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T Atkinson

Publications and source records attributed to T Atkinson.

At least 127 records · Page 7Linked to original sources
PubMed1987

Nucleotide sequence analysis of the gene for protein A from Staphylococcus aureus Cowan 1 (NCTC8530) and its enhanced expression in Escherichia coli.

Nucleotide sequence analysis of the gene (spa) for staphylococcal protein A (SPA) from Staphylococcus aureus strain Cowan 1 (NCTC8530) shows that the sequence differs from previously reported SPA nucleotide sequences, especially in the number of repeat units in the cell-wall-binding region of the gene. Dot matrix comparison with streptococcal protein G and the macrophage receptor for the constant fragment (Fc) of immunoglobulins shows a limited but significant homology. The homology to the latter probably identifies the Fc-binding region in the immunoglobulin-binding domains of SPA. Enhanced production of SPA in Escherichia coli was achieved using the lac promoter immediately upstream from the spa gene.

Amino Acid Sequence↗
PubMed1987

Melatonin lowers plasma prolactin levels in female red deer (Cervus elaphus).

The aim of the study was to determine the effect of exogenous melatonin treatment on circulating prolactin levels in red deer. Melatonin was administered from 12 June 1984 (day 1) to lactating and non-lactating hinds in the feed daily at 1600 h, and to non-lactating hinds by a subcutaneous implant. Average concentrations (ng/ml) of prolactin in plasma taken serially over 15-h periods were significantly higher for untreated hinds than for melatonin-treated animals on day 15 whether lactating (66-133 v. 23-28, P less than 0.05) or non-lactating (28-174 v. 8-13, P less than 0.01), remained higher on day 36 (lactating: 41-152 v. 15-21, P less than 0.05; non-lactating: 21-50 v. 1-7, P less than 0.001) but had decreased to similar levels on day 72 (lactating: 5-24 v. 7-17; non-lactating: 2-9 v. 0-4). The advanced reduction in plasma prolactin for all melatonin-treated hinds was associated with an advanced onset of seasonal breeding activity.

Animals↗
PubMed1986

Purification of homo- and hetero-oligonucleotides using high-performance charge-transfer chromatography.

Oligonucleotides employed in molecular biology have previously been purified by gel electrophoresis, gravity flow chromatography and more recently, high-performance liquid chromatography. However, these techniques have a number of problems and for this reason we investigated high-performance charge-transfer chromatography using the dye acriflavin coupled to silica as the stationary phase. Numerous oligonucleotides were purified using this technique and in this report we present data on four such oligonucleotides two homo-oligonucleotides and two hetero-oligonucleotides. Homogeneity of oligonucleotides eluted from the acriflavin matrices was determined by electrophoresis on 20% polyacrylamide gels and in each case they were greater than 90% pure.

Buffers↗
PubMed1986

Is there evidence for a common amino acid sequence in proteins with membrane attaching ability?

A comparison between the primary sequence of staphylococcal protein A (SpA) and a wide range of published protein sequences revealed a limited, but striking homology in approximately two-thirds of them. The region in the SpA sequence with the common homology was identified as the octapeptide repeats comprising the cell wall peptidoglycan-binding domain. Available structural information and the known location of the proteins within their host cells suggests that this common octapeptide may be important in interaction of the protein with the cell surface (either membrane or wall).

Amino Acid Sequence↗
PubMed1986

The purification and characterization of glucokinase from the thermophile Bacillus stearothermophilus.

Homogeneous glucokinase (EC 2.7.1.2) from the thermophile Bacillus stearothermophilus was isolated on the large scale by using four major steps: precipitation of extraneous material at pH 5.5, ion-exchange chromatography on DEAE-Sepharose, pseudo-affinity chromatography on Procion Brown H-3R-Sepharose 4B and gel filtration on Ultrogel AcA 34. The purified enzyme had a specific activity of about 330 units/mg of protein and was shown to exist as a dimer of subunit Mr 33,000. Kinetic parameters for the enzyme were determined with a variety of substrates. The glucokinase was highly specific for alpha-D-glucose, and the only other sugar substrate utilized was N-acetyl-alpha-D-glucosamine. The enzyme shows Michaelis-Menten kinetics, with a Km value of 150 microM for alpha-D-glucose. The glucokinase was maximally active at pH 9.0.

Carbohydrates↗
PubMed1986

An enzyme mediated, colorimetric method for the measurement of salicylate.

A novel enzymatic assay for salicylate in serum has been developed. Salicylate monooxygenase and NADH are used to convert the drug to catechol. This is reacted with 4-aminophenol at high pH to yield a blue product, which is detected colorimetrically. The assay is complete in less than seven minutes and requires no sophisticated equipment. The method is precise, sensitive and shows excellent accuracy in recovery experiments and when compared to a high performance liquid chromatography method. The assay is free from interference by coloured or turbid samples, salicylate metabolites, structurally related compounds such as benzoate and 4-hydroxybenzoate, and a range of drugs. The assay reagents demonstrate excellent stability. The formulation of the assay in two stages provides increased specificity and sensitivity compared to other emergency salicylate assays and allows the inclusion of reagents to greatly enhance the stability of the salicylate monooxygenase-NADH reagent, yet the method is simple and performs well.

Buffers↗
PubMed1986

A new rapid procedure for the preparation of plasmid DNA.

This report describes a simple and efficient procedure for the isolation of plasmid DNA free from chromosomal DNA, cellular RNA, and protein. The technique comprises a modified cleared lysate procedure of D.B. Clewell and D.R. Helinski (1969, Proc. Natl. Acad. Sci. USA, 62, 1159-1166) followed by high-performance liquid chromatography on a Dupont Bioseries GF250 surface stable diol-coated silica gel permeation column (Zorbax) for the final purification of the plasmid DNA. The use of HPLC facilitates rapid and high-resolution separations within 3-4 h. Plasmid DNA produced in this manner retains its biological activity and exhibits yields equal to those obtained by the conventional cesium chloride-ethidium bromide density centrifugation method.

Chromatography, High Pressure Liquid↗
PubMed1986

Nucleotide sequence of the Erwinia chrysanthemi NCPPB 1066 L-asparaginase gene.

The complete nucleotide sequence of the Erwinia chrysanthemi NCPPB 1066 gene coding for the chemotherapeutic enzyme L-asparaginase has been determined. The structural gene consists of an open reading frame commencing with an ATG start codon of 1044 bp followed by a TGA stop codon. Confirmation of the nucleotide sequence was obtained by comparing the predicted amino acid (aa) sequence with that derived by N-terminal aa sequencing of the purified protein. The gene has been shown to code for a 21-aa signal peptide at its N terminus which closely resembles the signal peptides of other secreted proteins. In common with highly expressed Escherichia coli genes, little use is made of modulator codons. The predicted aa sequence of the enzyme exhibits 46% identity with the determined primary sequence of the E. coli L-asparaginase, although the predicted secondary structure of both proteins indicates more extensive homology. Downstream of the TGA stop codon is a G + C-rich region of dyad symmetry (delta G = -25.4 kcal) characteristic of E. coli Rho-independent transcription terminators. Upstream of the structural gene there are no sequences which bear a strong resemblance to the consensus -35 and -10 regions of E. coli promoters. A sequence is present (CTGGCTCTCCTCTTGAT), however, which exhibits strong homology to the nif promoter consensus sequence (CTGGCACN5TTGCA). Upstream of this region is a sequence which strongly resembles the consensus sequence for promoter regions which are subject to catabolite repression.

Amino Acid Sequence↗
PubMed1986

Cloning and complete nucleotide sequence of the Bacillus stearothermophilus tryptophanyl tRNA synthetase gene.

The Bacillus stearothermophilus NCA 1503 tryptophanyl tRNA synthetase (WTS; EC 6.1.1.2) gene has been cloned in Escherichia coli and the amino acid (aa) sequence of the enzyme deduced unequivocally from the DNA sequence of the cloned gene. The predicted aa sequence of the WTS enzyme agrees with the previously determined aa sequence except that the DNA sequence indicates a third Arg residue at the C terminus of the enzyme over the two Arg residues indicated by sequencing the protein itself. The trpS gene consists of a 984-bp open reading frame commencing with an ATG start codon and ending with a TAA stop codon. Putative transcriptional promoters, a Shine-Dalgarno sequence and a transcription terminator have been identified. Thus the trpS gene probably constitutes a single transcriptional unit.

Amino Acid Sequence↗
PubMed1986

Cloning, expression and complete nucleotide sequence of the Bacillus stearothermophilus L-lactate dehydrogenase gene.

The structural gene for L-lactate dehydrogenase (LDH; EC 1.1.1.27) from Bacillus stearothermophilus NCA 1503 has been cloned in Escherichia coli and its complete nucleotide sequence determined. The predicted amino acid (aa) sequence of the LDH enzyme agrees with the previously determined aa sequence except to three positions: aa 125 and 126, Ser-Glu, are inverted whilst His at position 130 has been replaced by Ser in our sequence. The lct gene consists of an open reading frame (ORF) commencing from the ATG start codon of 951 bp followed by a TGA stop codon. Upstream from the start codon is a strong (delta G = -14.4 kcal) Shine-Dalgarno (SD) sequence, a feature typical of Gram-positive ribosome binding sites. Putative RNA polymerase recognition signals (-35 and -10 regions) have been identified upstream from the lct structural gene but there are no structures resembling Rho-independent transcription termination signals downstream from the TGA stop codon. Two further ORFs, preceded by SD sequences, are present downstream from the lct gene. Thus the lct gene may constitute the first gene of an operon. Subclones of the lct gene have been constructed in the expression plasmid pKK223-3 and the LDH enzyme produced in soluble form at levels of up to 36% of the E. coli soluble cell protein.

Amino Acid Sequence↗
PubMed1986

Persistence of virulent Semliki Forest virus in mouse brain following co-inoculation with defective interfering particles.

Semliki Forest virus (SFV) normally causes an acute lethal encephalitis in mice following intranasal inoculation. However, animals co-administered with 10 LD50 SFV and defective interfering (DI) SFV survive the infection without clinical signs of disease. In this report we demonstrate the isolation of infectious virus from the brains of 12/169 protected mice up to 6.5 months post-infection. Although, with one exception, mice were clinically normal, five of 12 of the SFV isolates were identical to the original virus as judged by plaque morphology, maximum temperature for growth, virulence in mice and pathology. Others were less virulent (although not any were plaque or temperature-sensitive mutants) and on re-inoculation into fresh mice caused a demyelinating pathology which was not an attribute of the original inoculum. How the virulent virus can persist in brain, sometimes in amounts in excess of 100 LD50, without causing disease remains to be determined.

Animals↗
PubMed1986

The isolation and characterization of bacteriophages infecting obligately thermophilic strains of Bacillus.

Twenty-four thermophilic bacteriophages have been isolated from diverse sources such as compost, soil, silage and rotting straw. Although considerable individual host specificity was observed, the phages were able to infect most of the major taxonomic groups of Bacillus thermophiles. The phages varied considerably in morphology and size; the phage heads were either cylindrical or polyhedral with tails varying in length between 15 and 500 nm. Most of the phages were stable at 50 degrees C for 4-5 h but at 70 degrees C the plaque-forming units decreased by between 10(2)- and 10(7)-fold in 2 h. The DNA of morphologically similar phages was examined by restriction enzyme analysis, and some differences in the DNA fragment patterns were found. Efficiency of plating data indicated that 'B. caldotenax' has a restriction and modification system. These phages may be valuable for the study of the genetics of thermophilic bacilli: transduction of 'B. caldotenax' and 'B. caldovelox' by phage JS017 has been observed.

Bacteriophages↗
PubMed1986

Toxicity of paraquat to microorganisms.

The biochemical response of the microorganisms Lipomyces starkeyi (Lod & Rij), Escherichia coli K-12 W3110, Bacillus subtilis 168 (Marburg) and Pseudomonas sp. strain TTO1 to the presence of growth-inhibitory concentrations of paraquat was studied. Paraquat was added to each culture at a concentration previously determined to reduce the culture growth rate by up to 50%. The changes in activity of a number of enzymes previously shown to be associated with the defense of the mammalian system against the action of paraquat were studied. While the response of E. coli was in agreement with that found in other studies of this microorganism and supports a commonly accepted mechanism for paraquat toxicity, the results obtained with L. starkeyi, B. subtilis, and Pseudomonas sp. strain TTO1 suggest that other mechanisms exist for protection against the toxicity of paraquat.

Bacillus subtilis↗
PubMed1986

Induction of early breeding in red deer (Cervus elaphus) by melatonin.

Lactating (N = 12) and non-lactating (N = 6) red deer hinds and one stag at pasture were given concentrates (500 g/head) containing melatonin (5 mg/head) daily at 16:00 h from 18 June to 16 October. The stag shed the antler velvet and started rutting 5 weeks ahead of untreated stags, and hinds had their first oestrus and ovulation of the breeding season in mid-September, 5 weeks in advance of control lactating (N = 9) and non-lactating (N = 5) hinds. Treated hinds were allowed to mate with the treated stag and control hinds ran in an adjacent paddock with an untreated stag. All hinds became pregnant during the study with all but 2 melatonin-fed hinds (1 lactating, 1 non-lactating) and 1 control hind (lactating) conceiving at the first oestrus. Melatonin-treated hinds lost slightly more live weight than did controls from June to November, but their suckled calves grew throughout at a rate similar to those of control hinds.

Animals↗
PubMed1986

Evaluation of epithelial tissue plasminogen activator as a thrombolytic agent in a rabbit model of venous thrombosis.

The thrombolytic effect of guinea pig keratocyte plasminogen activator was evaluated in rabbits with experimental jugular vein thrombosis and compared with human melanoma activator. Both the activators were infused locally at two dose levels in groups of three rabbits. Infusion of 6,000 and 16,000 I.U./rabbit over 4 hr resulted in 43 and 62% lysis with guinea pig keratocyte activator whereas melanoma activator induced 59 and 66% lysis with 6,000 and 12,000 I.U./rabbit. Thrombolysis with both these activators was not associated with systemic activation of the fibrinolytic system or fibrinogen breakdown. It is concluded that the guinea pig keratocyte activator is a specific thrombolytic agent with an in vivo potency very similar to the melanoma activator.

Animals↗