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Biomedical subjects

E Goldman

Publications and source records attributed to E Goldman.

At least 55 records · Page 3Linked to original sources
PubMed1992

Editing of errors in selection of amino acids for protein synthesis.

All living cells must conduct protein synthesis with a high degree of accuracy maintained in the transmission and flow of information from gene to finished protein product. One crucial "quality control" point in maintaining a high level of accuracy is the selectivity by which aminoacyl-tRNA synthetases furnish correctly activated amino acids, attached to tRNA species, as the building blocks for growing protein chains. During selection of amino acids, synthetases very often have to distinguish the cognate substrate from a homolog having just one fewer methyl group in its structure. The binding energy of a methyl group is estimated to contribute only a factor of 100 to the specificity of binding, yet synthetases distinguish such closely related amino acids with a discrimination factor of 10,000 to 100,000. Examples of this include methionine versus homocysteine, isoleucine versus valine, alanine versus glycine, and threonine versus serine. Many investigators have demonstrated in vitro the ability of certain aminoacyl-tRNA synthetases to edit, that is, correct or prevent incorrect attachment of amino acids to tRNA molecules. Several major editing pathways are now established from in vitro data. Further, at least some aminoacyl-tRNA synthetases have recently been shown to carry out the editing function in vivo. Editing has been demonstrated to occur in both Escherichia coli and Saccharomyces cerevisiae. Significant energy is expended by the cell for editing of misactivated amino acids, which can be reflected in the growth rate. Because of this, cellular levels of aminoacyl-tRNA synthetases, as well as amino acid biosynthetic pathways which yield competing substrates for protein synthesis, must be carefully regulated to prevent excessive editing. High-level expression of recombinant proteins imposes a strain on the biosynthetic capacity of the cell which frequently results in misincorporation of abnormal or wrong amino acids owing in part to limited editing by synthetases. Unbalanced amino acid pools associated with some genetic disorders in humans may also lead to errors in tRNA aminoacylation. The availability of X-ray crystallographic structures of some synthetases, combined with site-directed mutagenesis, allows insights into molecular details of the extraordinary selectivity of synthetases, including the editing function.

Amino Acids↗
PubMed1991

Low-usage codons in Escherichia coli, yeast, fruit fly and primates.

Codon usage is compared between four classes of species, with an emphasis on characterization of low-usage codons. The classes of species analyzed include the bacterium Escherichia coli (ECO), the yeast Saccharomyces cerevisiae (YSC), the fruit fly Drosophila melanogaster (DRO), and several species of primates (PRI) (taken as a group; includes eleven species for which nucleotide sequence data have been reported to GenBank, however, greater than 90% of the sequences were from Homo sapiens). The number of protein-coding sequences analyzed were 968 for ECO, 484 for YSC, 244 for DRO, and 1518 for PRI. Three methods have been used to determine low-usage codons in these species. The first and most common way of assessing codon usage is by summing the number of time codons appear in reading frames of the genome in question. The second way is to examine the distribution of usage in different genes by scoring the number of protein reading frames in which a particular codon does not appear. The third way starts with a similar notion, but instead considers combinations of codons that are missing from the maximum number of genes. These three methods give very similar results. Each species has a unique combination of eight least-used codons, but all species contain the arginine codons, CGA and CGG. The agreement between YSC and PRI is particularly striking as they share six low-usage codons. All six carry the dinucleotide sequence, CG. The eight least-used codons in PRI include all codons that contain the CG dinucleotide sequence. Low-usage codons are clearly avoided in genes encoding abundant proteins for ECO, YSC DRO. In all species, proteins containing a high percentage of low-usage codons could be characterized as cases where an excess of the protein could be detrimental. Low codon usage is relatively insensitive to gross base composition. However, dinucleotide usage can sometimes influence codon usage. This is particularly notable in the case of CG dinucleotides in PRI.

Animals↗
PubMed1991

Stress response protein (srp-27) determination in primary human breast carcinomas: clinical, histologic, and prognostic correlations.

Expression of an estrogen-regulated protein known as the 27,000-d heat-shock or stress-response protein (srp-27) was evaluated in human breast carcinomas and established breast cancer cell lines. Results obtained by Northern and Western blot analyses and immunohistochemical methods were concordant. Immunohistochemical assessment of srp-27 expression in 300 breast carcinomas (with median patient follow-up of 8 years) was performed. Twenty-six percent of lymph node-negative and 45% of lymph node-positive tumors were overexpressors. Univariate analysis demonstrated significant correlations between srp-27 overexpression and estrogen receptor (ER) content, pS2 protein expression, nodal metastases, advanced T stage, lymphatic/vascular invasion, and a shorter disease-free survival period (but not a shorter overall survival) for the study population as a whole. Regression tree analysis showed that srp-27 expression was an independent prognostic indicator for disease-free survival only in patients with one to three positive lymph nodes. The Cox proportional hazards model confirmed the independent prognostic significance of nodal involvement, T stage, and ER content but failed to recognize srp-27 overexpression as a significant independent parameter predictive of patient outcome in the patient population as a whole. The observed associations between srp-27 overexpression and more aggressive tumors suggest a biologic role for srp-27 in human breast carcinomas.

Blotting, Western↗
PubMed1991

Bacteriophage T7 morphogenesis and gene 10 frameshifting in Escherichia coli showing different degrees of ribosomal fidelity.

Bacteriophage T7 infection has been studied in Escherichia coli strains showing both increased and decreased ribosome fidelity and in the presence of streptomycin, which stimulates translational misreading, in an effort to determine effects on the apparent programmed translational frameshift that occurs during synthesis of the gene 10 capsid protein. Quantitation of the protein bands from SDS-PAGE failed to detect any significant effects on the amounts of the shifted 10B protein relative to the in-frame 10A protein under all fidelity conditions tested. However, any changes in fidelity conditions led to inhibition of phage morphogenesis in single-step growth experiments, which could not be accounted for by reduced amounts of phage protein synthesis, nor, at least in the case of decreased accuracy, by reduced amounts of phage DNA synthesis. Reduction in phage DNA synthesis did appear to account for a substantial proportion of the reduction in phage yield seen under conditions of increased accuracy. Similar effects of varying ribosomal fidelity on growth were also seen with phage T3, and to a lesser extent with phage T4. The absence of change in the high-frequency T7 gene 10 frameshift differs from earlier reports that ribosomal fidelity affects low-frequency frameshift errors.

Capsid↗
PubMed1991

Munchausen syndrome by proxy: mother fabricates infant's hearing impairment.

Munchausen syndrome by proxy is a form of child abuse in which a mother causes or simulates her child's symptoms and presents the child for diagnosis and treatment. All previously reported cases have involved acute illnesses. This case study describes the ways in which a mother obtained a diagnosis of sensorineural hearing loss as well as amplification for her normally hearing infant.

Female↗
PubMed1991

Recovery of costal and crural diaphragmatic contractility from partial paralysis.

Since the two muscles (costal and crural) that constitute the diaphragm are separate and histologically different, their individual recovery pattern from neuromuscular blockade also may be different. Therefore, we studied the recovery of force and shortening in the in vivo diaphragm from atracurium-induced neuromuscular blockade in seven pentobarbital anesthetized dogs to assess segmental differences. Transdiaphragmatic pressure (Pdi), shortening of costal and crural segments, integrated electromyogram (EMG), and tidal volume (VT) were measured during spontaneous breathing. After atracurium had reduced VT to 30% of control, breathing parameters were followed until recovered to 90% of control values. In addition, force-frequency curves generated by supramaximal tetanic stimuli of the phrenic nerve were measured. Recovery times for tidal Pdi, tidal EMG, tidal shortening, low-frequency shortening, and twitch Pdi were twice as fast as for VT (40 +/- 4 min), reflecting a slower rate of recovery of accessory inspiratory muscles. High-frequency recovery was typically slower than that of VT. During tidal breathing and tetanic stimulation, costal and crural shortening recovered simultaneously. On the other hand, comparison between costal and crural by analysis of pressure-shortening relationships showed a segmental difference (crural shortened 30% more than costal at the same Pdi), which implied reduced afterload on the crural segment. However, since shortening and pressure were linearly related during paralysis and recovery, measurements of Pdi alone can accurately reflect changes in contractile mass when heterogeneity and afterload are controlled.

Animals↗
PubMed1991

Respiratory muscle coordination and diaphragm length during expiratory threshold loading.

Active expiration is produced by the abdominal muscles and the rib cage expiratory muscles. We hypothesized that the relative contribution of these two groups to expiration would affect diaphragmatic length and, hence, influence the subsequent inspiration. To address this question we measured the respiratory muscle response to expiratory threshold loading in spontaneously breathing anesthetized dogs. Prevagotomy, the increase in lung volume (functional residual capacity) and decrease in initial resting length of the diaphragm were attenuated by greater than 50% of values predicted by the passive relationships. Diaphragmatic activation (electromyogram) increased and tidal volume (VT) was preserved. Postvagotomy, effective expiratory muscle recruitment was abolished. The triangularis sterni muscle remained active, and the increase in lung volume was attenuated by less than 15% of that predicted by the passive relationship. Diaphragmatic length was shorter than predicted. VT was not restored, even though costal diaphragmatic and parasternal intercostal electromyogram increased. During expiratory threshold loading with abdominal muscles resected and vagus intact, recruitment of the rib cage expiratory muscles produced a reduction in lung volume comparable with prevagotomy; however, diaphragmatic length decreased markedly. Both the rib cage and abdominal expiratory muscles may defend lung volume; however, their combined action is important to restore diaphragmatic initial length and, accordingly, to preserve VT.

Animals↗
PubMed1991

Analysis of bacteriophage T7 gene 10A and frameshifted 10B proteins.

Bacteriophage T7 capsid protein 10B has previously been proposed to arise by a translational frameshift near the 3' end of the capsid gene 10A coding sequence, adding an additional 53 amino acid residues to the carboxyl-terminal end of the protein. Here we show by peptide mapping experiments as well as by direct partial sequence analysis of an overlapping "junction" peptide, that 10B is in fact related to 10A by a -1 switch in reading frame in a narrow region near the carboxy terminus of 10A. Peptide mapping experiments demonstrate that 10A and 10B have the same amino terminus as well as virtually identical methionine-labeled peptide maps. However, the predicted unique carboxyl-terminal peptide from 10B was also identified. An overlapping peptide was isolated from 10B which spans the junction region in which the proposed translational frameshift is thought to occur. Partial sequencing of this junction peptide confirms a -1 frameshift within the last few codons of 10A.

Amino Acid Sequence↗
PubMed1990

Relationship between protein synthesis and concentrations of charged and uncharged tRNATrp in Escherichia coli.

We have continuously monitored Trp-tRNA(Trp) concentrations in vivo and, in the same cultures, measured rates of protein synthesis in isogenic stringent and relaxed strains. We have also manipulated cellular charged and uncharged [tRNA(Trp)] by two means: (i) the strain used contains a Trp-tRNA synthetase mutation that increases the Km for Trp; thus, varying exogenous Trp varies cellular Trp-tRNA(Trp); and (ii) we have introduced into the mutant strain a plasmid containing the tRNA(Trp) gene behind an inducible promoter; thus, total [tRNA(Trp)] also can be varied depending on length of induction. The use of these conditions, combined with a previously characterized assay system, has allowed us to demonstrate that (i) the rate of incorporation of Trp into protein is proportional to the fraction of tRNA(Trp) that is charged; for any given total [tRNA(Trp)], this rate is also proportional to the [Trp-tRNA(Trp)]; (ii) uncharged tRNA(Trp) inhibits incorporation of Trp into protein; and (iii) rates of incorporation into protein of at least two other amino acids, Lys and Cys, are also sensitive to [Trp-tRNA(Trp)] and are inhibited by uncharged tRNA(Trp). Our results are consistent with models of translational control that postulate modulating polypeptide chain elongation efficiency by varying concentrations of specific tRNAs.

Amino Acyl-tRNA Synthetases↗
PubMed1990

Uncharged tRNA, protein synthesis, and the bacterial stringent response.

Uncharged tRNA has been shown in vivo to have an active role both in the stringent response, and in modulating the rate of translational elongation. Both of these effects appear to be mediated by codon-anticodon interactions on the ribosome. Although the involvement of uncharged tRNA in the stringent response was expected from in vitro experiments, it has only recently been confirmed in vivo. Inhibition of translation by cognate uncharged tRNA was not expected, and a model is proposed in which excess uncharged tRNA competes with charged tRNA (in ternary complex) for the 30S component of the ribosomal A site. When uncharged tRNA is in sufficient excess over charged tRNA, interaction of uncharged tRNA with the 50S component of the A site occurs as well, leading to a stringent response. The cell has a continuum of responses to decreasing aminoacyl-tRNA levels: in moderately limited conditions, the proportion of uncharged tRNA increases, and the translation rate is slowed; under more severe limitations, uncharged tRNA provokes a stringent response, with pleiotropic consequences for the cell.

Bacterial Proteins↗
PubMed1990

Mechanisms of pulsus paradoxus in airway obstruction.

To assess the mechanisms of pulsus paradoxus (i.e., inspiratory decline of greater than or equal to 10 Torr in systolic pressure) in airway obstruction, we studied 12 patients with chronic airflow obstruction before and during breathing through an external resistance that provided loads during both inspiration and expiration. Esophageal pressure (Ppl) and brachial artery pressure, relative to either atmospheric (Pa) or esophageal pressure (Patm), were measured simultaneously during normal and loaded breathing. It was assumed that changes in intrathoracic systemic arterial transmural pressure were adequately represented by Patm. During control, no significant difference between systolic fluctuation (delta Pa) and pleural swings (delta Ppl) was found. Concurrently, inspiratory and expiratory Patm were nearly identical. By contrast, under maximally loaded conditions, higher magnitudes of delta Ppl than delta Pa were found and consequently Patm rose with inspiration. In this connection, the plot of delta Pa against delta Ppl showed that the slopes for delta Ppl less than or equal to 15 Torr (1.2 Torr delta Pa/delta Ppl) and delta Ppl greater than 15 Torr (0.4 Torr delta Pa/delta Ppl) were significantly different. Under all experimental conditions we found during inspiration a rise in diastolic Patm that is consistent with an increase in left ventricular afterload.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1989

Ibuprofen and acetaminophen in the relief of acute pain: a randomized, double-blind, placebo-controlled study.

To determine the relative analgesic efficacy of ibuprofen 400 mg and acetaminophen 1000 mg, we conducted a single-dose, double-blind, placebo-controlled, randomized clinical trial using a standard assay for analgesic agents, the dental pain model. At regular intervals over 6 hours, 184 patients who had undergone dental impaction surgery rated pain intensity and relief on categorical scales and pain half-gone on a dichotomous nominal scale; a categorical overall evaluation was completed at the end of 6 hours. Both active agents were effective compared to placebo. Ibuprofen 400 mg was more effective than acetaminophen 1000 mg for Sum Pain Intensity Difference (SPID), Total Pain Relief (TOTPAR), sum pain half-gone, and overall evaluation (P less than .05 to P less than .001). The time-effect curves demonstrated a greater peak effect and longer duration of action for ibuprofen 400 mg compared to acetaminophen 1000 mg. Side effects were reported in five ibuprofen patients, 11 acetaminophen-treated patients, and seven placebo patients. Based on the results of this clinical study, we conclude that ibuprofen 400 mg is a safe and more effective analgesic than acetaminophen 1000 mg for patients with acute pain.

Acetaminophen↗
PubMed1989

Absence of BK virus sequences in transformed hamster cells transfected by human tumor DNA.

In an attempt to gain insight into the mechanism of oncogenic transformation by BK virus (BKV), a human papovavirus, we have probed for BKV sequences in transformed hamster cells in which oncogenic transformation had occurred as a result of transfection by human tumor DNA positive for BKV sequences. Even though the sources of the transfecting DNA contained BKV sequences, the transformed hamster cells which arose from the transfection for the most part did not retain BKV sequences. In only one barely detectable case was BKV-specific DNA found associated with chromosomal DNA, and in only a small minority of the transformed cells was BKV DNA detected in the Hirt supernatant, indicating an episomal configuration. Even in these few cases where BKV sequences were present in an episomal form, altered migration on gels of some BKV-positive bands (compared to bands derived from cloned viral DNA) suggested deletions and rearrangements of BKV DNA. We employed several different probe methodologies for these studies, including nick-translation, random primer and a non-isotopic biotinylated probe which gave a sensitivity that could detect better than 0.01 copy of viral genome per diploid cell. We conclude that transformation by transfection with human tumor DNA does not require persistence of the BKV viral genome, suggesting that either BKV virus was irrelevant to original oncogenesis, in analogy with models proposed by others for herpesvirus oncogenesis.

Animals↗
PubMed1989

AIDS and children: some of the issues in haemophilia care and how to address them.

Haemophilia is a life-long, life-threatening inherited condition for which there is treatment but no cure. It affects not only the patient but all members of the family. This paper focuses on how AIDS/HIV, which shares these characteristics with haemophilia, has affected children with HIV, their siblings, those who have remained antibody negative and those with HIV infected fathers. The impact of haemophilia and AIDS on the family is considered and the ripple effect of AIDS on health care staff is addressed.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Treatment dilemmas for HIV infected haemophiliacs.

A description of a structured group, held for anti-HIV positive haemophiliacs and their relatives, to discuss the implications of participating in the MRC/INSERM double-blind control trial of zidovudine is given. The group discussion is used to define, evaluate and rank the many treatment dilemmas inherent in participating in this drug trial.

Double-Blind Method↗
PubMed1989

Effect of variation of charged and uncharged tRNA(Trp) levels on ppGpp synthesis in Escherichia coli.

We introduced into a stringent Escherichia coli tryptophan auxotroph a plasmid bearing the tRNA(Trp) gene under the control of an inducible promoter. This allows us to manipulate the total concentration of tRNA(Trp) in the cell according to whether and when inducer is added to the culture. We also manipulated the concentration of Trp-tRNA(Trp) in vivo since the strain used bears a mutation in the Trp-tRNA synthetase affecting the Km for tryptophan, such that varying the exogenous concentration of tryptophan led to variation in the level of Trp-tRNA(Trp) in the cell. With this system, we found that the signal eliciting ppGpp synthesis during a stringent response triggered by tryptophan limitation did not depend on the absolute concentration of either charged or uncharged tRNA(Trp) but rather depended on a decline in the ratio of charged/uncharged tRNA(Trp). In addition, we found that the amplitude of the response, once triggered by tryptophan limitation, was determined by the total concentration of tRNA(Trp) present in the cell (which is mostly uncharged at that point in time). However, excess uncharged tRNA(Trp) did not amplify ppGpp synthesis triggered by limitation of a different amino acid. These data provide in vivo support for the in vitro-derived model of ppGpp synthesis on ribosomes.

Bacterial Proteins↗
PubMed1988

Atypical malignant hyperthermia with persistent hyperkalaemia during renal transplantation.

A 35-year-old 110 kg male developed marked hyperkalaemia, hyponatraemia, hypercapnia and hyperthermia during living-related renal transplantation under anaesthesia with oxygen-nitrous oxide, isoflurane and muscle relaxation with atracurium. This is the first report of successfully treated malignant hyperthermia triggered by isoflurane during renal transplantation with early appearance and persistent (to 12 hours after surgery) electrolyte abnormalities.

Adult↗