Anomalous Raman spectra from La2CuO4.
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Journal Article↗
Biomedical subjects
G Burns
Publications and source records attributed to G Burns.
Opioid effects on intake of sweet solutions depend both on prior drug experience and on prior ingestive experience.
Two experiments investigated the effect of opioids on ingestion of sweet solutions in non-deprived rats. Experiment 1 replicated previous work from our laboratory showing virtually complete inhibition of sucrose and saccharin intake during 10 days of daily naloxone treatment. During recovery, prior naloxone experience significantly stimulated sucrose intake but had no effect on saccharin intake. In the absence of naloxone treatment, ingestive experience alone reduced naloxone's typical intake-suppressant effect. These findings suggest that drug experience and ingestive experience may interact to determine the intake-suppressant effect of naloxone. Experiment 2 examined the effects of opioid agonists on sucrose ingestion during 10 days of initial drug treatment and 5 days of recovery. A low dose of the kappa agonist U-50,488H significantly stimulated sucrose ingestion during the drug treatment period and this effect persisted for several days after treatment ended. Initial (non-significant) intake suppressant effects of the mu agonist morphine or a high dose of U-50,488H tended to decrease with repeated testing and did not reappear during recovery. These data suggest that in addition to immediate, direct effects on motivation, opioids may affect long-term changes in responsiveness to sweet tastes.
Albinism↗
Stimulation of hypothalamic opioid peptide release by lithium is mediated by opioid autoreceptors: evidence from a combined in vitro, ex vivo study.
The effect of both chronic and acute lithium treatment on hypothalamic opioid peptides was investigated. Acute treatment with lithium was found to stimulate the release of beta-endorphin, dynorphin and Met-enkephalin from perfused rat hypothalamic slices. Application of tetrodotoxin was found to have no effect upon the stimulation indicating it to be mediated at the nerve terminal level. The release of hypothalamic opioid peptides is known to be under the chronic control of a system of inhibitory autoreceptors. Blockade of these autoreceptors with, for example, the opioid receptor antagonist naloxone causes a release of all three opioid peptides. Simultaneous addition of naloxone and lithium was found to have no additive effect on the release of any opioid, suggesting lithium acts via an inhibition of the inhibitory autoreceptor. Preincubation with pertussis toxin prevented the lithium stimulation of dynorphin and Met-enkephalin, but not beta-endorphin, release, indicating lithium interacts with a G-protein to affect the autoreceptor controlling the release of dynorphin and Met-enkephalin. Chronic treatment with lithium in vivo (10 days) had no effect on the basal release or hypothalamic content of any of the opioids, but prevented the naloxone-stimulated release of dynorphin and Met-enkephalin in vitro. Long-term treatment with lithium would thus appear to inactivate the autoreceptor(s) controlling their release. These data demonstrate a lithium-stimulated release of hypothalamic beta-endorphin, Met-enkephalin and dynorphin, apparently mediated via an inhibition of the autoreceptors controlling their release. Chronic treatment with lithium permanently inactivated the autoreceptor(s) controlling the release of dynorphin and Met-enkephalin but not beta-endorphin. Lithium would appear to mediate its effects upon Met-enkephalin and dynorphin release via an interaction with a pertussis toxin-sensitive G-protein. The mechanisms underlying its release of beta-endorphin are at present uncertain.
Animals↗
Transcriptional analysis of the promoter region of the Pseudomonas putida branched-chain keto acid dehydrogenase operon.
Branched-chain keto acid dehydrogenase is a multienzyme complex produced by Pseudomonas putida when it is grown in a minimal medium containing branched-chain amino acids. A 1.87-kilobase (kb) DNA fragment was cloned and sequenced which contained 0.24 kb of the E1 alpha structural gene and 1.6 kb of upstream DNA. There were 854 base pairs (bp) of noncoding DNA upstream of bkdA1, the first gene of the bkd operon, and 592 bp between the transcriptional and translational starts. The G + C content of the noncoding region was 56.7% compared with 65.2% for all the structural genes of the operon. A partial open reading frame was found on the strand opposite that of the bkd operon beginning at base 774. When the bkd promoter was cloned into the promoter probe vector pKT240, streptomycin resistance was obtained in P. putida but not Escherichia coli with the promoter in both orientations, which indicates either that the bkd promoter is bidirectional or that there are two promoters in this region. A series of ordered deletions on both sides of the proposed site of the start of transcription revealed that almost 700 bp upstream of the start of translation were required for expression. Streptomycin resistance was also obtained in an rpoN mutant of P. putida KT2440 containing constructs with the intact bkd promoter, indicating that the bkd operon does not require the rpoN sigma factor for expression. Another construct containing the bkd promoter, bkdA1, and bkdA2 in pKT240 was used to transform P. putida JS113, a mutant which was unable to produce the E1 subunits of the branched-chain keto acid dehydrogenase. In this case, very high inducible expression of the bkd operon was obtained.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
Lectin binding in murine tumour lines with different malignant characteristics. Effect of enzyme (pronase, neuraminidase) treatment on lectin labelling.
We used 5 syngeneic murine tumour systems for studying quantitative lectin surface binding of intact viable tumour cells. We also investigated, for 3 of the tumours, whether the lectin binding sites were susceptible to proteolytic enzyme (pronase) or neuraminidase treatment. There were significant differences between two of the tumour lines in the binding of wheat germ agglutinin (WGA), concanavalin A (ConA). peanut agglutinin (PNA), soybean agglutinin (SBA) and Ulex europeus agglutinin (UEA). There were also variations in lectin binding between the other tumor lines, but these differences were not statistically significant. Lectin binding showed no evident relationship to the malignancy or the metastasis-forming capacity of the respective tumour cell line. Proteolytic treatment, which drastically affects intravenously induced experimental metastasis formation by one of the tumours, caused a decreased binding of SBA, ConA and WGA. Neuraminidase treatment increased both PNA and SBA binding in three different cell lines, presumably by removing sialic acid masking D-galactose and N-acetyl-D-galactose-amine groups.
Animals↗
Raman-active phonons in the high-temperature superconductors and results in Bi2Sr2CuO6.
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Journal Article↗
Sequence analysis of the lpdV gene for lipoamide dehydrogenase of branched-chain-oxoacid dehydrogenase of Pseudomonas putida.
The production of two lipoamide dehydrogenases by Pseudomonas is so far unique. One, LPD-val, is the specific E3 component of the branched-chain-oxoacid dehydrogenase and the second, LPD-glc, is the E3 component of 2-oxoglutarate dehydrogenase and the L-factor of the glycine oxidation system. The objective of the present research was to determine the nucleotide sequence of the structural gene for LPD-val in order to compare its deduced amino acid structure with that of other redox-active disulfide flavoproteins. Northern blots using mRNA isolated from P. putida grown in media with branched-chain amino acids identified a transcript of 6.2 kb which is long enough to encode all the structural genes for the complex. The nucleotide sequence of the structural gene for LPD-val, lpdV, was determined and consists of 459 codons plus the stop codon. The open reading frame begins two bases after the stop codon for the E2 subunit and is composed of 66.3% G + C. Codon usage is characteristic of moderately strongly expressed genes. There is a ribosome-binding site preceding the ATG start codon and a strong candidate for a rho-independent terminator at the 3' end of the reading frame. The Mr of the protein encoded is 48,164 and when the Mr of FAD is added, the total Mr is 48,949, which is very close to the value of 49,000 obtained by SDS-polyacrylamide gel electrophoresis. Similarity comparisons of LPD-val with sequences of three other lipoamide dehydrogenases showed that LPD-val was somewhat more distantly related. It is probable that the lipoamide dehydrogenases and the glutathione and mercuric reductases evolved from a common ancestral flavoprotein.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
Temperature dependence of the Raman spectra for Bi2CaSr2Cu2O8.
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Journal Article↗
Viewing distance affects stereoscopic tilt created with spatial frequency disparity.
A controversy exists as to whether stereoscopic tilt created with interocular differences in spatial frequency is based on perception of spatial frequency disparity or positional disparity. To determine which hypothesis is correct, we investigated the influence of viewing distance on perceived tilt. Tilt was induced by having observers view, at three viewing distances, dichoptic spatial frequency grating patterns differing in frequency by 25%. By appropriate physical scaling of the size of the patterns, their spatial frequency and angular width remained unchanged as distance varied. Under such conditions, the spatial frequency disparity hypothesis predicts no effect of distance, whereas the positional disparity hypothesis predicts a significant effect of distance (due to stereoscopic depth constancy) on the magnitude of tilt. The results showed that perceived tilt does covary with distance, a result consistent with only the positional disparity hypothesis.
Adolescent↗
Isolation of a third lipoamide dehydrogenase from Pseudomonas putida.
Pseudomonads are the only organisms so far known to produce two lipoamide dehydrogenases (LPDs), LPD-Val and LPD-Glc. LPD-Val is the specific E3 component of branched-chain oxoacid dehydrogenase, and LPD-Glc is the E3 component of 2-ketoglutarate and possibly pyruvate dehydrogenases and the L-factor of the glycine oxidation system. Three mutants of Pseudomonas putida, JS348, JS350, and JS351, affected in lpdG, the gene encoding LPD-Glc, have been isolated; all lacked 2-ketoglutarate dehydrogenase, but two, JS348 and JS351, had normal pyruvate dehydrogenase activity. The pyruvate and 2-ketoglutarate dehydrogenases of the wild-type strain of P. putida were both inhibited by anti-LPD-Glc, but the pyruvate dehydrogenase of the lpdG mutants was not inhibited, suggesting that the mutant pyruvate dehydrogenase E3 component was different from that of the wild type. The lipoamide dehydrogenase present in one of the lpdG mutants, JS348, was isolated and characterized. This lipoamide dehydrogenase, provisionally named LPD-3, differed in molecular weight, amino acid composition, and N-terminal amino acid sequence from LPD-Glc and LPD-Val. LPD-3 was clearly a lipoamide dehydrogenase as opposed to a mercuric reductase or glutathione reductase. LPD-3 was about 60% as effective as LPD-Glc in restoring 2-ketoglutarate dehydrogenase activity and completely restored pyruvate dehydrogenase activity in JS350. These results suggest that LPD-3 is a lipoamide dehydrogenase associated with an unknown multienzyme complex which can replace LPD-Glc as the E3 component of pyruvate and 2-ketoglutarate dehydrogenases in lpdG mutants.
Amino Acid Sequence↗
Effect of sublethal concentration of antibiotics on the adherence of Staphylococcus epidermidis to eukaryotic cells.
Adherence of Staphylococcus epidermidis to pharyngeal epithelial cells as measured by [3H]-leucine-labelled bacteria was found to be a simple, objective and highly reproducible technique. Sublethal concentrations of cephalothin, rifampicin and vancomycin significantly reduced the bacterial adherence to epithelial cells, the effect being most marked with 1/2 MIC. Cloxacillin increased the adherence for one strain while it decreased it for the second. The effect was dose-related and the maximum reduction was observed when bacteria were grown with an antibiotic overnight.
Animals↗
A two-step mechanism by which corticotropin-releasing hormone releases hypothalamic beta-endorphin: the role of vasopressin and G-proteins.
It is well established that in the pituitary gland corticotropin-releasing hormone (CRH) stimulates the release of beta-endorphin (beta-E) via a cAMP-linked mechanism. Studies of the mechanisms underlying the CRH stimulation of beta-E release from rat hypothalamic slices perifused in vitro are reported in this paper. The data indicate that both a cAMP-dependent and non-cAMP-dependent mechanism mediate the action of CRH in the hypothalamus. The presence of a cAMP-linked mechanism was suggested by the finding that cholera toxin (0.1-10 nM) and forskolin (2.5 x 10(-6) M), both of which act to raise intracellular cAMP levels, stimulated the release of beta-E. In both cases, no further stimulation was seen upon addition of CRH (10(-8)M). However, it was also found that preincubation of the tissue with pertussis toxin (PTX; 100 ng/ml) prevented both the CRH- and forskolin-stimulated release of beta-E. This indicated that, in addition to the cAMP-linked mechanism, a further messenger system which is connected to a PTX-sensitive G-protein may also play a role. The latter observation also implied that a further substance, which utilizes a separate second messenger system, might be involved in the CRH stimulation of beta-E release. In this regard the role of arginine vasopressin (AVP) was investigated due to the known interaction between CRH and AVP in the pituitary gland. AVP (10(-12) to 10(-6)M) itself potently and dose-dependently stimulated beta-E release, producing a maximal increase of 220% above basal levels. The AVP-induced release of beta-E was abolished in PTX-pretreated hypothalami. The apparently obligatory requirement of AVP for the CRH-stimulation of beta-E release was illustrated by the finding that blockade of AVP receptors using the AVP antagonist d(CH2)5 [Tyr(OEt)2,Val4]-AVP almost completely attenuated the CRH-stimulated release of beta-E. Furthermore, in the presence of a high concentration of AVP (10(-6)M) no further stimulation of release was seen with CRH (10(-8)M). These data therefore strongly indicate that CRH acts via the intermediacy of AVP to release beta-E from hypothalamic slices in vitro and that two separate second messenger systems are involved: a cAMP-linked mechanism connected to a cholera toxin-sensitive G-protein (CRH) and a second system linked to a PTX-sensitive G-protein (AVP).
Animals↗
Absence of a single repeat from the coding region of the human involucrin gene leading to RFLP.
The human involucrin gene has been mapped to the region q21-q22 of chromosome 1. Three of six Utah families examined were polymorphic for a PstI fragment of the involucrin gene. In one individual, the variant PstI fragment was found by DNA sequencing to be missing one of the 39 repeats that make up two-thirds of the coding region.
Alleles↗
Similarity of the E1 subunits of branched-chain-oxoacid dehydrogenase from Pseudomonas putida to the corresponding subunits of mammalian branched-chain-oxoacid and pyruvate dehydrogenases.
The genes encoding proteins responsible for activity of the E1 component of branched-chain-oxoacid dehydrogenase of Pseudomonas putida have been subcloned and the nucleotide sequence of this region determined. Open reading frames encoding E1 alpha (bkdA1, 1233 bp) and E1 beta (bkdA2, 1020 bp) were identified with the aid of the N-terminal sequence of the purified subunits. The Mr of E1 alpha was 45,158 and of E1 beta was 37,007, both calculated without N-terminal methionine. The deduced amino acid sequences of E1 alpha and E1 beta had no similarity to the published sequences of the E1 subunits of pyruvate and 2-oxoglutarate dehydrogenases of Escherichia coli. However, there was substantial similarity between the E1 alpha subunits of Pseudomonas and rat liver branched-chain-oxoacid dehydrogenases. In particular, the region of the E1 alpha subunit of the mammalian branched-chain-oxoacid dehydrogenase which is phosphorylated, was found to be highly conserved in the Pseudomonas E1 alpha subunit. There was also considerable similarity between the E1 beta subunits of Pseudomonas branched-chain-oxoacid dehydrogenase and human pyruvate dehydrogenase.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
Comparison of the amino acid sequences of the transacylase components of branched chain oxoacid dehydrogenase of Pseudomonas putida, and the pyruvate and 2-oxoglutarate dehydrogenases of Escherichia coli.
The nucleotide sequence of bkdB, the structural gene for E2b, the transacylase component of branched-chain-oxoacid dehydrogenase of Pseudomonas putida has been determined and translated into its amino acid sequence. The start of bkdB was identified from the N-terminal sequence of E2b isolated from branched-chain-oxoacid dehydrogenase of the closely related species, P. aeruginosa. The reading frame was composed of 65.5% G + C with 82.3% of the codons ending in G or C. There was no intergenic space between bkdA2 and bkdB. No codons requiring minor tRNAs were utilized and the codon bias index indicated a preferential codon usage. The bkdB gene encoded 423 amino acids although the N-terminal methionine was absent from E2b prepared from P. aeruginosa. The relative molecular mass of the encoded protein was 45,134 (45,003 minus methionine) vs 47,000 obtained by SDS/polyacrylamide gel electrophoresis. There was a single lipoyl domain in E2b compared to three lipoyl domains in E2p, and one domain in E2o, the transacylases of pyruvate and 2-oxoglutarate dehydrogenases of Escherichia coli respectively. There was significant similarity between the lipoyl domain of E2b and of E2p and E2o as well as between the E1-E2 binding domains of E2b, E2p and E2o. There was no similarity between the E3 binding domain of E2b to E2p and E2o which may reflect the uniqueness of the E3 component of branched-chain-oxoacid dehydrogenase of P. putida. The conclusions drawn from these comparisons are that the transacylases of prokaryotic pyruvate, 2-oxoglutarate and branched-chain-oxoacid dehydrogenases descended from a common ancestral protein probably at about the same time.
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
Translation of globin RNA spliced in vitro.
Pre-mRNA (precursor mRNA) enriched for globin-gene transcription products was prepared from murine erythroleukaemia cells induced to differentiate with dimethyl sulphoxide. The pre-mRNA was prepared from nuclear RNA by oligo(dT)-cellulose chromatography followed by sucrose-gradient centrifugation. The pre-mRNA thus obtained was utilized as the substrate for a splicing assay based on sequential splicing and translation reactions. Small nuclear ribonucleoprotein particles, which were shown to inhibit translation, were removed by an intermediate high-speed centrifugation step. The assay developed is based on a three-step procedure: (1) incubation of pre-mRNA with HeLa-cell nuclear extract in the presence of ATP; (2) incorporation of 3H-labelled amino acid into proteins by wheat-germ extract by using RNA from step 1; and (3) immunoprecipitation of globin chains with rabbit anti-(mouse globin) antibody. No incorporation of label into immune complexes was observed in the absence of HeLa-cell extract or ATP. Preincubation with these components was unnecessary for the incorporation of label into immune complexes when mature cytoplasmic RNA was used as a template for protein synthesis. The assay was further validated by fluorography after polyacrylamide-gel electrophoresis, when the only band detectable corresponded to globin chains. These results demonstrate, for the first time, translational activity of transcripts spliced in vitro. The current assay can be utilized with a few micrograms of pre-mRNA and may have potential for further characterization of the splicing system.
Cell Line↗
Low-temperature specific heat in ferroelectric (Pb,Ba)5Ge3O11 crystals.
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Journal Article↗