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R C Warner

Publications and source records attributed to R C Warner.

At least 19 recordsLinked to original sources
PubMed1996

Preparation of figure 8 and cruciform DNAs and their use in studies of the kinetics of branch migration.

We have re-examined the kinetics of the branch migration of double-stranded DNA that is mediated by the stepwise movement of the Holliday junction. This work revises and extends our previous treatment (Thompson, B. J., Camien, M. N., and Warner, R. C. (1976) Proc. Natl. Acad. Sci. U.S.A. 73, 2299-2303). New methodology and new highly purified substrates have been used. The latter include figure 8s prepared from phage G4 DNA by annealing single-stranded components and two sizes of a novel cruciform. We treat the process as a one-dimensional diffusion based on the random walk, the mathematical basis of which is discussed in detail. The step rate is shown to be 3 orders of magnitude slower than we reported previously. The most important contribution to the erroneously high rate was a result of the presence of EDTA in the spreading solution used for electron microscopy at that time. A second contribution of about 4-fold resulted from catalysis by EcoRI and other proteins. The rates reported here are for the uncatalyzed reaction.

Base Sequence↗
PubMed1986

Denaturation of covalently closed circular DNA. Kinetics, comparison of several DNAs, mechanism and ionic effects.

The rates of the alkaline denaturation of the covalently closed, circular DNAs (form I) of the replicative forms (RF) of phages G4, phi X174, and fd, and of plasmid pBR322 and phage PM2 have been measured at 0 degrees C and some at higher temperatures. These rates are orders of magnitude slower than the denaturation of linear DNA because of the increased stability of the helix to deprotonation that results from the accumulating positive superhelicity during denaturation. Denaturation reactions were initiated by rapid, infrasonic mixing (Camien, M.N., and Warner, R.C. (1984) Anal. Biochem. 138, 329-334), and their progress was measured by analytical ultracentrifugal analysis for the amounts of form I and denatured (Id) DNA after neutralization of the alkaline reaction. The comparative rates of the five DNAs varied over a wide range; the fastest, G4-RF, denatured at 500-fold the rate of the slowest, fd-RF. The differences are accounted for by the interaction of positive superhelicity with the sequence-dependent regions of relative helix stability in the various DNAs. Renaturation rates of Id DNAs varied similarly for Ids prepared at 0 degrees C, but only a few-fold for Ids prepared at 50 degrees C. The rate of denaturation of G4-RF was determined over a wide range of NaOH and NaCl concentrations at 0 degrees C, and the pHm was determined as a function of ionic strength and temperature. The effects of ionic strength have been analyzed in an application of the Manning ion condensation-screening theory (Manning, G.S. (1978) Q. Rev. Biophys. 11, 179-246) which is shown to account for the large destablizing effect of salts on the helix. The pH region of transition at 50 degrees C from renaturation to denaturation was examined, and it was found that the maximum rate of renaturation occurred at a pH about 0.05 units below the pHm.

Coliphages↗
PubMed1986

Novel dimeric configurations from bacteriophage G4 replicative form DNA.

The oligomeric fraction of the replicative form of phage G4 was prepared by sedimentation on three successive CsCl velocity gradients followed by resolution on CsCl-propidium diiodide equilibrium gradients and subfractions through the equilibrium gradients were examined by electron microscopy. The most frequent dimer species were the circular dimer, the singly linked catenane and the figure 8; these occurred in a ratio of 10:3:1. The high enrichment for dimers and other oligomers made possible the observation and the determination of the frequency of occurrence of a number of minor species, some of them of novel configuration. These are (a) dimers similar to figure 8s except containing long, apparently four-stranded junctions common to the two halves (theta forms); (2) dimers similar to those in (1) except that the long junctions separate the two halves (dumbbell forms); (3) multiply catenated dimers with apparent right-handed intertwines; and (4) dimers containing a knot. Theta forms cleaved by EcoRI were shown to be stable under conditions in which EcoRI-treated figure 8s were resolved by branch migration.

Coliphages↗
PubMed1984

An apparatus for sequentially combining microvolumes of reagents by infrasonic mixing.

A method employing high-speed infrasonic mixing for obtaining timed samples for following the progress of a moderately rapid chemical reaction is described. Drops of 10 to 50 microliter each of two reagents are mixed to initiate the reaction, followed, after a measured time interval, by mixing with a drop of a third reagent to quench the reaction. The method was developed for measuring the rate of denaturation of covalently closed, circular DNA in NaOH at several temperatures. For this purpose the timed samples were analyzed by analytical ultracentrifugation. The apparatus was tested by determination of the rate of hydrolysis of 2,4-dinitrophenyl acetate in an alkaline buffer. The important characteristics of the method are (i) it requires very small volumes of sample and reagents; (ii) the components of the reaction mixture are pre-equilibrated and mixed with no transfer outside the prescribed constant temperature environment; (iii) the mixing is very rapid; and (iv) satisfactorily precise measurements of relatively short time intervals (approximately 2 sec minimum) between sequential mixings of the components are readily obtainable.

Biochemistry↗
PubMed1984

Effect of consuming fiber from corn bran, soy hulls, or apple powder on glucose tolerance and plasma lipids in type II diabetes.

The effects of consuming corn bran, soy hulls, or apple powder on glucose and lipid metabolism were investigated in two studies of persons with type II diabetes. Fiber sources, completely or partially added to bread, were incorporated into subjects' self-selected diets. Low fiber white bread served as a control. In study A, 10 subjects consumed 26 g fiber source daily; in study B, eight subjects consumed 52 g fiber source. Biochemical tests, including a 2-h postprandial glucose test using a low fiber formula meal, were scheduled after 2 and 4 wk of each dietary treatment. Soy hull consumption slightly improved some measures of glucose tolerance, with results varying between the studies. Consumption of 52 g corn bran decreased very low-density lipoprotein cholesterol, triglycerides, and glycosylated Hb, but subject tolerance was poor with the particle size used. Consumption of 52 g apple powder increased low-density lipoprotein and total cholesterol levels.

Aged↗
PubMed1981

Renaturation of denatured, covalently closed circular DNA.

The rate of renaturation of denatured, covalently closed, circular DNA (form Id DNA) of the phi X174 replicative form has been investigated as a function of pH, temperature, and ionic strength. The rate at a constant temperature is a sharply peaked function of pH in the range of pH 9 to 12. The position on the pH scale of the maximum rate decreases as the temperature is increased and as the ionic strength is increased. The kinetic course of renaturation is pseudo-first order: it is independent of DNA concentration, but falls off in rate from a first order relationship as the reaction proceeds. The rate of renaturation depends critically on the temperature at which the denaturation is carried out. Form Id, prepared at an alkaline pH at 0 degrees C, renatures from 5 to more than 100 times more rapidly than that similarly prepared at 50 degrees C. Both the heterogeneity in rate and the effect of the temperature of denaturation depend, in part, on the degree of supercoiling of the form I DNA from which the form Id is prepared. However, it is concluded that a much larger contribution to both arises from a configurational heterogeneity introduced in the denaturation reaction. The renaturation rate was determined by neutralization of the alkaline reaction and analytical ultracentrifugal analysis of the amounts of forms I and Id. The nature of the proximate renatured species at the temperature and alkaline pH of renaturation was investigated by spectrophotometric titration and analytical ultracentrifugation. It is concluded that the proximate species are the same as the intermediate species defined by an alkaline sedimentation titration of the kind first done by Vinograd et al. ((1965) Proc. Natl. Acad. Sci. U. S. A. 53, 1104-1111). Observations are included on the buoyant density of form Id and on depurination of DNA at alkaline pH values and high temperatures.

Bacteriophage phi X 174↗
PubMed1981

Replication of double-stranded RNA of the virus-like particles in Saccharomyces cerevisiae.

The mode of replication of the L double-stranded RNA (dsRNA) present in virus-like particles in Saccharomyces cerevisiae was examined by density transfer experiments. After transfer to light medium, significant amounts of fully heavy dsRNA persisted over a number of cell doublings. In addition, very little material of hybrid density was ever formed, and the accumulation of fully light material began as early as 0.5 doubling after transfer to light medium. Our results are compatible with a conservative mode of replication or with a semiconservative mode of replication carried out by a small portion of the total dsRNA population. In additional experiments the synthesis of dsRNA relative to the cell cycle was studied. This was done by determining the ratio of short-term to long-term radioactive label in size-separated cell fractions of a prelabeled exponential culture. The ratio of short-term to long-term label remained constant for all fractions, implying that dsRNA is synthesized throughout the cell cycle, increasing through the cell cycle at an exponential rate.

Cell Cycle↗
PubMed1979

Heteroduplexes of phiX174 and G4 DNAs: orientation to genetic map and comparison with predictions from nucleotide sequences.

Heteroduplexes between the viral DNA of phiX174 and DNA from the replicative form (RF) of phage G4 were examined by electron microscopy. The single Eco RI site of G4-RF was utilized as a physical marker by preparing the heteroduplexes from the denatured, linear DNA obtained by restricting G4-RF with Eco RI endonuclease. Restriction fragments of phiX were used in a separate series of heteroduplexes to align the heteroduplex map and the G4 Eco RI site with the similar genetic maps of the two phages. The positions of the branch migrating junctions of recombinant phiX-G4 figure-8s, previously located only with respect to the G4-Eco RI site, have now been located with high proability within the gene A region of the two genomes. The degree of mismatch between the known nucleotide sequences of phi X and G4 accounts for positions of all of the regions of single-strandedness in the observed heteroduplexes, but unexplained discrepancies were also found.

Base Sequence↗