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Biomedical subjects

B Ferguson

Publications and source records attributed to B Ferguson.

At least 109 records · Page 6Linked to original sources
PubMed1985

Interactions of aminoacyl-tRNA synthetases in high-molecular-weight multienzyme complexes from rat liver.

The functional interaction of Arg-, Ile-, Leu-, Lys- and Met-tRNA synthetases occurring within the same rat liver multienzyme complex are investigated by examining the enzymes catalytic activities and inactivation kinetics. The Michaelis constants for amino acids, ATP and tRNAs of the dissociated aminoacyl-tRNA synthetases are not significantly different from those of the high-Mr multienzyme complex, except in a few cases where the Km values of the dissociated enzymes are higher than those of the high-Mr form. The maximal aminoacylation velocities of the individual aminoacyl-tRNA synthetases are not affected by the presence of simultaneous aminoacylation by another synthetase occurring within the same multienzyme complex. Site-specific oxidative modification by ascorbate and nonspecific thermal inactivation of synthetases in the purified rat liver 18 S synthetase complex are examined. Lys- and Arg-tRNA synthetases show remarkably parallel time-courses in both inactivation processes. Leu- and Met-tRNA synthetases also show parallel kinetics in thermal inactivation and possibly oxidative inactivation. Ile-tRNA synthetase shows little inactivation in either process. The oxidative inactivation of Lys- and Arg-tRNA synthetases can be reversed by addition of dithiothreitol. These results suggest that synthetases within the same high-Mr complex catalyze aminoacylation reactions independently; however, the stabilities of some of the synthetases in the multienzyme complex are coupled. In particular, the stability of Arg-tRNA synthetase depends appreciably on its association with fully active Lys-tRNA synthetase.

Amino Acids↗
PubMed1985

Isolation and analysis of an Abelson murine leukemia virus-encoded tyrosine-specific kinase produced in Escherichia coli.

A segment of the coding sequence of the Abelson murine leukemia virus transforming gene (v-abl) has been inserted into a plasmid vector that allows its efficient and regulated expression in Escherichia coli. The product of the v-abl-derived coding sequence, designated p60v-abl, accumulated to a level of approximately 10% of total E. coli protein. A procedure is described for the isolation of p60v-abl from E. coli that yields about 50 micrograms of p60v-abl/g wet weight of E. coli. p60v-abl was capable of autophosphorylation and phosphorylating certain E. coli proteins specifically at tyrosine residues. The E. coli-expressed p60v-abl specifically phosphorylated tyrosine residues on casein and angiotensin II. The Km and Vmax values for ATP, casein, and angiotensin II in the p60v-abl kinase reaction have been determined and compared to values reported for other tyrosine-specific kinases. The expression system and isolation procedure described here permit the preparation of functional p60v-abl in quantities sufficient for detailed physical and biochemical characterization and examination of its biological action(s).

Abelson murine leukemia virus↗
PubMed1985

Mapping of functional domains in adenovirus E1A proteins.

We have modified the E1A gene of human subgroup C adenovirus by introducing deletions in its coding sequence. Various truncated E1A proteins were expressed in Escherichia coli, purified, and microinjected via glass capillaries into Vero cells. We monitored their movement from the cell cytoplasm to the nucleus and their ability to induce expression of H5dl312, an adenovirus E1A deletion mutant. Our results show that the carboxyl terminus of E1A contains sequences essential for rapid and efficient nuclear localization. Essential information for efficient H5dl312 complementation is contained in an internal region, comprising sequences of both exons of the E1A gene. A first exon-encoded region, however, is sufficient to induce low levels of adenovirus gene expression. Information for nuclear localization and for H5dl312 complementation are therefore encoded by distinct domains of the E1A gene. In addition, we determined that the human c-myc product was unable to complement H5dl312.

Adenoviridae↗
PubMed1985

A first exon-encoded domain of E1A sufficient for posttranslational modification, nuclear-localization, and induction of adenovirus E3 promoter expression in Xenopus oocytes.

The purified Escherichia coli-expressed human subgroup C adenovirus E1A 13S mRNA product induces expression from the adenovirus type 5 E3 promoter when injected into Xenopus oocytes. In the present communication, the E. coli-expressed E1A 13S and 12S mRNA products are shown to undergo a posttranslational modification in microinjected Xenopus oocytes, which causes a 2- to 4-kDa increase in apparent molecular size, identical to that occurring in HeLa cells expressing the E1A gene. The E. coli-expressed E1A proteins are similarly modified in vitro in a soluble rabbit reticulocyte lysate. The modified form of the E1A proteins preferentially localizes to the oocyte nucleus following cytoplasmic microinjection. The use of various deleted forms of E1A protein synthesized in E. coli shows that a first exon-encoded domain of E1A, residing between amino acid residues 23 and 120, is sufficient for the posttranslational modification and nuclear localization of E1A and also for the trans-activation of the E3 promoter by E1A in Xenopus oocytes. These results suggest that the posttranslational modification of E1A protein may be important for its function.

Acetyltransferases↗
PubMed1985

E1A 13S and 12S mRNA products made in Escherichia coli both function as nucleus-localized transcription activators but do not directly bind DNA.

We previously purified and characterized functionally the Escherichia coli-expressed product of the human subgroup C adenovirus E1A 13S mRNA (B. Ferguson, N. Jones, J. Richter, and M. Rosenberg, Science 224:1343-1346, 1984; B. Krippl, B. Ferguson, M. Rosenberg, and H. Westphal, Proc. Natl. Acad. Sci. USA 81:6988-6992, 1984). We have now expressed in E. coli and purified the protein product encoded by the human subgroup C adenovirus E1A 12S mRNA and have compared the functional properties of this protein with those of the E1A 13S mRNA product. Using microinjection techniques to introduce these proteins into mammalian cells, we found that the E1A 12S mRNA product, like the 13S mRNA product, localized rapidly to the cell nucleus and induced adenovirus gene expression. Although both E1A gene products localized to the nucleus and stimulated adenovirus gene transcription, these proteins did not directly bind to DNA under conditions in which a known DNA-binding protein, the human c-myc gene product, bound DNA efficiently. Thus, the E1A and myc gene products, which have been related both structurally and functionally, exhibit distinctly different biochemical properties.

Adenovirus Early Proteins↗
PubMed1984

Adenovirus E1a gene product expressed at high levels in Escherichia coli is functional.

The human type C adenovirus E1a 13S messenger RNA encodes a gene product, that positively regulates the transcription of viral genes and certain cellular genes and is involved in the transformation of primary mammalian cells. The E1a gene product was expressed at high levels in Escherichia coli. In a Xenopus oocyte microinjection assay, the purified Escherichia coli-produced protein activated the E1a-responsive adenovirus E3 promoter and functioned as efficiently as the E1a gene itself.

Adenoviruses, Human↗
PubMed1984

Functions of purified E1A protein microinjected into mammalian cells.

We have purified the human type C adenovirus E1A 13S mRNA gene product, expressed in Escherichia coli, and demonstrate that the protein exhibits genuine viral functions upon microinjection into mammalian cells. We show that the E1A protein activates expression of the adenovirus E2A gene and induces expression from the major late transcription unit of the adenovirus E1A deletion mutant, H5dl312. We use this functional assay to examine the stability of E1A protein microinjected into cells and find that E1A exhibits full function for at least 18 hours after its injection. In addition, the purified E1A protein was used to generate a high-titer monospecific rabbit antiserum. This antiserum was used to detect and localize E1A proteins within adenovirus-infected cells as well as within microinjected cells. The E1A protein is found to rapidly and quantitatively localize to the cell nucleus following microinjection into the cell cytoplasm. Thus, nuclear localization is an intrinsic property of the E1A polypeptide. The ability of the E1A protein to localize to the cell nucleus and to induce expression from the H5dl312 major late transcription unit is shown to be highly heat stable.

Adenoviruses, Human↗
PubMed1983

Efficient expression of influenza virus NS1 nonstructural proteins in Escherichia coli.

RNA segment 8 of the influenza A virus genome codes for two nonstructural proteins, NS1 and NS2, for which the functions are unknown. Cloned cDNA copies of this gene from three different influenza A virus strains were inserted into an Escherichia coli plasmid expression vector, pAS1, carrying the strong regulatable lambda phage promoter, PL. After induction, the NS1 proteins were overproduced to levels of 20-25% of total cellular protein. This was surprising in that the codon composition for these eukaryotic genes is similar to that for weakly expressed proteins in E. coli. Thus, under the appropriate conditions, it appears that high level expression of genes containing a relatively large proportion of minor codons can be obtained. The NS1 protein produced in bacteria from a cloned cDNA copy of the A/PR/8/34 virus NS gene was purified to apparent homogeneity and used to generate a high-titer monospecific rabbit antiserum. Immunoprecipitation studies showed this antibody to be crossreactive against the NS1 proteins produced by several different influenza A virus strains. Immunofluorescence experiments in Madin-Darby canine kidney cells showed the NS1 proteins to be located in the nucleoplasm early in infection for all strains examined. With some of the strains, NS1-specific immunofluorescence was observed predominantly in the nucleoli later in infection. This technology can be used to obtain other viral proteins in pure form for structural, functional, and immunological studies.

Animals↗
PubMedDate not supplied

Transaction costs, externalities and information technology in health care.

This paper discusses some of the economic issues which underpin the rationale for investment in information and communications technologies (ICTs). Information imperfections lead to significant transaction costs (search, negotiating and monitoring) which in turn confer a negative externality on parties involved in exchange. This divergence in private and social costs leads to a degree of resource misallocation (efficiency loss) which, uncorrected, results in a sub-optimal outcome. Traditional solutions to this problem are to rely upon direct government action to reduce the costs of transacting between market agents, or to employ tax/subsidy measures and other legislative action to achieve the desired market outcome. Three key policy questions are raised in the context of the NHS purchaser/provider relationship. Firstly, what is the optimum level of transaction costs; secondly, can ICTs assist in lowering the level of transaction costs to the optimum level; thirdly, who should bear the investment cost in reducing the level of transaction costs? The issue of property rights in different information systems is discussed and raises interesting policy questions about how much investment should be undertaken centrally rather than devolved to a more local level. In some ways this economic framework offers a post hoc justification of why different ICT systems have been introduced at various levels of the NHS. Essentially this reduces to the problem of externalities: providing good information confers a positive externality: not providing relevant, timely and accurate information confers a negative externality, by increasing further the level of transaction costs. The crucial role which ICT systems can play lies in attempting to reduce the level of transaction costs and driving the market towards what Dahlman has described as the transaction-cost-constrained equilibrium.

Costs and Cost Analysis↗
PubMedDate not supplied

Expenditure on physicians' services in Canada: was Medicare a structural change?

This paper uses cointegration analysis to investigate the question of whether the introduction of national health insurance for physicians' services (Medicare) in the late 1960s caused a structural change in the relation driving national expenditure on physicians' services in Canada. The results support the existence of a single cointegration equation applying to both the Medicare and pre-Medicare periods. Structural change tests do not support the presence of a fundamental structural change associated with the introduction of Medicare.

Bias↗
PubMedDate not supplied

Seven-year follow-up of speech/language-impaired and control children: speech/language stability and outcome.

OBJECTIVE: This study examined the 7-year outcome of speech/language (S/L) impaired and control children selected from a community sample at age 5 years. METHOD: Two hundred fifteen children completed a variety of speech and language tests at age 12 years. Children with S/L impairment were further classified as "speech only," "language only," or "speech and language impaired." RESULTS: More than 72% of children who had S/L impairment at age 5 remained impaired at age 12. Children with both speech and language problems were most likely to remain S/L impaired; 81% had some kind of S/L impairment at follow-up. Similarly, children with both expressive and receptive language impairment were more likely to show expressive or receptive impairment at follow-up than children with expressive impairment alone. One third of time 1 controls had S/L problems at follow-up, and of these 82% had speech impairment only. CONCLUSIONS: S/L impairment identified at age 5 has long-lasting effects. More pervasive problems were associated with poorer outcomes. Screening at age 5 may be useful, as most serious S/L problems that emerged by middle childhood could be identified at age 5. The effects of S/L treatment require further study.

Adolescent↗
PubMedDate not supplied

A model of mixed-market healthcare delivery: the British Columbia Transplant Society.

Driven by the twin imperatives of cost and quality, healthcare providers must innovate and adapt new management models which continuously improve performance. Focusing specifically on the British Columbia Transplant Society (BCTS), this article describes a successful mixed-market management model for delivering healthcare services to a relatively well-defined patient population which is suffering from chronic disease and has predictable healthcare needs. The BCTS is an example of a healthcare organization which combines the best aspects of both the Canadian and American healthcare systems.

British Columbia↗