence similarity with the multiply hydrophobic glycoprotein M conserved in the genomes of all herpesviruses sequenced to date. ORF 19R encodes a protein which shares a significant degree of amino acid sequence conservation with the origin-binding protein homologues encoded by members of the alphaherpesvirus subgroup, but does not share detectable amino acid sequence homology with positionally analogous open reading frames present in the genomes of other betaherpesviruses or in the genomes of gammaherpesviruses.

Amino Acid Sequence↗

Spliced transcripts of human cytomegalovirus.

The availability of the human cytomegalovirus (HCMV) genomic sequence has resulted in more extensive knowledge of the overall coding capacity of the virus. Using polymerase chain reaction and rapid sequencing techniques, we have studied the splicing of mRNAs from a number of the predicted open reading frames (ORFs). Splicing was found between the UL122(IE2) ORF present within major immediate-early (MIE) region 2 and the downstream ORF (UL118) predicted to encode an incomplete glycoprotein. This locates the IE2 3' donor site and provides evidence of a link between the MIE region and downstream ORFs. The downstream UL119-UL118-UL115 ORFs also undergo differential splicing, further increasing the known complexity of this region of the genome. A detailed map of the differential splicing within the region encoding the MIE ORF is presented. Also described are several previously unidentified spliced ORFs found in the long repeats and long unique regions, including one encoding a transcript with a large (4-kb) intron. The results show that spliced transcripts are encoded from throughout the genome at immediate-early, early, and late times postinfection.

Amino Acid Sequence↗

Identification of homologues to the human cytomegalovirus US22 gene family in human herpesvirus 6.

The sequence of 10079 bp corresponding to the overlapping SalI H and SmaI G restriction fragments of the genome of human herpesvirus 6 (HHV-6) strain U1102 was determined. The sequence contains six complete open reading frames (ORFs) and two incomplete ORFs located at the 5' and 3' ends of the SalI H and SmaI G fragments respectively. Seven of these ORFs have recognizable homologues only in the beta-herpesvirus human cytomegalovirus (HCMV), no obvious counterparts being detectable in the genomes of the human alpha-herpesviruses, varicella-zoster virus and herpes simplex virus type 1 or the gamma-herpesvirus Epstein-Barr virus. The DNA sequenced is located proximal to the left repeat of the HHV-6 genome outside the well recognized region encompassing conserved herpesvirus gene blocks. A close colinear relationship is evident between the HHV-6 ORFs identified in this study and their counterparts in HCMV, ORFs UL23, UL24 and UL27 to UL31. Four of the HHV-6 ORFs, SHL1, SHL2, SFL1 and SSL2, are related to members of the HCMV US22 family of proteins, which are themselves tandemly arranged and located predominantly within the unique short and the left end of the unique long region of the prototype HCMV strain AD169 genome. Two adjacent HHV-6 ORFs, SSL1 and SHL3, are related to HCMV UL27. The identification of this gene set in addition to the HHV-6 ORFs with amino acid sequence similarity to the HCMV US22 family indicates a particularly close relationship between these two human herpesviruses, and suggests that the clustering of these related tandemly arranged genes may be a general feature of beta-herpesvirus-type genomes.

Amino Acid Sequence↗

Cloning of a yeast U1 snRNP 70K protein homologue: functional conservation of an RNA-binding domain between humans and yeast.

We have cloned and sequenced a gene encoding a yeast homologue of the U1 snRNP 70K protein. The gene, SNP1, encodes a protein which has 30% amino acid identity with the human 70K protein and has a predicted molecular weight of 34 kDa. The yeast and human sequences are more closely related to each other than to other (non-U1) RNA-binding proteins, but diverge considerably in their C-terminal portions. In particular, SNP1 lacks the charged carboxy terminus of the human 70K protein. A yeast strain, a alpha 115, was constructed in which one allele of the SNP1 gene contained a 554 bp deletion. Tetrad analysis of a alpha 115 showed that the SNP1 gene is essential for the viability of yeast cells. The complete human 70K gene did not complement snp1, but the lethal snp1 mutation was rescued by plasmids bearing a chimera in which over half the yeast gene was replaced with the homologous region of the human 70K gene, including the RNA-binding domain. These results suggest that SNP1 encodes a functional homologue of the U1 snRNP 70K protein.

Amino Acid Sequence↗

Sequence of the HindIII T fragment of human cytomegalovirus, which encodes a DNA helicase.

The DNA sequence of the HindIII T fragment of human cytomegalovirus strain AD169 has been determined. This 6225 bp sequence has been analysed for transcription signals and probable open reading frames. Similarities with herpes simplex virus, varicella-zoster virus and Epstein-Barr virus genes were observed for three of the predicted open reading frames; a virion protein and a unique DNA helicase are believed to be the functional products of two of these open reading frames. Two other open reading frames are novel in that no homologues could be found, either in the known herpesvirus sequences or in the Protein Identification Resource database. Both of these open reading frames also lie in the genomic coding region of a 5.0 kb RNA which is transcribed throughout the infectious cycle.

Amino Acid Sequence↗

Human cytomegalovirus encodes three G protein-coupled receptor homologues.

Human cytomegalovirus (HCMV) is a herpesvirus with a genome of 230 kilobases (Kb) encoding about 200 genes. Although infection is generally innocuous, HCMV causes serious congenital and neonatal disease, and is a dangerous opportunistic pathogen in immune-deficient individuals. We have identified a family of three HCMV genes which encode polypeptides containing seven putative membrane-spanning domains, and a series of well-defined motifs characteristic of the rhodopsin-like G protein-coupled receptors (GCRs). By these criteria all three of the HCMV sequences are homologous to cellular GCRs. Members of this receptor family function in visual signal transduction, regulation of homeostasis, and development, and include known and potential oncogenes. These receptors are activated by photons or small molecules such as neurotransmitters, and glycoprotein hormones. The finding of viral-encoded GCR homologues implies a further level of complexity in the interactions between HCMV and its host, and may provide a potential pathway for virally transformed cell proliferation. Their identification could permit the development of a novel class of antiviral drugs analogous to beta-adrenergic receptor antagonists.

Amino Acid Sequence↗