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At least 1,261 records · Page 70Linked to original sources
PubMed1991

Structure, chromosomal localization and evolutionary conservation of the gene encoding human U1 snRNP-specific A protein.

Three specific proteins, called A, 70K and C, are present in the U1 small nuclear ribonucleoprotein (snRNP) particle, in addition to the common proteins. The human U1 snRNP-specific A protein is, apart from a proline-rich region, highly similar to the U2 snRNP-specific protein B". To examine the homologous regions at the genomic level, we isolated and characterized the human U1-A gene. The human U1-A protein appears to be encoded by a single-copy gene and its locus has been mapped to the q arm of chromosome 19. The gene, about 14-16 kb in length, consists of six exons. The regions homologous to the U2-B" gene are not limited to single exons and are mostly not confined by exon-exon junctions in the corresponding U1-A mRNA. However, the proline-rich region of U1-A, absent in U2-B", is encoded by a single exon, suggesting a specific function for this domain of U1-A. The region of the cap site and upstream sequences contain interesting similarities to the promoter region of other snRNP protein-encoding genes and several housekeeping genes, in particular the vertebrate ribosomal protein-encoding genes. Hybridization experiments with various vertebrate genomic DNAs revealed that U1-A sequences are evolutionarily conserved in all tested vertebrate genomes, except for chicken, duck and pigeon. The divergence of these avian genomes is probably typical for the class of birds.

Amino Acid Sequence↗
PubMed1991

Organization of the human hepatocyte growth factor-encoding gene.

Human genomic phage libraries were screened for the human hepatocyte growth factor (HGF)-encoding gene (HGF) using a cDNA encoding the human protein as a probe. Characterization of the clones revealed that this gene is composed of 18 exons interrupted by 17 introns spanning approx. 70 kb. The first exon contains the 5'-untranslated region and the signal peptide. The next ten exons encode the alpha-chain which contains four kringle structures. Each kringle domain is encoded by two exons as observed in other kringle-containing proteins. The twelfth exon contains the short spacer region between the alpha- and beta-chains and the remaining six exons comprise the beta-chain. The beta-chain is structurally similar to the catalytic domains of serine proteases; amino acid substitutions in the active site were found. The organization of the HGF gene is highly homologous to those of the serine proteases involved in blood coagulation and fibrinolysis, especially with that of plasminogen. This suggests that the human HGF gene is evolutionally related to these genes.

Amino Acid Sequence↗
PubMed1991

Sequence of the plasmid-encoded catechol 1,2-dioxygenase-expressing gene, pheB, of phenol-degrading Pseudomonas sp. strain EST1001.

Phenol monooxygenase (PMO) and catechol 1,2-dioxygenase (C12O), the two first enzymes of the phenol-degradation pathways, are encoded by a 3.4-kb DNA fragment cloned from Pseudomonas sp. EST1001 plasmid DNA. We have previously shown that activation of the cloned genes in Pseudomonas putida PaW85 is controlled by insertion of the 17-kb transposon, Tn4652, from the host chromosome into the plasmid carrying these genes [Kivisaar et al. Plasmid 24 (1990) 25-36]. Transcription of the DNA encoding PMO (pheA) and C12O (pheB) is activated by a promoter located on a 0.2-kb SacI-ClaI fragment from Tn4652. We have determined the nucleotide sequence of pheB. The 906-bp gene encodes a protein product with a deduced Mr of 33,362. The relationship between the pheB gene and other C12O-encoding genes has been shown: comparison of the pheB sequence with sequences of catA of Alcaligenes calcoaceticus, tfdC of A. eutrophus and clcA of P. putida demonstrated that there are conserved residues in all the four protein products of these genes.

Amino Acid Sequence↗
PubMed1991

Expression of the chicken 'delta 2-crystallin' gene in mouse cells: evidence for encoding of argininosuccinate lyase.

In the chicken genome, downstream from the delta 1 gene, encoding delta-crystallin, a highly homologous gene called delta 2 is found, but neither its expression nor its function has been clarified. Close similarity of the delta 2 gene to mammalian argininosuccinate lyase (ASL)-encoding genes in organization and deduced amino acid sequence of the encoded protein suggested that this gene may encode chicken ASL. To test this possibility, we cloned the delta 2 gene on a plasmid, introduced it into mouse cells, and assessed expression and ASL activity in comparison with delta 1. We found that delta 2 was expressed well in fibroblastic L cells, which resulted in synthesis of a protein with a size (50 kDa) and antigenicity very close to delta-crystallin, but it was repressed in lens cells in which expression of delta 1 was activated. Transfectant L cell lines expressing delta 2 exhibited significantly higher ASL activity than those without. We conclude that the delta 2 gene codes for chicken ASL, but has a tissue specificity distinct from delta 1 in its expression.

Animals↗
PubMed1991

De novo purine nucleotide biosynthesis: cloning, sequencing and expression of a chicken PurH cDNA encoding 5-aminoimidazole-4-carboxamide-ribonucleotide transformylase-IMP cyclohydrolase.

The purH cDNA, encoding 5-aminoimidazole-4-carboxamide-ribonucleotide (AICAR) transformylase-inosine monophosphate cyclohydrolase (ATIC), was cloned by functional complementation of an Escherichia coli purH mutant using a chicken liver cDNA expression library. This represents the first report of the cloning of any eukaryotic ATIC-encoding cDNA (PurH). The avian ATIC mRNA is 2.3 kb long and encodes a protein with an Mr of 64,422. The deduced amino acid sequence is 36% identical to the bacterial purH-encoded enzymes from Bacillus subtilis and E. coli. The avian cDNA was expressed as a glutathione S-transferase (GST) fusion protein that was purified in a single step by affinity chromatography. A novel vector was employed which permits rapid and highly efficient cleavage of the GST fusion protein yielding 10 mg of purified PurH product per liter of bacterial culture. Km values were determined with the purified fusion protein utilizing AICAR and (6-R)N10-formyl-tetrahydrofolate as substrates. These values compare favorably with the isolated avian enzyme, supporting the idea that kinetic, as well as other physical properties of the recombinant fusion protein are similar to the native avian enzyme. Large quantities of purified enzyme and the ability to generate site-directed mutations should make mechanistic studies possible. The recombinant enzyme also affords a simple and reliable approach to identifying new antifolates.

Acyltransferases↗
PubMed1991

Juxtaposition of the genes encoding Mycoplasma pneumoniae cytadherence-accessory proteins HMW1 and HMW3.

The loss and reacquisition of high-Mr (HMW) proteins, HMW1, 2, 3, 4 and 5, by Mycoplasma pneumoniae correlates with cytadherence phase variation. We are cloning and characterizing the genes encoding HMW1-5 to understand the mechanism regulating their coordinate expression. HMW1 was purified by polyacrylamide-gel electrophoresis. Amino acid (aa) sequence data were obtained from enzymatically generated peptide fragments from HMW1. A degenerate 17-mer probe synthesized based upon the aa sequence of one peptide clearly identified a single 4.75-kb BamHI fragment of M. pneumoniae DNA under stringent hybridization conditions. This fragment was cloned into pUC19 to generate pKV16. Restriction mapping of the 4.75-kb BamHI fragment in pKV16 revealed a possible overlap with the 9.4-kb EcoRI fragment containing the gene encoding protein HMW3. Southern blotting and reciprocal hybridization studies confirmed this overlap, establishing the juxtaposition of the genes encoding HMW1 and HMW3. Finally, physical mapping analysis by probing restriction fragments of M. pneumoniae DNA resolved by pulsed-field gel electrophoresis with the cloned genes encoding HMW1 and HMW3 revealed definitively that the hmw locus maps to a 106.8-kb ApaI fragment, rather than a 117.5-kb ApaI fragment, as had been reported previously for hmw3 [Krause and Mawn, J. Bacteriol. 172 (1990) 4790-4797].

Amino Acid Sequence↗
PubMed1991

Cloning and analysis of a cDNA encoding a human liver carboxylesterase.

A human liver carboxylesterase (CE)-encoding cDNA has been cloned using synthetic oligodeoxyribonucleotides (oligos) based on the known amino acid (aa) sequences of rabbit and rat liver CEs. The oligos hybridize specifically to DNA encoding liver CEs. The longest cDNA obtained from screening several cDNA libraries encodes about 80% of the protein and translates into an aa sequence which has a high degree of similarity with the sequences of liver CEs from other species. On hybridization to mRNA isolated from human liver, the cDNA gave a single band of about 2.0 kb consistent with its encoding a protein of less than 68 kDa. DNA obtained from a number of human livers and probed with the CE cDNA gave identical hybridization patterns. These patterns were moderately complex by comparison with published data.

Amino Acid Sequence↗
PubMed1991

Cloning, expression and characterization of a cDNA encoding a lipase from Rhizopus delemar.

A lambda gt11 cDNA library was constructed in Escherichia coli using poly(A)-selected mRNA from the fungus, Rhizopus (Rp.) delemar. Lipase-producing members of the library were identified by means of a phenotypic score wherein the release of fatty acids by lipase causes a characteristic color change in the growth medium. One such isolate contained a 1287-bp insert (LIP cDNA) which hybridizes to 1.25- to 1.35-kb mRNA species from Rp. delemar. The lipase produced in E. coli containing the LIP cDNA exhibits the same substrate selectivity as the authentic fungal enzyme, hydrolyzing ester bonds at the stereospecific numbering (sn) sn-1 and sn-3, but not the sn-2, positions of triglycerides. The complete nucleotide sequence of the LIP cDNA was determined. By reference to the N-terminal sequence of authentic Rp. delemar lipase, the lipase-encoding region was identified within this fragment. The LIP cDNA encodes a putative preprolipase consisting of a 26-amino-acid(aa) signal sequence, a 97-aa propeptide, and a 269-aa mature enzyme. The predicted mature lipase has the same molecular weight and aa composition as that of Rp. delemar, is highly homologous to that produced by the fungus Rhizomucor miehei, and contains the consensus pentapeptide (Gly-Xaa-Ser-Yaa-Gly) which is conserved among lipolytic enzymes. It is concluded that the LIP cDNA is an essentially full-length analogue of the lipase-encoding gene of Rp. delemar. The lipase encoded by the LIP cDNA occupies a cytoplasmic location when synthesized in E. coli. Unprocessed forms of the lipase accumulate in E. coli.

Amino Acid Sequence↗
PubMed1992

Evolution and differential expression of the (1-->3)-beta-glucan endohydrolase-encoding gene family in barley, Hordeum vulgare.

The (1-->3)-beta-D-glucan glucanohydrolases [(1-->3)-GGH; EC 3.2.1.39] of barley (Hordeum vulgare L., cv Clipper) are encoded by a small gene family. Amino acid sequences deduced from cDNA and genomic clones for six members of the family exhibit overall positional identities ranging from 44% to 78%. Specific DNA and oligodeoxyribonucleotide (oligo) probes have been used to demonstrate that the (1-->3)-GGH-encoding genes are differentially transcribed in young roots, young leaves and the aleurone of germinated grain. The high degree of sequence homology, coupled with characteristic patterns of codon usage and insertion of a single intron at a highly conserved position in the signal peptide region, indicate that the genes have shared a common evolutionary history. Similar structural features in genes encoding barley (1-->3,1-->4)-beta-glucan 4-glucanohydrolases [(1-->3,1-->4)-GGH; EC 3.2.1.73] further indicate that the (1-->3)-GGHs and (1-->3,1-->4)-GGHs are derived from a single 'super' gene family, in which genes encoding enzymes with related yet quite distinct substrate specificities have evolved, with an associated specialization of function. The (1-->3,1-->4)-GGHs mediate in plant cell wall metabolism through their ability to hydrolyse the (1-->3,1-->4)-beta-glucans that are the major constituents in barley walls, while the (1-->3)-GGHs, which are unable to degrade the plant (1-->3,1-->4)-beta-glucans, can hydrolyse the (1-->3)- and (1-->3,1-->6)-beta-glucans of fungal cell walls.

Amino Acid Sequence↗
PubMed1992

Cloning, expression and tissue distribution of the gene encoding rat fibroblast growth factor receptor subtype 4.

The rat homologue of the gene encoding the fibroblast growth factor receptor subtype 4 (FGFR4) was cloned from rat lung mRNA, and the cDNA sequence was found to be 95% similar and 92% identical to the human homologue. Northern blot analysis of adult rat tissues demonstrated that a 3.1-kb mRNA encoding FGFR4 is detectable only in the lung and kidney. The receptor variant described here encodes two potential immunoglobulin-like domains, 21 hydrophobic amino acids encoding a potential transmembrane domain, and a split tyrosine kinase motif. However, the acidic box and hydrophobic signal peptide domains are not present in this cDNA isolate.

Amino Acid Sequence↗
PubMed1992

Transcriptional regulation of the Tn5276-located Lactococcus lactis sucrose operon and characterization of the sacA gene encoding sucrose-6-phosphate hydrolase.

The Lactococcus lactis sucrose operon was located on the conjugative transposon Tn5276 and the nucleotide sequence of the sacA gene, encoding sucrose-6-phosphate hydrolase, and its surrounding regions was determined. Northern blot analysis showed that the sucrose operon contains two divergent transcriptional units of 3.2 and 3.6 kb, the expression of which is considerably higher in cells grown on sucrose than in cells grown on glucose. This was confirmed by primer extension studies which demonstrated that transcription is initiated at two sucrose-inducible promoters with a back-to-back organization. The 3.2-kb transcriptional unit includes the sacB gene which most probably encodes the sucrose-specific enzyme II of the phosphotransferase system, and may contain the gene encoding fructokinase. The 3.6-kb transcriptional unit includes genes sacA and sacR. The protein encoded by the sacR gene is likely to be involved in the regulation of the sac operon expression, since its deduced N terminus is homologous to helix-turn-helix DNA-binding domains found in several regulatory proteins.

Amino Acid Sequence↗
PubMed1992

Characterization of a highly expressed lignin peroxidase-encoding gene from the basidiomycete Phanerochaete chrysosporium.

The genomic clone, LG2, encoding LiP2, the major lignin peroxidase (LiP) isozyme from Phanerochaete chrysosporium strain OGC101, was isolated and characterized. The 5'-untranslated region of LG2 contains sequences similar to CRE and XRE promoter elements. Comparison with its transcript indicates that eight introns, each less than 59 bp, interrupt the coding sequence. Comparison with genes encoding other LiP isozymes shows five related patterns of intron location, whose incidence coincides with described LiP structural subfamilies. Codon bias indices calculated for all known P. chrysosporium genes, including trpC and genes encoding LiP, MnP, and exo-cellobiohydrolase I, demonstrate that LG2 has the most biased codon usage. We conclude that subdivisions of the LiP family may be based on intron location in the encoding genes, and that ranking of isozyme production levels can be estimated by the extent of bias in codon usage in the cognate gene.

Amino Acid Sequence↗
PubMed1992

The structure and complete sequence of the gene encoding chicken ribosomal protein L5.

The nucleotide (nt) sequence of the gene encoding chicken ribosomal protein L5, which associates with 5S rRNA, was determined. The gene contains eight exons and seven introns which spread over 7252 bp. The transcription start point (tsp) is a C residue in a tract of 13 pyrimidines, and its 5'-flanking region lacks canonical TATA and CAAT boxes. The G+C content of the region around the tsp is calculated as high as 78%, and nine GC boxes exist within the 5'-flanking region and the first intron. The nt sequence at positions -36 to -22 is similar to an internal sequence at positions 56-70 of the gene encoding chicken 5S rRNA, which corresponds to the transcriptional internal control region of the 5S rRNA-encoding gene of Xenopus laevis. This region might contribute to the coordinate expression of the genes encoding protein L5 and 5S rRNA.

Amino Acid Sequence↗
PubMed1992

Characterization of the gene encoding the most abundant in vitro translation product from virus-infected Chlorella-like algae.

The gene (33kDa) encoding a 33-kDa peptide from Chlorella virus, PBCV-1, was cloned and sequenced. This gene encodes the most abundant in vitro translation product synthesized from viral mRNAs isolated beginning at 20 min post-infection. The message persisted throughout the remainder of the viral life cycle. An open reading frame (ORF) of 717 bp, which encodes a polypeptide of 238 amino acids with a predicted M(r) or 26,613, was found on a 2752-bp cloned fragment from PBCV-1 HindIII restriction fragment 9. Transcriptional analysis of this ORF indicated that it was expressed both early and late, and as the viral life cycle progressed, the mRNA increased in size and abundance. Three other ORFs were also found; the largest of which (741 bp) hybridized to a low-abundance transcript which would encode a polypeptide with a predicted M(r) of 27,854.

Amino Acid Sequence↗
PubMed1992

Isolation and expression of a murine purine nucleoside phosphorylase-encoding cDNA and sequence similarity with the human message.

To isolate murine purine nucleoside phosphorylase-encoding cDNA sequences (PNP), a murine BALB/c liver cDNA library in lambda gt10 was screened for recombinants hybridizing to a human PNP cDNA probe. Two of three clones recovered included inserts large enough to contain the full-length coding sequence. Sequence analysis of the largest clone revealed an 867-nucleotide open reading frame encoding 289 amino acids with 84% residue identity to that encoded by human PNP and 351 bp of 3'-untranslated region. The 5' end of the murine PNP message was specifically amplified by PCR using the RACE (rapid amplification of cDNA ends) protocol, revealing a 5'-untranslated region of 78 bp. Northern hybridization using the murine PNP cDNA sequence as a probe identified a message of approx. 1.6 kb in mouse NIH3T3 cells which was slightly smaller than the human message observed in HeLa cells. The cloned murine PNP cDNA coding sequence was inserted into a mammalian expression vector under transcriptional regulation of the Moloney murine leukemia virus long terminal repeat. Transfection of this plasmid into human 293 cells resulted in the expression of PNP activity which co-focused with murine PNP activity extracted from NIH3T3 cells, verifying that the isolated murine PNP cDNA clone encoded catalytically active PNP protein.

3T3 Cells↗
PubMed1992

Expression and genomic structure of the genes encoding FdI, the major allergen from the domestic cat.

The genes encoding chain 1 (Ch1) and chain 2 (Ch2) of the major allergen of the domestic cat, Felis domesticus I, have been analyzed by genomic cloning and by polymerase chain reaction (PCR). Ch1 has two potential leader sequences, designated A and B. Analysis of a genomic clone encoding Ch1 demonstrated that one structural gene contains sequences corresponding to both leaders, which utilize different Met start codons. PCR analysis showed that genes encoding Ch1 and Ch2 are co-expressed in both the salivary glands and the skin, and that leader sequence A of Ch1 is utilized preferentially in both tissues. Ch2 was shown to have two dominant forms that are differentially expressed in the aforementioned tissues. The long form (Ch2L), composed of 92 amino acids (aa), is preferentially expressed in the salivary glands, while the short form (Ch2S), composed of 90 aa, is preferentially expressed in the skin. There is minor sequence polymorphism in both forms of Ch2. A genomic clone for Ch2 only contained sequences for Ch2S, suggesting that Ch2L is encoded by an exon not contained within this genomic clone.

Allergens↗
PubMed1992

Structural organization of the gene encoding rat cystatin beta.

A genomic DNA clone encompassing the gene (cy beta) encoding rat cystatin beta (Cy beta) was isolated by screening with a rat cy beta cDNA as a probe. The gene spans about 2.6 kb and comprises three exons. The first intron is located between Lys22 and Val23 and the second between Lys56 and Val57 in the deduced amino acid sequence of Cy beta. The second exon contains the highly conserved QVVAG sequence which, unlike the sequence of other cystatin family members, is not split by an intron. In the 5'-upstream region, three SP-1-binding sites exist, but no typical TATA-box or CAAT-box sequences are found. The difference in the organization of the rat cy beta gene, encoding a family-1 cystatin, from that encoding members of the other cystatin families, suggests that cy beta diverged from a common ancestral gene earlier than the separation of genes encoding family-2 and family-3 cystatins.

Amino Acid Sequence↗
PubMed1993

The complete sequence of the gene encoding bovine alpha s2-casein.

From a bovine genomic library, five overlapping clones, spanning some 50 kb, have been isolated. These clones contain the complete alpha s2-casein-encoding gene (alpha s2ca) and its 5' and 3' flanking regions. The nucleotide (nt) sequence of the complete gene including 2510 bp of the 5' flanking region and 276 bp of the 3' region has been determined. The total length of alpha s2ca appears to be 18483 bp and, therefore, it is the longest of the four bovine casein-encoding genes. The alpha s2ca gene is comprised of 18 exons ranging in size from 21 to 266 nt. There are 16 Alu-like artiodactyla retroposons inserted at ten different locations within the gene. About 14% of the gene is composed of these repetitive sequences. Although the organization of alpha s2ca appears to be similar to that of the alpha s1-casein-encoding gene (alpha s1ca), sequence comparisons and the length of the exons indicate that it is more closely related to the beta-casein-encoding gene. Furthermore, it is shown that both genes could have evolved from a common ancestor by means of internal duplications.

Animals↗