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Query: UNIPROT:P06889 (Mol)
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In the silkworm, Bombyx mori, a group of structurally related proteins, termed 30K proteins, accumulate in the hemolymph of the last instar larvae. We have isolated and characterized three genes, each of which encodes a distinct 30K protein component. Each 30K protein gene is composed of a short first exon and a protein-coding second exon interspersed by a single intron. The transcription initiation site of the 30K protein mRNA was identified at the nucleotide level. A typical TATA box exists some 30 base-pairs upstream from the transcription initiation site. The 5'-flanking region of each gene also contains octamer-like sequences. Restriction mapping analyses revealed that the cloned 46 x 10(3) base-pair region of the chromosomal DNA bears three 30K protein genes. Several copies of highly reiterated retrotransposon-like sequences are present around the 30K protein genes. S1 nuclease protection analysis provided evidence that the biosynthesis of 30K protein is regulated in a stage-specific manner at the transcriptional level in the fat body.
J Mol Biol 1991 Mar 05
PMID:Structures and organization of major plasma protein genes of the silkworm Bombyx mori. 200 9

A full length (25,000 base-pair) myosin heavy chain gene completely contained within a single cosmid clone was isolated from a Syrian hamster cosmid genomic library. Sequence comparison of the 3' untranslated region indicated the presence of a 75% homology with the rat embryonic myosin heavy chain gene. Extensive 5' flanking region regulatory element conservation was also found when the sequence was compared to the rat myosin heavy chain gene. S1 nuclease digestion analysis, however, indicated that the Syrian hamster myosin heavy chain gene exhibited expression in adult Syrian hamster ventricular tissue, as well as the adult vastus medialis, a fast twitch skeletal muscle. Expression also appears to be enhanced in myopathic relative to control hearts. This myosin heavy chain gene is neither the alpha nor beta cardiac myosin heavy chain gene, but is a unique, previously unrecognized, myosin heavy chain gene present in both myocardial and skeletal muscle tissues.
J Mol Biol 1991 Apr 20
PMID:Isolation and characterization of a previously unrecognized myosin heavy chain gene present in the Syrian hamster. 202 40

An open reading frame (ORF) was found upstream of the mdh gene in Thermus flavus by computer-aided analysis. It was identified as the gene encoding the alpha subunit of succinyl-CoA synthetase (SCS) and termed scsA. Nucleotide sequencing of a further upstream region revealed the presence of another ORF, corresponding to the sequence of the beta subunit of SCS. The latter gene was termed scsB. The scsB gene was found to contain an unusual translational initiation codon, TTG. S1 nuclease mapping indicates that transcription starts at the nucleotide at position--31 upstream of the initiation codon of the beta gene. The scsB and scsA genes along with the mdh gene appear to form an operon and are most likely co-transcribed in this order, because the intercistronic regions between them are very short; in fact, the termination codon of scsB overlaps the initiation codon of scsA. A stretch characteristic of the--10 region of a typical prokaryotic promoter was found upstream of scsB, whereas no sequence characteristic of a typical--35 region was present. Escherichia coli harboring a plasmid containing scsA and scsB did not produce thermostable SCS activity, even when a synthetic promoter for E. coli was attached. However, when an inverted repeat present in front of scsB, which covers the putative ribosome-binding site and is capable of forming a stable stem-loop structure, was altered by site-directed mutagenesis, overproduction of heat-stable SCS was observed.
Mol Gen Genet 1991 Apr
PMID:Characterization of an operon encoding succinyl-CoA synthetase and malate dehydrogenase from Thermus flavus AT-62 and its expression in Escherichia coli. 203 8

We have determined the complete nucleotide sequence of the linear DNA plasmid, pSKL, isolated from Saccharomyces kluyveri. Sequence analysis showed that pSKL has a high (A + T) content of 71.7%, and that there are 10 open reading frames (ORFs) larger than 250 nucleotides. All 10 ORFs were shown to be transcribed in S. kluyveri cells by S1 nuclease mapping analysis. The localization of ORFs, direction of transcription, and the predicted amino acid sequences of each ORF were quite similar to that of pGKL2, one of the killer plasmids found in Kluyveromyces lactis. The amino acid sequences of the largest two ORFs (ORF2 and ORF6) have homology with several DNA polymerases and RNA polymerases, respectively.
Mol Gen Genet 1991 Apr
PMID:Genome organization of the linear plasmid, pSKL, isolated from Saccharomyces kluyveri. 203 32

To better understand the regulation of interleukin-7 receptor (IL-7R) expression, we have pursued a detailed analysis of the structure of the murine and human IL-7R genes. The genes consist of eight exons, the sizes of which are conserved in mouse and human cells, spread out over 24 kbp (murine) and 19 kbp (human). A differential splicing event results in an mRNA encoding a secreted form of the human IL-7R gene. Primer extension and S1 nuclease analysis show a single transcriptional start site for the murine IL-7R gene. The 5'-flanking region of the murine IL-7R gene contains TATA- and CAAT-like sequences. The promoter region also contains a functional interferon regulatory element, to which the interferon-induced nuclear factors IRF-1 and IRF-2 are capable of binding and which is able to confer interferon-inducible expression on a heterologous gene. There are also potential binding sites for the transcription factors AP-1 and AP-2 as well as multiple glucocorticoid response elements. A fusion gene containing 2.5 kb of murine IL-7R 5' regulatory sequence linked to the bacterial chloramphenicol acetyltransferase gene directed expression of chloramphenicol acetyltransferase activity in murine pre-B-cell line 70Z/3 but not in the mouse fibroblast cell line NIH 3T3. Comparison of the murine and human IL-7R exon/intron boundaries with those of other hematopoietin receptor superfamily members whose exon/intron boundaries are also known reveals a conserved evolutionary structure.
Mol Cell Biol 1991 Jun
PMID:Organization of the murine and human interleukin-7 receptor genes: two mRNAs generated by differential splicing and presence of a type I-interferon-inducible promoter. 203 16

We have analyzed 572 bp in the 28S rDNA of the human blood fluke Schistosoma mansoni which correspond to expansion segment 5 of domain IV as defined by Clark et al. for the Xenopus laevis 28S rRNA. S1 nuclease mapping and primer extension analysis comparing this region with the mature 28S rRNA indicate that there are 54 nucleotides present in the 28S rDNA which are absent from the mature rRNA. This defines a gap that creates two 28S rRNA subunits (28S alpha and 28S beta). Comparison of the S. mansoni sequence with rDNAs of other organisms which contain gaps in their 28S rRNA shows that the overall features are conserved except that the S. mansoni gap is less A + T-rich. The conserved features include: (1) the location of the gap within the 28S rRNA; (2) the predicted secondary structure of the gap, containing a stem-loop with a UAAU sequence within the loop; and (3) a conserved CGAAAGGG on the 3' side of the gap.
Mol Biochem Parasitol 1991 Apr
PMID:Characterization of a 54-nucleotide gap region in the 28S rRNA gene of Schistosoma mansoni. 203 56

Maximum expression of the Corynebacterium glutamicum lysA gene is dependent upon the presence of a 2.3 kb region immediately 5' of the lysA reading frame. Subcloning and functional analysis of the upstream region implied that this region contained the lysA promoter. Sequence determination of the upstream region revealed a single open reading frame, orfX, in the same orientation as lysA. The orfX coding sequence exhibited all the sequence characteristics of a gene with the potential for a 550-amino-acid polypeptide product. Expression of lysA is coupled to that of orfX via a common promoter located immediately 5' of orfX. The RNA start site has been determined by S1 nuclease mapping. Both the orfX and the lysA gene are expressed as a single 3.0 kb RNA transcript. These data indicate that orfX and lysA are genes within a two-gene operon. Expression of the lysA gene is not subject to regulation by lysine. The orfX gene product was shown not to be directly linked to the lysine biosynthetic pathway, nor is it the enzyme incorporating DAP into the peptidoglycan precursor.
Mol Microbiol 1990 Nov
PMID:Nucleotide sequence and organization of the upstream region of the Corynebacterium glutamicum lysA gene. 849 94

The structural gene encoding human 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase (3 beta HSD) was isolated from a human EMBL3 genomic library. The gene encompasses approximately 8 kilobases of DNA and is comprised of two large introns and three exons encoding amino acid residues 1-48, 49-103, and 104-373, respectively. The exonic sequence is identical to that of the cDNA that we previously isolated and expressed in COS 1 cells. DNA sequence analysis reveals a putative TATA (TATATAA) motif 26 basepairs up-stream of the beginning of exon I, as determined by S1 nuclease protection analysis. However, primer extension analysis using poly(A)+ RNA isolated from both placenta and corpora lutea indicates that the RNA initiates up-stream of the putative TATA motif, and that an additional 53-basepair exon, which is untranslated, is present 5' to the first coding exon. Southern hybridization analysis of genomic DNA using a single exon probe suggests that there may be more than one copy of the gene in the human genome. In addition, we confirm from Southern analysis of genomic DNA isolated from human x hamster somatic cell hybrids that the gene is located on human chromosome 1. These findings will provide a foundation for the characterization of apparent 3 beta HSD clinical deficiencies when these are due to a mutation in the structural gene.
Mol Endocrinol 1990 Dec
PMID:Structural analysis of the gene encoding human 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase. 208 86

The pea plastocyanin gene in a 3.5 kbp Eco RI fragment of pea nuclear DNA was introduced into tobacco by Agrobacterium-mediated transformation. Regenerated plants contained pea plastocyanin located within the chloroplast thylakoid membrane system. Analysis of seedlings from a self-pollinated transgenic plant containing a single copy of the pea plastocyanin gene indicated that seedlings homozygous for the pea gene contained almost twice as much pea plastocyanin as seedlings hemizygous for the pea gene. Homozygous seedlings contained approximately equal amounts of pea and tobacco plastocyanins. The amount of tobacco plastocyanin in leaves of transgenic plants was unaffected by the expression of the pea plastocyanin gene. The mRNA from the pea gene in tobacco was indistinguishable by northern blotting and S1 nuclease protection from the mRNA found in pea. In both pea and transgenic tobacco, expression of the pea plastocyanin gene was induced by light in leaves but was suppressed in roots. Pea plastocyanin free of contaminating tobacco plastocyanin was purified from transgenic tobacco plants and shown to be indistinguishable from natural pea plastocyanin by N-terminal protein sequencing and 1H NMR spectroscopy.
Plant Mol Biol 1990 Feb
PMID:Synthesis and accumulation of pea plastocyanin in transgenic tobacco plants. 210 92

A 1.8 kb transcript corresponding to a region of the Anabaena 7120 chromosome 4 kb downstream of the nifHDK operon appears 12-18 h after heterocyst induction. The DNA corresponding to this transcript was sequenced and found to contain two open reading frames, designated ORF 1 and ORF 2. Two polypeptides, of 30 kDa and 13 kDa, encoded by these ORFs were expressed in Escherichia coli. An apparent start site for the transcript, detected by S1 nuclease protection, was located 42 bp upstream of the ATG start codon of ORF 1. ORF 2 shows strong sequence similarity to ORF 6 in the nif gene region of Azotobacter vinelandii. ORF 1 was interrupted using a 1.4 kb neomycin resistance cassette and the resulting mutant grew very slowly on medium lacking combined nitrogen. The mutant had 45% of wild-type acetylene reduction activity, which could be complemented by a 2.8 kb EcoRI fragment of wild-type Anabaena DNA containing only ORF 1 and ORF 2. Thus, one or both of these ORFs is required for efficient nitrogen fixation in Anabaena.
Mol Gen Genet 1990 Apr
PMID:Expression, nucleotide sequence and mutational analysis of two open reading frames in the nif gene region of Anabaena sp. strain PCC7120. 211 11


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