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At least 145 records · Page 8Linked to original sources

PubMed1990

Characterization of multiple mRNAs originating from the chicken progesterone receptor gene. Evidence for a specific transcript encoding form A.

The structure of the 42-kilobase (kb) long chicken progesterone receptor (cPR) gene and of all six transcripts that are detectable on Northern blots was determined. The first of 8 exons encodes the N-terminal region A/B which is highly divergent among different species and contains a constitutive transcription activation function. The DNA (DBD)- and hormone-binding domains (HBD) are assembled from 2 and 5 exons, respectively, with the individual "zinc fingers" of the DBD encoded by separate exons. In addition to the previously described 4.5-kb cPR mRNA species, alternative polyadenylation, splicing variation, and "5'-truncation" lead to the generation of 5 further mRNAs. Most importantly, this 5'-truncation produces, by an as yet unidentified mechanism, an abundant transcript which encodes form A but not form B of cPR. Lack of splicing at the exon 2 splice-donor and polyadenylation due to a signal site in the second intron generates a previously undetected 3.4-kb mRNA species. The corresponding cDNA was sequenced in its entirety and shown to encode only region A/B and the N-terminal "finger" of the DBD. Alternative polyadenylation upstream of the signal site for the 4.5-kb mRNA is responsible for the appearance of a 3.3-kb mRNA. The longest cPR mRNA (8.2 kb) originates from a transcription termination point more than 3 kb downstream of the 4.5-kb mRNA 3'-end. Finally, the primary sequence of more than 2 kb upstream sequences of the cPR gene, containing several consensus hexamer progestin/glucocorticoid receptor-binding sites (PRE/GRE and putative Sp1 binding motifs, is discussed.

Amino Acid Sequence↗

PubMed1994

DNA recognition by intercalators and hybrid molecules.

Experiments are described which probe the role of the 2-amino group of guanine as a critical determinant of the recognition of nucleotide sequences in DNA by specific ligands. Homologous samples of tyrT DNA substituted with inosine or 2,6-diaminopurine residues in place of guanosine or adenine respectively yield characteristically modified footprinting patterns when challenged with sequence-selective antibiotics such as echinomycin, actinomycin or netropsin. The capacity of small molecules to recognise particular DNA sequences is exploited in the 'combilexin' strategy to target small molecules to defined sites in DNA. A composite molecule containing a distamycin moiety linked to an intercalating ellipticine derivative has been synthesised and shown to bind tightly to DNA but without much sequence-selectivity. Refinement of this molecule based on predictions from molecular modelling has led to the synthesis of a second generation derivative bearing an additional positive charge: this new hybrid molecule is strongly selective for binding to AT-rich tracts in DNA.

Antineoplastic Agents↗

PubMed1993

A human heart cDNA library--the development of an efficient and simple method for automated DNA sequencing.

A direct polymerase chain reaction (PCR) amplification of the human heart cDNA clones was used to generate lambda PCR product. By the use of the first set of primers derived from lambda gt11, each cDNA insert can readily be obtained. Using millipore filters, primers and nucleotides are removed and this purified PCR product can then be subjected to a second set of fluorescent primers in the generation of nucleotides in the auto-cycle reactions for automated DNA sequencing.

Base Sequence↗

PubMed1995

Recombinant vesicular stomatitis viruses from DNA.

We assembled a DNA clone containing the 11,161-nt sequence of the prototype rhabdovirus, vesicular stomatitis virus (VSV), such that it could be transcribed by the bacteriophage T7 RNA polymerase to yield a full-length positive-strand RNA complementary to the VSV genome. Expression of this RNA in cells also expressing the VSV nucleocapsid protein and the two VSV polymerase subunits resulted in production of VSV with the growth characteristics of wild-type VSV. Recovery of virus from DNA was verified by (i) the presence of two genetic tags generating restriction sites in DNA derived from the genome, (ii) direct sequencing of the genomic RNA of the recovered virus, and (iii) production of a VSV recombinant in which the glycoprotein was derived from a second serotype. The ability to generate VSV from DNA opens numerous possibilities for the genetic analysis of VSV replication. In addition, because VSV can be grown to very high titers and in large quantities with relative ease, it may be possible to genetically engineer recombinant VSVs displaying foreign antigens. Such modified viruses could be useful as vaccines conferring protection against other viruses.

Animals↗

PubMed1994

Molecular cloning of HCV and clinical application.

Fifty-five clones encoding epitopes of HCV were isolated from Japanese patients. Their amino acid homology (AAH) to the sequence of prototype (HCV-1) ranged from 47% to 94%. These sequences cover 60% of the HCV genome lacking M/E and NS2 regions suggesting a very low or lacking immunogenicity for these regions. Two test kits for detection of anti-HCV antibody were developed using a combination of a synthetic peptide (AR142) containing the epitope of N14 (QRKTKRSTNRR) having a homology to the core of HCV of 8/11AA and a non-fusion recombinant protein Y19 starting from amino acid number (AAN) 1380 to 1507 in the NS3 region showing a AAH to the HCV-1 of 90%, and a combination of a mixture of three synthetic peptides of S29 AAN of 1-30, 38-65 and 47-74 of the core and a non-fused recombinant protein S4 AAN of 1287-1506 having a 93% AAH of the NS3 region. They showed almost the same order of sensitivity and specificity of the second-generation kits when tested with serum from blood donors and patients with non-A, non-B hepatitis. It should also be stressed that in all of the complete responders of a recombinant alpha-interferon therapy, the antibody levels against AR142 gradually decreased during and after the treatment. In 1992, studies performed for 125 patients with hepatocellular carcinoma in our clinic shows that of these 16 patients might developed from either chronic non-B, non-C liver diseases or chronic liver diseases caused by mutant(s) of HCV as their serum were negative for HBsAg and second-generation of anti-HCV.

Adolescent↗

PubMed1996

Antisense oligonucleotides: towards clinical trials.

Antisense oligonucleotides have the ability to selectively block disease-causing genes, thereby inhibiting production of disease-associated proteins. The specificity and application of antisense oligonucleotides have been strongly validated in animal models for various disease targets. Based on the pharmacological, pharmacodynamic and pharmacokinetic profiles, the first generation of antisense oligonucleotides--phosphorothioates--have reached the stage of human clinical trials for various diseases. While ongoing human clinical trials are being carried out to further establishing the safety and efficacy of these oligonucleotides, the experience gained is providing a basis for designing a second generation of antisense oligonucleotides.

Administration, Oral↗

PubMed1981

Primary structure of the Fc region of human immunoglobulin D: implications for evolutionary origin and biological function.

We have determined the complete amino acid sequence of a tryptic Fc delta fragment generated from an intact human IgD (WAH); it is 226 residues long and includes the second (C delta 2) and the third (C delta 3) constant domains of the delta chain. Comparison of the homology of the Fc sequence of the five human immunoglobulin classes suggests that either the delta-chain gene evolved from the alpha-chain gene soon after the divergence of a mu-alpha common ancestor or it evolved from an ancestral gene distinct from both the mu-alpha and the gamma-epsilon common ancestors. Comparative study using a spatial model of the Fc region indicates that the structure of the C delta 3 domain differs extensively from that of the carboxy-terminal domains of other heavy chain classes; this, together with the unique hinge region structure, probably reflects the biological role of IgD as a receptor molecule on the B-lymphocyte surface.

Amino Acid Sequence↗

PubMed1986

Programs to produce high quality dichotic tapes for central auditory testing.

Dichotic stimulation, the simultaneous presentation of two different acoustic signals to the right and left ears, respectively, is used routinely in the clinical assessment of speech lateralization as well as in other central auditory testing procedures in the clinic and laboratory. At present, most researchers and clinicians depend on a few commercial sources for dichotic tapes, because there are a limited number of facilities which are able to produce such tapes. Moreover, some of the commercial tapes currently offered for sale contain important stimulus errors. In the present paper, we describe a general set of programs for the PDP 11 series of computers, which permits sequences of dichotic stimuli to be generated and output with a high degree of precision. These programs therefore allow researchers to generate any desired sequence of dichotic stimulation, for recording and taping, for use in their research. The programs operate in two stages to generate the required dichotic sequences. In the first stage, the constrained random ordering of the stimuli is generated as specified by the user. In the second stage, after the preliminary stimulus preparation has been completed (for example, using a waveform editing package; cf., D.G. Jamieson and D. A. Naugler, Comput. Biomed. Res., 1985, 18, 480), an audio tape is generated with stimuli presented dichotically, with timing and sequencing precisely as specified.

Auditory Cortex↗

PubMed1984

The light-harvesting polypeptides of Rhodospirillum rubrum. I. The amino-acid sequence of the second light-harvestng polypeptide B 880-beta (B 870-beta) of Rhodospirillum rubrum S 1 and the carotenoidless mutant G-9+. carotenoidless mutant G-9+.

The light-harvesting complex B 880 from Rhodospirillum rubrum S 1 (wild type) and B 870 from the carotenoidless mutant G-9+ was shown to consist mainly of an organic solvent-(chloroform/methanol-) soluble and an organic solvent-insoluble polypeptide. The isolation and separation of these two low-molecular-mass polypeptides (Mr 6101 and Mr 6079) were achieved by a two-step extraction procedure of chromatophores using in the first step chloroform/methanol containing 0.1M ammonium acetate. Following Sephadex LH-60 chromatography of this first extract a light-harvesting polypeptide (B 870-alpha) was isolated and its complete amino acid sequence was determined (R. Brunisholz et al. (1981) FEBS Lett. 129/1, 150-154, B 880-alpha: G. Gogel et al. (1983) Biochim. Biophys. Acta 746, 32-39). Upon reextraction of the chromatophore pellet with chloroform/methanol/ammonium acetate containing in addition acetic acid a second low-molecular-mass polypeptide (B 880-beta of B 870-beta) was generated. The complete amino acid sequences of the chloroform/methanol-insoluble light-harvesting polypeptide of Rs. rubrum S 1 (B 880-beta) and of Rs. rubrum G-9+ (B 870-beta) were determined. They are identical and consist of 54 amino acid residues. The conserved histidine residue within the hydrophobic stretch raises more evidence for ligand complexation of bacteriochlorophyll to this specific histidine residue which therefore possibly plays the key role in pigment-protein interactions. Both polypeptides (B 880-alpha and B 880-beta) are part of the light-harvesting complex B 880 in an apparent ratio of 1:1. Based on the primary structure data a possible arrangement of both light-harvesting polypeptides within the membrane will be discussed.

Amino Acid Sequence↗

PubMed1995

Determination of cyclic nucleotide-dependent protein kinase substrate specificity by the use of peptide libraries on cellulose paper.

An iterative approach to the a priori determination of the substrate specificity of cAMP- and cGMP-dependent protein kinases (PKA and PKG) by the use of peptide libraries on cellulose paper is described. The starting point of the investigation was an octamer library with the general structure Ac-XXX12XXX, where X represents mixtures of all 20 natural amino acids and 1 and 2 represent individual amino acid residues. The library thus contained all possible 2.56 x 10(10) octamers, divided into 400 sublibraries with defined amino acids 1 and 2 each consisting of 6.4 x 10(7) sequences. After phosphorylation with the kinases in the presence of [gamma-32P]ATP, the sublibrarys Ac-XXXRRXXX and Ac-XXXRKXXX were identified as the best substrates for PKA and PKG, respectively. The second-generation libraries had the structures Ac-XXXRR12X and Ac-XXXRK12X for PKA and PKG and resulted in the most active sequence pools Ac-XXXRRASX and Ac-XXXRKKSX. After delineation of every position in the octameric sequence and extension of the investigation to decameric peptides, the best sequences, Ac-KRAERKASIY and Ac-TQKARKKSNA, were obtained for PKA and PKG, respectively. Promising octameric and decameric peptides were assembled 5 or 10 times each and assayed in order to determine the experimental scatter inherent in the approach. The kinetic data of several octameric and decameric sequences were determined in solution and compared to data for known substrates. The recognition motif of PKA was confirmed by this approach, and a novel substrate sequence for PKG was identified.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗

PubMed2026

Employing Metagenomics Capture targeted next-generation sequencing for the etiological diagnosis of bloodstream infections.

BACKGROUND: Bloodstream infections (BSIs) represent a significant public health concern. Metagenomic Capture targeted next-generation sequencing technology, as a newly emerging method for pathogen detection, has been applied in the etiological diagnosis of various infectious diseases and demonstrates good diagnostic efficacy. However, there is relatively limited research on the diagnostic value of this technology for the etiological diagnosis of BSIs. METHODS: A comprehensive retrospective analysis was performed on patients suspected of having BSIs who were admitted to the Affiliated Guangdong Second Provincial General Hospital of Jinan University in 2024. These patients underwent both blood culture analysis and Metagenomic Capture targeted next-generation sequencing technology for diagnostic testing, and a detailed comparison of the results was conducted. RESULTS: It was found that the Metagenomic Capture-targeted next-generation sequencing method has a shorter time to result [1.33 (1.18 - 1.69) vs 2.73 (1.89 - 3.84) days, p < 0.001], more pathogenic microbial species detected, higher positive detection rate and higher sensitivity than blood culture. CONCLUSIONS: Metagenomic Capture targeted next-generation sequencing technology is a promising tool for pathogen identification in BSIs, offering substantial methodological advantages in terms of turnaround time, detection breadth, and sensitivity. These diagnostic performance characteristics support its potential utility in clinical microbiology practice.

Humans↗

PubMed1989

Computer-controlled sequencing of peptides by tandem mass spectrometry.

A fully automated computer-controlled system was used to generate series of different linked scans at constant B2E and constant neutral loss in the second field-free region. This system has been shown to be suitable for deriving the amino acid sequence of oligopeptides.

Amino Acid Sequence↗

PubMed1995

Detection of hepatitis C virus antibodies and specific hepatitis C virus ribonucleic acid sequences in cord bloods from a heterogeneous prenatal population.

OBJECTIVE: Our aim was to quantify the prevalence of at-risk pregnancies for maternal-fetal hepatitis C virus transmission in a heterogeneous prenatal population by detection of both hepatitis C virus-specific antibody and hepatitis C virus ribonucleic acid sequences in cord bloods from their deliveries. STUDY DESIGN: An anonymous serosurvey of 1432 consecutive umbilical cord blood samples were analyzed for hepatitis C virus antibodies with a second-generation enzyme immunoassay with all hepatitis C virus antibody-positive samples batched and analyzed for both human immunodeficiency virus antibodies and hepatitis C virus ribonucleic acid sequences by polymerase chain reaction. RESULTS: Forty-seven of the samples (3.2%) were positive for hepatitis C virus antibodies; seropositivity rates differed significantly by socioeconomic status but not by race. Significantly more of the antibody-positive women underwent cesarean section for delivery (31.9% vs 21.9%, p = 0.03). Three (6.4%) hepatitis C virus antibody-positive samples were also human immunodeficiency virus-antibody positive, whereas nine (19.1%) were hepatitis C virus ribonucleic acid positive. CONCLUSION: As many as 19% of hepatitis C virus antibody-positive women in this study also had hepatitis C virus ribonucleic acid isolated from their delivery cord blood samples, which may indicate an increased risk of vertical hepatitis C virus transmission in those pregnancies. Hepatitis C virus-specific antibody and ribonucleic acid detection may also be markers for other pregnancy complications that result in higher rates of cesarean section for these women.

Base Sequence↗

PubMed1984

Repeats of base oligomers as the primordial coding sequences of the primeval earth and their vestiges in modern genes.

Three outstanding properties uniquely qualify repeats of base oligomers as the primordial coding sequences of all polypeptide chains. First, when compared with randomly generated base sequences in general, they are more likely to have long open reading frames. Second, periodical polypeptide chains specified by such repeats are more likely to assume either alpha-helical or beta-sheet secondary structures than are polypeptide chains of random sequence. Third, provided that the number of bases in the oligomeric unit is not a multiple of 3, these internally repetitious coding sequences are impervious to randomly sustained base substitutions, deletions, and insertions. This is because the recurring periodicity of their polypeptide chains is given by three consecutive copies of the oligomeric unit translated in three different reading frames. Accordingly, when one reading frame is open, the other two are automatically open as well, all three being capable of coding for polypeptide chains of identical periodicity. Under this circumstance, a frame shift due to the deletion or insertion of a number of bases that is not a multiple of 3 fails to alter the down-stream amino acid sequence, and even a base change causing premature chain-termination can silence only one of the three potential coding units. Newly arisen coding sequences in modern organisms are oligomeric repeats, and most of the older genes retain various vestiges of their original internal repetitions. Some of the genes (e.g., oncogenes) have even inherited the property of being impervious to randomly sustained base changes.

Animals↗

PubMed1996

A protein class database organized with ProSite protein groups and PIR superfamilies.

A protein class (ProClass) database is developed as a "value-added" "second-generation" database organized according to family relationships. The database collects non-redundant protein sequence entries from SwissProt and PIR databases, and classifies them in families defined collectively by the ProSite protein groups and PIR superfamilies. The major objectives of the database are to maximize family information retrieval, to provide speedy family identification, and to help organizing existing protein sequence databases. The database has two sub-databases: PCFam (ProClass Family) to define protein families and provide links to ProSite patterns and PIR superfamilies, and PCSeq (ProClass Sequence) to describe sequence entries and provide links to PCFam, SwissProt, PIR, and ProSite databases. The current ProClass release has a total of 85,165 sequence entries, about half of which are classified in 3072 ProClass families; it also contains 10,431 newly established SwissProt-PIR links. The database can help reveal domain structures of related families, define new ProSite and PIR families, and provide family assignments for unclassified sequence entries. New ProSite and PIR family members are readily identified via database cross-reference, including 9437 SwissProt entries and 8522 PIR entries. False negative family members missed by both ProSite and PIR are detected using a neural network family identification system. The newly identified superfamily memberships are being incorporated into the current PIR database releases in a collaborative effort with the PIR. The ProClass database is accessible through anonymous FTP and on-line search on the World Wide Web.

Amino Acid Sequence↗

PubMed1993

Gene transfer and cardiovascular disorders.

Within the past four years, basic recombinant techniques (such as molecular cloning, sequencing, site-directed mutagenesis, PCR, and transfection) have been combined to yield a "second generation" of recombinant DNA technology with experimental potential which could barely have been envisioned only a decade ago. This review will focus upon the genesis and cardiovascular application of two recent developments in gene transfer technology: gene targeting by homologous recombination and direct in vivo gene transfer. Gene targeting evolved from transgenic mouse technology but is distinguished by its ability to precisely disrupt or "knock-out" specific genes in the murine genome. This not only provides decisive answers to functional questions, but also produces accurate models of human genetic disorders. In vivo gene transfer provides for the direct introduction of genetic information into living tissues. In vivo gene transfer not only facilitates basic research by providing a simple and direct way to analyze gene structure and function in intact animals, but may also find direct clinical application in the treatment of genetic and acquired disorders such as familial hypercholesterolemia and restenosis.

Animals↗

PubMed1997

Imaging of shifted stimulated echoes and multiple spin echoes.

It is shown that a repetitive pulse sequence consisting of two 90 degrees pulses and gradients in a 1:2 ratio around the second 90 degrees pulse generates interscan shifted stimulated echoes (SSTEs) and intrascan multiple spin echoes (MSEs). Separation of these two types of signals is accomplished using specific gradient crusher schemes. The intensity of the SSTEs is an order of magnitude larger than that of the MSEs and determines the signal contrast if both effects are selected simultaneously. The SSTE sequence generates improved contrast between gray and white matter, even at high field, which is explained in terms of increased inverse T1-weighting for the interscan echo. The MSE image has low signal to noise and no detectable contrast. The effect of interscan diffusion weighting is also discussed.

Animals↗

PubMed1986

Localization and DNA sequence analysis of the C gene of bacteriophage Mu, the positive regulator of Mu late transcription.

The C gene of bacteriophage Mu, required for transcription of the phage late genes, was localized by construction and analysis of a series of deleted derivatives of pKN50, a plasmid containing a 9.4 kb Mu DNA fragment which complements Mu C amber mutant phages for growth. One such deleted derivative, pWM10, containing only 0.5 kb of Mu DNA, complements C amber phages and transactivates the mom gene, one of the Mu late genes dependent on C for activation. The DNA sequence of the 0.5 kb fragment predicts a single long open reading frame coding for a 140 amino acid protein. Sequence analysis of DNA containing a C amber mutation located the base change to the second codon of this reading frame. Generation of a frameshift mutation by filling in a BglII site spanning codon 114 of this reading frame resulted in the loss of C complementation and transactivation activity. These results indicate that this open reading frame encodes the Mu C gene product. Comparison of the predicted amino acid sequence of the C protein with those of other transcriptional regulatory proteins revealed some similarity to a region highly conserved among bacterial sigma factors.

Amino Acid Sequence↗