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Biomedical subjects

K A Edwards

Publications and source records attributed to K A Edwards.

29 records · Page 2Linked to original sources
PubMed1985

Purification and characterization of a type II DNA topoisomerase from bovine calf thymus.

We report here the large scale purification of DNA topoisomerase II from calf thymus glands, using the unknotting of naturally knotted P4 phage DNA as an assay for enzymatic activity. Topoisomerase II was purified more than 1300-fold as compared to the whole cell homogenate, with 22% yield. Analysis of the purified enzyme by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed two bands of apparent molecular masses of 125 and 140 kDa. Tryptic maps of the two bands indicated that they derive from the same protein. Using these fragments, specific polyclonal antisera to topoisomerase II were raised in rabbits. Immunoblotting of whole cell lysates from various species indicated that topoisomerase II is well conserved among mammals and has a native subunit molecular mass of 180 kDa. Analytical sedimentation and gel filtration were used to determine a sedimentation coefficient of 9.8 S and a Stokes radius of 68 A. The calculated solution molecular mass of 277 kDa implies a dimer structure in solution. The purified topoisomerase II unknots P4 DNA in an ATP-dependent manner and is highly stimulated in its relaxation activity by ATP. A DNA-stimulated ATPase activity, as has been found with other type II topoisomerases, is associated with the purified enzyme. Approximate kinetic parameters for the ATPase reaction were determined to be: a Vmax of 0.06 nmol of ATP/(micrograms of protein) (min) and Km of 0.2 mM in the absence of DNA, and a Vmax of 0.2 nmol of ATP/(micrograms of protein) (min) and Km of 0.4 mM ATP in the presence of supercoiled plasmid DNA.

Adenosine Triphosphate↗
PubMed1985

Inverse effects of ethidium bromide on superoxide dismutase and lactate dehydrogenase of Artemia salina embryos.

Development of Artemia salina embryos in the presence of ethidium bromide, an inhibitor of mitochondrial transcription, results in a dose dependent increase in the specific activity of lactate dehydrogenase, and a concomitant decrease in the specific activity of a cyanide-resistant superoxide dismutase. The inhibition of mitochondrial function by ethidium bromide appears to exert opposite effects on the nuclear cistrons encoding lactate dehydrogenase and superoxide dismutase, and suggests that a common mitochondrial signal may exert diametric effects on nuclear cistrons whose products are characteristic of alternate states of respiration.

Animals↗
PubMed1982

Recognition sites of eukaryotic DNA topoisomerase I: DNA nucleotide sequencing analysis of topo I cleavage sites on SV40 DNA.

Eukaryotic DNA topoisomerase I introduces transient single-stranded breaks on double-stranded DNA and spontaneously breaks down single-stranded DNA. The cleavage sites on both single and double-stranded SV40 DNA have been determined by DNA sequencing. Consistent with other reports, the eukaryotic enzymes, in contrast to prokaryotic type I topoisomerases, links to the 3'-end of the cleaved DNA and generates a free 5'-hydroxyl end on the other half of the broken DNA strand. Both human and calf enzymes cleave SV40 DNA at the identical and specific sites. From 827 nucleotides sequenced, 68 cleavage sites were mapped. The majority of the cleavage sites were present on both double and single-stranded DNA at exactly the same nucleotide positions, suggesting that the DNA sequence is essential for enzyme recognition. By analyzing all the cleavage sequences, certain nucleotides are found to be less favored at the cleavage sites. There is a high probability to exclude G from positions -4, -2, -1 and +1, T from position -3, and A from position -1. These five positions (-4 to +1 oriented in the 5' to 3' direction) around the cleavage sites must interact intimately with topo I and thus are essential for enzyme recognition. One topo I cleavage site which shows atypical cleavage sequence maps in the middle of a palindromic sequence near the origin of SV40 DNA replication. It occurs only on single-stranded SV40 DNA, suggesting that the DNA hairpin can alter the cleavage specificity. The strongest cleavage site maps near the origin of SV40 DNA replication at nucleotide 31-32 and has a pentanucleotide sequence of 5'-TGACT-3'.

Animals↗
PubMed1982

Intra- and intermolecular strand transfer by HeLa DNA topoisomerase I.

The major type I DNA topoisomerase (topo I) has been purified from HeLa cell nuclei to a homogeneous, monomeric protein (Mr = 100,000). Similar to the nicking-closing enzyme (Mr = 67,000) from rat liver (Been, M. D., and Champoux, J. J. (1981) Proc. Natl. Acad. Sci. U. S. A. 78, 2883-2887), HeLa topo I has the following properties: (a) HeLa topo I breaks down single-stranded DNA to smaller fragments, each with an enzyme-linked 3'-phosphoryl end and a free 5'-OH end. This cleavage is not dependent upon protein denaturant or protease treatment. (b) HeLa topo I produces single-stranded DNA circles from linear single-stranded DNA. Such DNA circles are believed to be produced by the intramolecular cyclization of topo I-linked, single-stranded DNA fragments. (c) HeLa topo I-linked, single-stranded fragments (donors) can join covalently to double-stranded DNA possessing a 5'-OH group (acceptors). The donor is transferred to the 5'-OH end of the acceptor, independent of the position of the end (internal nick or end of linear DNA) or the configuration of the end (flush, 5'-protruding, or 5'-recessed end) of the acceptor. (d) HeLa topo I cleavage of single-stranded DNA is site-specific, but no special sequence at the ends of the acceptor molecule is apparently required for a successful heterologous strand transfer. These results suggest that HeLa topo I may be involved in DNA sequence rearrangements in addition to its possible role as a swivelase for transcription and replication.

DNA Topoisomerases, Type I↗
PubMed1978

An index for assessing weight change in children: weight/height ratios.

Weight and height data from two studies were recomputed, and original data were computed using a Weight Index formula that accounts for the interaction of actual weight/height changes in growing children and compares this ratio with that of normed weight/height ratios for equal-aged children. Recomputing the data of one study showed that the Weight Index is a more sensitive procedure for assessing long-term behavioral treatment of obesity for girls than weight alone. Recomputing the data of a second study showed that the Weight Index formula is consistent for boys and girls and is a more sensitive procedure for assessing long-term effects of a variety of medical treatments for obesity than the Ponderal Index or weight alone. Computation of the data for 17 "normal" children in a preschool class showed a zero Weight Index score before and after a six-month interval elapsed without treatment. The procedure may be useful in assessing ponderosity or anorexia over intervals of six months or more with growing children or difference between actual and normed weight over shorter intervals.

Adolescent↗
PubMedDate not supplied

Informed decision making in outpatient practice: time to get back to basics.

CONTEXT: Many clinicians have called for an increased emphasis on the patient's role in clinical decision making. However, little is known about the extent to which physicians foster patient involvement in decision making, particularly in routine office practice. OBJECTIVE: To characterize the nature and completeness of informed decision making in routine office visits of both primary care physicians and surgeons. DESIGN: Cross-sectional descriptive evaluation of audiotaped office visits during 1993. SETTING AND PARTICIPANTS: A total of 1057 encounters among 59 primary care physicians (general internists and family practitioners) and 65 general and orthopedic surgeons; 2 to 12 patients were recruited from each physician's community-based private office. MAIN OUTCOME MEASURES: Analysis of audiotaped patient-physician discussions for elements of informed decision making, using criteria that varied with the level of decision complexity: basic (eg, laboratory test), intermediate (eg, new medication), or complex (eg, procedure). Criteria for basic decisions included discussion of the nature of the decision and asking the patient to voice a preference; other categories had criteria that were progressively more stringent. RESULTS: The 1057 audiotaped encounters contained 3552 clinical decisions. Overall, 9.0% of decisions met our definition of completeness for informed decision making. Basic decisions were most often completely informed (17.2%), while no intermediate decisions were completely informed, and only 1 (0.5%) complex decision was completely informed. Among the elements of informed decision making, discussion of the nature of the intervention occurred most frequently (71 %) and assessment of patient understanding least frequently (1.5%). CONCLUSIONS: Informed decision making among this group of primary care physicians and surgeons was often incomplete. This deficit was present even when criteria for informed decision making were tailored to expect less extensive discussion for decisions of lower complexity. These findings signal the need for efforts to encourage informed decision making in clinical practice.

Colorado↗
PubMedDate not supplied

Research methodology. Part IV: Understanding canonical correlation analysis.

Canonical correlation is presented as a technique to determine how sets of dependent variables are related with sets of independent variables. Canonical correlation reveals the strength of the relationship between the clusters using case data as illustration, three pairs of clusters (factors or profiles) emerged. Interpretation of the clusters are presented. As indicated in the case presentation, Canonical Correlation (CA) is the fourth in a series of methodologies selected for illustration as precursors to advanced statistics and modeling. In this paper, background will be given, a schematic example presented, sample size and CA, SPSS procedure to perform CA, and interpretation of CA and possible uses of CA in nursing research.

Analysis of Variance↗