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Biomedical subjects

B M Ross

Publications and source records attributed to B M Ross.

At least 55 records · Page 3Linked to original sources
PubMed1994

Characterization of lysophospholipid metabolizing enzymes in human brain.

Lysophospholipids are generated during the turnover and breakdown of membrane phospholipids. We have identified and partially characterized three enzymes involved in the metabolism of lysophospholipids in human brain, namely, lysophospholipase, lysophospholipid:acyl-CoA acyltransferase (acyltransferase), and lysophospholipid:lysophospholipid transacylase (transacylase). Each enzyme displayed comparable levels of activity in biopsied and autopsied human brain, although in all cases the activity was somewhat lower in human than that in rat brain. All three enzymes were localized predominantly in the particulate fraction, with lysophospholipase possessing the greatest activity followed by acyltransferase and transacylase. Lysophosphatidylcholine possessed a Km in the micromolar range for lysophospholipase and transacylase, and in the millimolar range for acyltransferase, whereas arachidonyl-CoA displayed a Km in the micromolar range for acyltransferase. The three enzymes differed in their pH optima, with lysophospholipase being most active at pH 8.0, transacylase at pH 7.5, and acyltransferase at pH 6.0. Both bromophenacyl bromide and N-ethylmaleimide inhibited lysophospholipase activity and, to a lesser extent, that of acyltransferase and transacylase. None of the enzyme activities were affected by the presence of dithiothreitol or EDTA, although particulate lysophospholipase was activated approximately two-fold by the addition of 5 mM MgCl2 or CaCl2 but not KCl. Transacylating activity was stimulated by CoA, the EC50 of activation being 6.8 microM. Acyltransferase displayed an approximately threefold preference for arachidonyl-CoA over palmitoyl-CoA, whereas the acylation rate of different lysophospholipids was in the order lysophosphatidylinositol > 1-palmitoyl lysophosphatidylcholine > 1-oleoyl lysophosphatidylcholine >> lysophosphatidylserine > lysophosphatidylethanolamine. This, and the preference of human brain phospholipase A2 for phosphatidylinositol, suggests that this phospholipid may possess a higher turnover rate than the other phospholipid classes examined. Human brain homogenates also possessed the ability to transfer fatty acid from lysophosphatidylcholine to lysophosphatidylethanolamine. In addition, we also present evidence that diacylglycerophospholipids can act as acyl donors for the transacylation of lysophospholipids. We have therefore demonstrated the presence of, and partially characterized, three enzymes that are involved in the metabolism of lysophospholipids in human brain. Our results suggest that lysophospholipase may be the major route by which lysophospholipids are removed from the cell membrane in human brain. However, all three enzymes likely play an important role in the remodeling of membrane composition and thereby contribute to the overall functioning of membrane-associated processes.

Acetophenones↗
PubMed1994

Characterization of the Wilson disease gene encoding a P-type copper transporting ATPase: genomic organization, alternative splicing, and structure/function predictions.

Wilson disease is an autosomal recessive disorder of copper transport. Disease symptoms develop from the toxic build-up of copper primarily in the liver, and subsequently in the brain, kidney, cornea and other tissues. A candidate gene for WD (ATP7B) has recently been identified based upon apparent disease-specific mutations and a striking amino acid homology to the gene (ATP7A) responsible for another human copper transport disorder, X-linked Menkes disease (MNK). The cloning of WD and MNK genes provides the first opportunity to study copper homeostasis in humans. A preliminary analysis of the WD gene is presented which includes: isolation and characterization of the 5'-end of the gene; construction of a genomic restriction map; identification of all 21 exon/intron boundaries; characterization of extensive alternative splicing in brain; prediction of structure/function features of the WD and MNK proteins which are unique to the subset of heavy metal-transporting P-type ATPases; and comparative analysis of the six metal-binding domains. The analysis indicates that WD and MNK proteins belong to a subset of transporting ATPases with several unique features presumably reflecting their specific regulation and function. It appears that the mechanism of alternative splicing serves to regulate the amount of functional WD protein produced in brain, kidney, placenta, and possibly in liver.

Adenosine Triphosphatases↗
PubMed1993

Alterations in the activity of adenylate cyclase and high affinity GTPase in Alzheimer's disease.

The aim of this study was to assess the effect of Alzheimer's disease has on the functional integrity of several signal transduction proteins. The relative levels of the G-protein alpha subunits Gs alpha-L, Gs alpha-S, Gi alpha-2 and G(o) alpha were measured by western blotting and found to be unchanged in membranes prepared from Alzheimer-diseased frontal cortex or hippocampus compared to control brains. However the activity of the G-protein associated enzyme, high affinity GTPase, was found to be reduced in the frontal cortex (reduced by 25%) and by a similar magnitude in the hippocampus (reduced by 27%) of Alzheimer subjects. The same membrane preparations were also assayed for the activity of adenylate cyclase. Basal enzyme activity was not significantly altered in Alzheimer diseased hippocampus, but was markedly reduced (by 45%) in the frontal cortex. The ability of fluoride and aluminium ions to stimulate adenylate cyclase was not significantly changed in either brain region. This suggests that G-proteins, especially Gs, are still able to interact with this enzyme. These results indicate that although the presence of Alzheimer's disease does not significantly alter G-protein levels, changes have taken place in the overall activity of these proteins. However this alteration does not affect their ability to stimulate adenylate cyclase activity.

Adenylyl Cyclases↗
PubMed1993

Construction of a yeast artificial chromosome contig spanning the spinal muscular atrophy disease gene region.

The childhood spinal muscular atrophies (SMAs) are the most common, serious neuromuscular disorders of childhood second to Duchenne muscular dystrophy. A single locus for these disorders has been mapped by recombination events to a region of 0.7 centimorgan (range, 0.1-2.1 centimorgans) between loci D5S435 and MAP1B on chromosome 5q11.2-13.3. By using PCR amplification to screen yeast artificial chromosome (YAC) DNA pools and the PCR-vectorette method to amplify YAC ends, a YAC contig was constructed across the disease gene region. Nine walk steps identified 32 YACs, including a minimum of seven overlapping YAC clones (average size, 460 kb) that span the SMA region. The contig is characterized by a collection of 30 YAC-end sequence tag sites together with seven genetic markers. The entire YAC contig spans a minimum of 3.2 Mb; the SMA locus is confined to roughly half of this region. Microsatellite markers generated along the YAC contig segregate with the SMA locus in all families where the flanking markers (D5S435 and MAP1B) recombine. Construction of a YAC contig across the disease gene region is an essential step in isolation of the SMA-encoding gene.

Base Sequence↗
PubMed1992

On the stability of messenger RNA and ribosomal RNA in the brains of control human subjects and patients with Alzheimer's disease.

The levels of the mRNAs encoding the G protein subunits GS alpha, G beta 1, and G beta 2 were measured by northern blotting in the frontal cortex and hippocampus of control subjects and of patients with a clinical and histopathological diagnosis of dementia of the Alzheimer type (DAT). There was no significant difference, in either brain region, between the control and DAT groups for any of the G protein mRNAs measured. The degree of intersubject variability was very high, e.g., GS alpha mRNA in the frontal cortex (mean optical density +/- SD) was 405 +/- 342 in the control group versus 305 +/- 207 in the DAT group. The extent of generalised RNA degradation was assessed by detecting the breakdown products of 28S rRNA. RNA degradation was present in tissue samples from every human subject studied. The extent of 28S rRNA degradation in each subject was found to be related to the levels of G protein mRNA detected. The degree of RNA degradation in human subjects was found to be very variable and unaffected by the presence of DAT. RNA degradation correlated poorly with postmortem interval and this was confirmed by a controlled study of postmortem degradation in rat tissue. The possibility that the relative hypoxia and ischaemia in patients immediately before death could influence RNA degradation is discussed. The variable extent of RNA degradation means that great care must be taken to ensure the validity of RNA analyses undertaken in human postmortem brain, particularly when techniques are employed (such as in situ hybridisation) that themselves give no indication of RNA integrity.

Aged↗
PubMed1991

Transcriptional termination sequence at the end of the Escherichia coli ribosomal RNA G operon: complex terminators and antitermination.

We have examined the termination region sequence of the rrnG operon and have observed its properties in vivo using a fusion plasmid test system. Transcription of rrnG terminator fragments was also studied in vitro. We found that termination of rrnG transcription is a complex process controlled by a tandem Rho-independent and Rho-dependent terminator arrangement which we designate rrnG-tt'. Together, these two elements were 98% efficient at terminating transcription initiated at the rrnG-P2 promoter. When the two elements were separated, however, we found that the Rho-independent structure was only 59% efficient while the Rho-dependent fragment alone could account for total transcriptional termination of the tandem arrangement. The rrnG termination region was resistant to rrn antitermination and, therefore, possesses some means of stopping antiterminated transcription. The distal rrnG sequence contains several additional noteworthy features; the rrnGt' fragment contains a REP (repetitive extragenic palindromic) sequence and homology with a small unidentified reading frame following rrnE. This sequence is followed by witA, which is homologous to a citrate transport gene, citB. Finally, our sequence, obtained from plasmid pLC23-30, contains a Tn1000 insertion that is absent from the E. coli chromosome. This insertion lies 975 bp beyond the 5S gene and is not involved in the termination events examined in this study.

Amino Acid Sequence↗
PubMed1991

Robustness of G proteins in Alzheimer's disease: an immunoblot study.

Many of the neurotransmitter systems that are altered in senile dementia of the Alzheimer type are known to mediate their effects via G proteins, yet the integrity of guanine nucleotide-binding proteins (G proteins) in Alzheimer's diseased brains has received minimal investigation. The aim of this study was to establish whether the level of G alpha subunits of five G proteins was altered in Alzheimer's disease. We used immunoblotting (Western blotting) to compare the amounts of Gi1, Gi2, GsH (heavy molecular weight), GsL (light molecular weight), and Go in the frontal cortex and hippocampus, two regions severely affected by the disease, and the cerebellum, which is less severely affected. The number of senile plaques was also quantified. We report that there was no significant difference in the level of these G alpha subunits between Alzheimer's diseased and age-matched postmortem brains. These results suggest that alterations in the amount of G protein alpha subunits are not a feature of Alzheimer's disease.

Alzheimer Disease↗
PubMed1991

ClpB is the Escherichia coli heat shock protein F84.1.

ClpB is thought to be involved in proteolysis because of its sequence similarity to the ClpA subunit of the ClpA-ClpP protease. It has recently been shown that ClpP is a heat shock protein. Here we show that ClpB is the Escherichia coli heat shock protein F84.1. The F84.1 protein was overproduced in strains containing the clpB gene on a plasmid and was absent from two-dimensional gels from a clpB null mutation. Besides possessing a slower growth rate at 44 degrees C, the null mutant strain had a higher rate of death at 50 degrees C. We used reverse transcription of in vivo mRNA to show that the clpB gene was expressed from a sigma 32-specific promoter consensus sequence at both 37 and 42 degrees C. We noted that the clpB+ gene also caused the appearance of a second protein spot, F68.5, on two-dimensional gels. This spot was approximately 147 amino acids smaller than F84.1 and most probably is the result of a second translational start on the clpB mRNA. F68.5 can be observed on many published two-dimensional gels of heat-induced E. coli proteins, but the original catalog of 17 heat shock proteins did not include this spot.

Base Sequence↗
PubMed1990

Induction of the heat shock response in rats modulates heart rate, creatine kinase and protein synthesis after a subsequent hyperthermic treatment.

STUDY OBJECTIVE - The aim of the study was to examine the effect of prior induction of the heat shock response on heat shock protein synthesis and physiological variables relevant to the shock response. DESIGN - Synthesis of heat shock protein (SP71, molecular mass 71,000) was induced in rats by 15 min hyperthermia (42 degrees C). Protein synthesis, heart rate, blood pressure and creatine kinase activity were determined in comparison with controls (no heat shock) and a group receiving two heat shock treatments 24 h apart (prior induction group). SUBJECTS - 24 male Sprague-Dawley rats (125-150 g) were used, divided into three groups: controls (n = 4), heat shock X 1 (HS, n = 11), heat shock X 2 (2 X HS, n = 9). Heat shock was induced under anaesthesia on a heating pad. MEASUREMENTS and RESULTS - Blood pressure and heart rate were measured at the beginning of the hyperthermia period, when body temperature first reached 42 degrees C (t = 0 min) and at the end of the hyperthermia treatment (t = 15 min). At t = 0 min systolic blood pressure and heart rate were increased compared to the control values in both HS and 2 X HS groups. At t = 15 min heart rate in the HS group was increased to 554 (SEM21) beats.min-1 v control 465(19) (p less than 0.05). In the 2 X HS group, heart rate of 494(14) beats.min-1 at t = 15 min was not significantly different from control. At t = 15 min, creatine kinase values in the hyperthermia treatment groups were not different from control. However at 2.5 h after hyperthermic treatment plasma creatine kinase was increased in the HS group to 481(83) mU.ml-1 (n = 6) v 223(20) in controls, but was not increased in the 2 X HS group [178(64), n = 4]. Rats were radiolabelled for 2 h with 1.0 mCi of [35S]-methionine 30 min after hyperthermic treatment in HS group and 30 min after the second hyperthermic treatment in 2 X HS group. Following the 2 X HS treatment, synthesis of SP71, though increased above control values, was lower than in the HS group. CONCLUSIONS - The reduction in heart rate, plasma creatine kinase and synthesis of SP71 following a second hyperthermic exposure could be caused by a protective influence of the first exposure.

Animals↗
PubMed1988

One type of gamma-turn, rather than the other gives rise to chain-reversal in proteins.

Gamma-turns may be defined by a hydrogen bond between the carbonyl group of one amino acid residue and the amino group of the acid two residues ahead in the sequence. They occur as two types, inverse gamma-turns and classic gamma-turns (classic gamma-turns are usually called just gamma-turns but we prefer to add the adjective classic to distinguish them from the word gamma-turn, referring collectively to both). Of the two, classic gamma-turns are less common and are considered by all authors to be extreme rarities in proteins. However, we find that a number do occur in a sample of proteins of known three-dimensional structure. One occurs at the edge of the second hypervariable region of the light chain in some immunoglobulins. All classic gamma-turns except one are associated with a reversal in the main chain direction. In most cases, the turn lies at the loop end of a beta-hairpin. By contrast, inverse gamma-turns, although giving rise to a kink in the chain, rarely occur within beta-hairpins and are seldom situated at a position of reversal, by 180 degrees, in chain direction.

Amino Acids↗
PubMed1979

Memorizing and copying visual patterns: a Piagetian interpretation.

Six-, 8-, 10-, and 12-year-old children (10 boys and 10 girls) reconstructed two visual patterns from immediate memory, while other 5- and 6-year-old children (10 boys and 10 girls) reconstructed the identical patterns by direct copying. Patterns were simple and composed entirely of circles or squares as component items. Four results were emphasized: (a) Numerous errors mady by the copying groups led to the conclusion that memory loss is often overestimated in young children. Since an independent estimate of perceptual encoding errors is rarely carried out, encoding mistakes are often included among forgetting errors. (b) One pattern was both copied and remembered more poorly than the other in accord with a Piagetian interpretation of a conceptual conflict inherent in the pattern design between spatial and numerical correspondence of component pattern items. (c) A memory strategy emphasizing configuration preservation was suggested for the 6-year-olds who made slightly fewer memory than copying errors for two configural scoring categories. (d) Performance in an unrelated planning-for-memory task significantly differentiated between better and worse performers on the visual pattern memory task.

Age Factors↗
PubMed1979

Three-dimensional structure of the central mitotic spindle of Diatoma vulgare.

Central mitotic spindles in Diatoma vulgare have been investigated using serial sections and electron microscopy. Spindles at both early stages (before metaphase) and later stages of mitosis (metaphase to telophase) have been analyzed. We have used computer graphics technology to facilitate the analysis and to produce stereo images of the central spindle reconstructed in three dimensions. We find that at prometaphase, when the nuclear envelope is dissassembling, the spindle is constructed from two sets of polar microtubules (MTs) that interdigitate to form a zone of overlap. As the chromosomes become organized into the metaphase configuration, the polar MTs, the spindle, and the zone of overlap all elongate, while the number of MTs in the central spindle decreases from greater than 700 to approximately 250. Most of the tubules lost are short ones that reside near the spindle poles. The previously described decrease in the length of the zone of overlap during anaphase central spindle elongation is clearly demonstrated in stereo images. In addition, we have used our three-dimensional data to determine the lengths of the spindle MTs at various times during mitotis. The distribution of lengths is bimodal during prometaphase, but the short tubules disappear and the long tubules elongate as mitosis proceeds. The distributions of MT lengths are compared to the length distributions of MTs polymerized in vitro, and a model is presented to account for our findings about both MT length changes and microtubule movements.

Anaphase↗
PubMed1976

Comparison of aggression and incongruity as factors in children's judgments of humor.

It was hypothesized that both aggression and incongruity are variables that will positively influence preschool children's judgments of humorous incidents. Past research has shown the influence of incongruity on humor only for school-age children, and--with the exception of studies of children's television viewing--the allegedly "antisocial" variable of aggression has been ignored. Two hundred and thirty boys and girls from ages three to eight each made six paired comparison judgments as to which of two brief incidents was funnier. The incidents were produced by the E manipulating either one or two hand puppets with or without accompanying dialogue. At all ages children significantly preferred aggressive and incongruous incidents to neutral incidents. When aggressive and incongruous nonverbal incidents were paired, there was no significant preference. Two major conclusions were reached: (a) Though neglected in the developmental theory of humor, aggression is an early occurring and potent factor in children's humor; (b) The clear-cut results obtained from preschoolers argue for the usefulness of cognitive choice as against such observational measures as smiling and laughing.

Aggression↗
PubMed1975

The attainment of formal operations: a comparison of probability concepts in deaf and hearing adolescents.

In a series of six experiments deaf and hearing adolescents were compared on attainment of formal operations in Piaget's theory as represented by performance on probability problems. Performance of deaf subjects provided a desirable control comparison, since their knowledge of probability concepts was most likely acquired outside school classrooms. The probability problems were usually binary choice situations that required no verbalizations and were designed to illuminate strategies, as well as to indicate conceptual stage. Two experiments, one a long-term training study of over two years' duration, used supplementary preadolescent subject groups. Most of the hearing adolescents and many of the deaf adolescents attained the formal operatory level for probability concepts according to the criteria described by Piaget and Inheldermtheir criteria can be criticized, however, as insufficiently demanding, although they do point up the difficulty of defining the place of computational abilities in formal operationsmresults from several experiments suggested a second and higher level of formal operatory attainment around the ages of 14 to 15. Further theoretical insights were obtained regarding the implicit rules that frequently appeared to guide subjects' choices; a surprising variety of strategies were reported. Analysis of choice tendendies showed consistent avoidance of calculations and selection of the choice with the greatest absolute rather than relative frequency (or area) even when these strategies produced systematic errors. Subjects were generally unable to combine probability information from two different sources. In the long-term training experiment spontaneous improvement was shown by children at formal operatory ages even without training, but not by younger children. Deaf adolescents performed as well as hearing adolescents on easy problems, but they made more errors on several types of more difficult problems; Deaf adolescents were also less consistent than the hearing in use of rules, and in several experiments they showed a two- to three-year performance lag.

Achievement↗