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PubMed1991

Bluetongue virus evolution: sequence analyses of the genomic S1 segments and major core protein VP7.

The S1 segments, encoding the group-specific antigen, VP7, from the five United States prototype BTV serotypes were cloned as full-length entities. The nucleotide and deduced amino acid sequences of segment S1 of BTV-2 were determined and compared with BTV-10, -11, -13, and -17, completing the sequencing of this cognate gene segment from all five US BTV serotypes. Each segment is 1156 bp long and contains an open reading frame encoding the 349-amino acid VP7 protein. Most (greater than 94%) of the amino acids of VP7 among the serotypes are conserved, including the location (position 255) of a single lysine residue. Secondary structure analyses of VP7 predict a putative eight-stranded beta-barrel between amino acid positions 150 and 250, a structure similar to that observed in ssRNA viruses. The S1 genes are flanked by conserved 5' and 3' noncoding regions. Stem-loop structures are predicted at the 3' end of each gene (nucleotide positions 1058-1097). The S1 segments of BTV-2, -10, -11, and -17 have greater than 93% of the nucleotides conserved, while less than 80% of their bases are identical with BTV-13. Analyses of nucleotide mismatches in each codon position of the VP7 open reading frame, transition frequencies, and evolutionary distances show that of the five, BTV-13 is the most distantly related and that BTV-10 and -17 are the most closely related serotypes. Evolutionary distance calculations of segment L2 from BTV-10, -11, and -17 concur with these observations. Comparison of this relationship with hybridization data of segment M3, which codes for VP5, suggests that BTV-17 has evolved by a combination of genetic drift and genomic reassortment. The data also indicate that the five US BTV serotypes are derived from two distinct gene pools. Evolution distances were used to estimate an evolution rate of 2.2 x 10(-3) nucleotide substitution/site/year for BTV segment S1. This rate is similar to the genes of retroviruses and implies an absence of RNA polymerase proofreading activity for dsRNA viruses.

Amino Acid Sequence↗
PubMedDate not supplied

[Analysis of the nucleotide sequence of a fragment (92-100%) of the CELO avian adenovirus genome].

The nucleotide sequence of 92-100% of the adenovirus CELO (FAV1), strain Phelps, genome has been determined. The computer analysis of the sequences revealed a ClaI site methylated by m*Ecodam. A recognition site for XbaI and two sites for PstI, not found in the corresponding genome of CELO, strain Ote, have been determined. Three extensive (more than 100 amino acid residues) open reading frames exist, coding for the polypeptides with molecular weights of 31.5, 19.3 and 14.5 kD (276, 178 and 128 amino acids, respectively). Some shorter open reading frames have been detected as well within the sequences studied.

Amino Acid Sequence↗
PubMed1994

Nucleotide sequence analysis of an 8887 bp region of the left arm of yeast chromosome XIV, encompassing the centromere sequence.

The nucleotide sequencing of 8887 bp of the left arm of chromosome XIV is described. The sequence includes the centromeric region. Both strands were sequenced with an average redundancy of 5.09 per base pair. The overall G+C content is 37.3% (39.2% for putative coding regions versus 32.5% for non-coding regions). Six open reading frames (ORFs) greater than 100 amino acids were detected, all of which are completely confined to the 8.9 kbp region. Codon frequencies of the six ORFs agree with codon usage in Saccharomyces cerevisiae and all show the characteristics of low-level expressed genes. Comparison of the translated sequences with protein sequences in data bases suggests the presence of two ORFs (N2014 and N2007) encoding ribosomal proteins, the latter of which is the previously sequenced MRP7 gene. Another ORF (N2012) could encode a membrane-associated protein since it contains secretory signal sequence and two presumed transmembrane helices. This protein might be involved in mitochondrial energy transfer. ORF N2016 is immediately adjacent to the centromere, suggesting that it corresponds to the SPO1 gene, which is very tightly linked to the centromere at the left arm side of chromosome XIV (Mortimer et al., 1989).

Base Composition↗
PubMed1992

The sequence of an 8 kb segment on the left arm of chromosome II from Saccharomyces cerevisiae identifies five new open reading frames of unknown functions, two tRNA genes and two transposable elements.

The DNA sequence of an 8079 bp ClaI fragment located at 40 kb from the centromere on the left arm of chromosome II from Saccharomyces cerevisiae has been determined. Sequence analysis reveals five new open reading frames, tRNA(Gly) and tRNA(Leu) genes as well as sigma and truncated delta elements. The disruption of the three larger open reading frames shows that they are not essential for mitotic growth.

Amino Acid Sequence↗
PubMedDate not supplied

Characterization of the radish mitochondrial orfB locus: possible relationship with male sterility in Ogura radish.

The orfB locus of the normal (fertile) and Ogura (male-sterile) radish mitochondrial genomes has been characterized in order to determine if this region, which has previously been correlated with cytoplasmic male sterility (CMS) in Brassica napus cybrids (Bonhomme et al. 1991; Temple et al. 1992), could also be involved in radish CMS. In normal radish, orfB is expressed as a 600-nucleotide (nt) transcript. In Ogura radish, orfB is present as the second gene of a 1200-nt transcript that also contains a 138-codon open reading frame (orf138). Sequences showing similarity to orf138 are present in normal radish, but are not expressed.

Amino Acid Sequence↗
PubMed1990

Open reading frames on plus strand genome of duck hepatitis B virus.

Hepadnaviruses have open reading frames of surface, core, polymerase and X protein on the minus strand of the genome. We analyzed a plus strand of duck hepatitis B virus (DHBV) for the presence of open reading frame with a computer program. We found one frame on the area that is complement to 3' end of polymerase gene and 5' end of precore gene. That open reading frame is conserved in three strains of DHBV, and the predicted protein is about 80 amino acids.

Amino Acid Sequence↗
PubMed1996

The LYS5 gene of Saccharomyces cerevisiae.

The LYS2 and LYS5 genes of Saccharomyces cerevisiae together encode the 180-kDa alpha-aminoadipate reductase (AAR) in the biosynthetic pathway of lysine. The 4.8-kb LYS2 gene encodes the 155-kDa subunit of AAR. The complete nucleotide (nt) sequence of the 1.1-kb LYS5 gene is presented in this report. It contains a single continuous open reading frame of 816 nt encoding a 272-amino-acid, 30.6-kDa polypeptide.

Aldehyde Oxidoreductases↗
PubMed1996

Cloning and genetic organization of the bacteriocin 31 determinant encoded on the Enterococcus faecalis pheromone-responsive conjugative plasmid pYI17.

The conjugative plasmid pYI17 (57.5 kb) isolated from Enterococcus faecalis YI717 confers a pheromone response on the host and encodes the bacteriocin 31 gene. Bacteriocin 31 is active against E. hirae 9790, E. faecium, and Listeria monocytogenes. pYI17 was mapped physically by restriction enzyme analysis and the relational clone method. Deletion mutant and sequence analyses of the EcoRI fragment B cloned from pYl17 revealed that a 1.0-kb fragment contained the bacteriocin gene (bacA) and an immunity gene (bacB). This fragment induced bacteriocin activity in E. faecalis OG1X and E. hirae 9790. The bacA gene is located on the pYI17 physical map between 3.37 and 3.57 kb, and bacB is located between 3.59 kb and 3.87 kb, bacA encodes 67 amino acids, and bacB encodes 94 amino acids. The deduced amino acid sequence of the bacA protein contained a series of hydrophobic residues typical of a signal sequence at its amino terminus. The predicted mature bacA protein (43 amino acids) showed sequence homology with the membrane-active class II bacteriocins of lactic acid bacteria. Analysis of Tn5 insertion mutants and the resulting transcripts indicated that these genes are transcribed as an operon composed of bacA, bacB, and an open reading frame located downstream of bacB designated ORF3.

Amino Acid Sequence↗
PubMed1992

Identification and expression of rpo19, a vaccinia virus gene encoding a 19-kilodalton DNA-dependent RNA polymerase subunit.

The vaccinia virus DNA-dependent RNA polymerase subunit gene rpo19 was identified, and its expression was examined at RNA and protein levels. Antibody to the multisubunit RNA polymerase purified from virions reacted with a polypeptide with an apparent Mr of 21,000 that was synthesized in reticulocyte lysates programmed with (i) mRNA from infected cells that was isolated by hybridization to DNA subclones of the viral genomic HindIII A fragment and (ii) mRNA made in vitro by transcription of the viral open reading frame A6R. Polyclonal antiserum, raised to a recombinant protein product of the A6R open reading frame which could encode an 18,996-Da protein with an acidic N terminus, reacted with Mr-21,000 and -22,000 polypeptides that cosedimented with purified RNA polymerase. Internal sequencing of the two polypeptides confirmed that both were encoded by A6R, and the gene was named rpo19 to indicate the predicted molecular mass of the polypeptide in kilodaltons. Immunoblotting and metabolic labeling of infected cell proteins indicated that synthesis of the Mr-21,000 polypeptide started early and continued throughout virus infection, whereas the Mr-22,000 form appeared late following DNA replication. RNA analyses suggested that the rpo19 mRNA was expressed from a dual early/late promoter and that the protein-coding region of the mRNA was directly preceded by a short 5' poly(A) leader, apparently initiated within the TAAATG motif at the beginning of the open reading frame.

Amino Acid Sequence↗
PubMed1992

Sequence and transcriptional analysis of the Escherichia coli rnt gene encoding RNase T.

The Escherichia coli rnt gene encoding the enzyme RNase T, which is responsible for the end-turnover of tRNA, was cloned on a 1.5-kilobase DNA fragment. When placed in pUC18 and pUC19 vectors this fragment led to approximately a 40-fold overexpression of RNase T activity. The cloned fragment was sequenced and was found to contain an open reading frame sufficient to encode a protein of 215 amino acids with a molecular weight of 23,521, which is close to the subunit molecular weight of RNase T; the fragment also contains a second incomplete open reading frame with some sequence similarity to RNA helicases. The derived sequence of RNase T showed no similarity to any of the other E. coli exoribonucleases sequenced to date. Primer extension analysis and deletion of part of the upstream region were used to identify the transcription start point and the promoter of the rnt gene. Northern and primer extension analysis revealed that the rnt message also included the second open reading frame, suggesting that rnt is part of an operon.

Amino Acid Sequence↗
PubMed1993

Sequence of the open reading frame of the FLO1 gene from Saccharomyces cerevisiae.

The cloned part of the flocculation gene FLO1 of Saccharomyces cerevisiae (Teunissen, A.W.R.H., van den Berg, J.A. and Steensma, H.Y. (1993). Physical localization of the flocculation gene FLO1 on chromosome I of Saccharomyces cerevisiae, Yeast, in press) has been sequenced. The sequence contains a large open reading frame of 2685 bp. The amino acid sequence of the putative protein reveals a serine- and threonine-rich C-terminus (46%), the presence of repeated sequences and a possible secretion signal at the N-terminus. Although the sequence is not complete (we assume the missing fragment consists of repeat units), these data strongly suggest that the protein is located in the cell wall, and thus may be directly involved in the flocculation process.

Amino Acid Sequence↗
PubMed1993

Characterization of the left 4 kb of conjugative transposon Tn916: determinants involved in excision.

The rate-limiting step in movement of the conjugative transposon Tn916, originally identified in Enterococcus faecalis, is believed to be an excision event that generates a non-replicative circular intermediate. When present on a plasmid vector in Escherichia coli, Tn916 generally excises at a high frequency. It was reported previously that insertion of Tn5 in a region near the left end of Tn916 eliminated the ability to excise; and the mutation could be complemented in trans. In this communication the nucleotide sequence of 4 kb of Tn916 DNA connecting the recently sequenced tet(M) determinant (Su et al., 1992; Burdett, 1990) with the left end of the transposon. Ten open reading frames (ORFs) were deduced, two of which (ORF3 and ORF4) were encoded in-frame within a third (ORF2). Mutants with Tn5 insertions in the ORF1 or ORF2 (ORF3 and ORF4) were defective in excision, but could be complemented in vivo by a co-resident plasmid containing the ORF1 or ORF2 determinant, respectively. The data support the view that both ORF1 and ORF2 are essential for excision. ORF1 and ORF2 are essentially identical to determinants designated xis-Tn and int-Tn, respectively, in the closely related Tn1545. A Tn5 insertion in ORF5 eliminated conjugative transfer between E. faecalis strains. Functions for the remaining ORFs (ORF6 through ORF10) remain unknown; however, nucleotide sequences within ORF6 and ORF9 had significant homology with sequence downstream of other tet(M) determinants.

Amino Acid Sequence↗
PubMed1993

Analysis of a region from the bacteriophage resistance plasmid pCI528 involved in its conjugative mobilization between Lactococcus strains.

A 10-kb HindIII fragment of pCI528 cloned into the nonconjugative shuttle vector pCI3340 could be transferred by conjugative mobilization from Lactococcus lactis subsp. lactis MG1363, whereas other HindIII fragments of pCI528 or the vector alone were nonmobilizable. Subcloning of this 10-kb region identified a 4.4-kb BglII-EcoRI fragment which contained all the DNA essential for transfer. Sequence analysis of a 2-kb region within this 4.4 kb-segment revealed a region rich in inverted repeats and two potential overlapping open reading frames, one of which demonstrated homology to mobilization proteins of two nonconjugative staphylococcal plasmids.

Amino Acid Sequence↗
PubMed1989

Identification of a T4 gene required for bacteriophage mRNA processing.

A ribonucleolytic activity that cleaves within the Shine/Dalgarno sequences of the bacteriophage T4 motA and ORF2 mRNAs was recently described. We have identified additional sites of processing within several other ribosome binding sites, including two sites in the polycistronic frd transcript. Deletion mutants (farP) that overproduce the product of frd are defective in this mRNA processing. The mutants were used to identify processing events dependent on the T4 activity including attack at nuclease-sensitive sites within the coding sequences of some genes and within the intercistronic region 5' of gene 43. All known processing sites lie within similar sequences. Another mutant in mRNA processing carries a point mutation in one of the open reading frames (orf61.9) removed by the farP deletions. Introduction of a cloned copy of this open reading frame into a unique site in the chromosome of farP phage is sufficient to restore mRNA processing capability. The open reading frame probably encodes the T4 regB protein.

Base Sequence↗
PubMed1994

Isolation of a yeast tropomyosin-related cDNA clone that encodes a novel transmembrane protein having a C-terminal highly basic region.

A novel cDNA clone was isolated from a yeast Saccharomyces cerevisiae lambda gt11 cDNA library using rabbit anti-rat tropomyosin (TM30nm) polyclonal antibody (RTM8-2). It consists of an open reading frame of 951 bp, encoding 317 amino acid residues. The putative sequence recognized by RTM8-2 was present in Asn-235 to Thr-250. The deduced amino acid sequence and hydropathy plot suggested that this protein has a tropomyosin-homologous sequence, a predicted transmembrane, and a C-terminal basic region. A search of the data bases (EMBL and GenBank) revealed that the 128bp sequence in the 3' untranslated region (3'UTR) is almost identical (96.9%) to the human cDNA clone 54E05 sequence (EMBL accession number Z15978).

Amino Acid Sequence↗
PubMed1993

Distribution and characterization of plasmid-related sequences in the chromosomal DNA of different thermophilic Methanobacterium strains.

The genomes of several thermophilic members of the genus Methanobacterium were analyzed for homology to the related restriction-modification plasmids pFV1 and pFZ1 from M. thermoformicicum strains THF and Z-245, respectively. Two plasmid regions, designated FR-I and FR-II, could be identified with chromosomal counterparts in six Methanobacterium strains. Multiple copies of the pFV1-specific element FR-I were detected in the M. thermoformicicum strains CSM3, FF1, FF3 and M. thermoautotrophicum delta H. Sequence analysis showed that one FR-I element had been integrated in almost identical sequence contexts into the chromosomes of the strains CSM3 and delta H. Comparison of the FR-I elements from these strains with that from pFV1 revealed that they consisted of two subfragments, boxI (1118 bp) and boxII (383 bp), the order of which is variable. Each subfragment was identical on the sequence level with the corresponding plasmid-borne element and was flanked by terminal direct repeats with the consensus sequence A(A/T)ATTT. These results suggest that FR-I represents a mobile element. FR-II was located on both plasmids pFV1 and pFZ1, and on the chromosome of M. thermoformicicum strains THF, CSM3 and HN4. Comparison of the nucleotide sequences of the two plasmid FR-II copies and that from the chromosome of strain CSM3 showed that the FR-II segments were approximately 2.5-3.0 kb in size and contained large open reading frames (ORFs) that may encode highly related proteins with an as yet unknown function.

Amino Acid Sequence↗
PubMed1993

IS870 requires a 5'-CTAG-3' target sequence to generate the stop codon for its large ORF1.

The TB regions of the Agrobacterium vitis octopine/cucumopine Ti plasmids constitute a family of related structures. All contain a bacterial insertion element downstream of the TB-iaaM gene, IS870.1. Whereas 43 isolates with octopine/cucumopine Ti plasmids carry only one IS870 copy, strain Ag57 carries a second copy (IS870.2) 3.9 kb to the right of IS870.1 and part of the same TB region. Two other octopine/cucumopine strains carry an IS870 copy on their chromosome (IS870.3). A study of the unmodified insertion sites of IS870.2 and IS870.3, cloned from closely related strains, enabled us to delimit the IS870 elements. IS870 has a size of 1,152 bp and is terminated by inverted repeats. It contains a large open reading frame without a stop codon. However, a stop codon is generated by insertion into the target sequence 5'-CTAG-3'. IS870 is related to five other insertion sequence elements. For two of these, the stop codon of the largest open reading frame is also created by insertion into a CTAG target site.

Amino Acid Sequence↗