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A Cummings

Publications and source records attributed to A Cummings.

31 records · Page 2Linked to original sources
PubMed1989

Multiple modifications in the phosphoproteins bound to stored messenger RNA in Xenopus oocytes.

Messenger RNA molecules accumulated in amphibian oocytes are stabilized and blocked from translation through association with a defined set of phosphoproteins. Phosphoproteins of 60 kDa and 56 kDa (pp60 and pp56) isolated from messenger ribonucleoprotein particles of Xenopus laevis oocytes can be bound in vitro to mRNA sequences. After phospholabelling in vitro, both pp60 and pp56 show a range of ionic forms, which resolve on two-dimensional gel electrophoresis as a series of pairs with identical charge. The similarities between pp60 and pp56 in their ionic properties suggest a common protein primary structure. This suggestion gains further support from proteinase digestion analysis of pp60 and pp56: practically identical size patterns of phospholabelled fragments are generated using a range of different proteinases. However, in spite of their structural similarities, pp60 and pp56 are recognised as antigenically distinct from each other by using polyclonal antibodies. It is concluded from these, and other, observations that pp60 and pp56 are members of a family of structurally similar polypeptides which are subjected to multiple secondary modifications. Of these modifications, phosphorylation appears to be instrumental in establishing tight binding to mRNA, while antigenicity appears to be determined by some other modification. The role of microheterogeneity in the structure of RNA-binding proteins is discussed in relation to the differential activation of mRNA sequences for translation during early development.

Animals↗
PubMed1988

5-Hydroxymethylcytosine DNA glycosylase activity in mammalian tissue.

The enzymatic release of 5-hydroxymethylcytosine from T2 bacteriophage DNA was effected by an extract of calf thymus. Like the previously described 5-hydroxymethyluracil DNA glycosylase, 5-hydroxymethylcytosine DNA glycosylase was not detectable in bacterial extracts. The phylogenetic distribution of these activities indicates that their primary function is the maintenance of methylcytosine residues in differentiated tissue.

5-Methylcytosine↗
PubMed1988

Protein kinase activity associated with stored messenger ribonucleoprotein particles of Xenopus oocytes.

As the oocytes of Xenopus laevis grow and develop they accumulate vast stores of mRNA for use during early embryogenesis. The stored mRNA is stabilized and may be prevented from being translated in oocytes by the binding of a defined set of oocyte-specific proteins to form messenger RNP (mRNP) particles. A key event in the interaction of protein with mRNA is the phosphorylation of those few polypeptides that bind directly to all classes of polyadenylated mRNA. In this study we show that the phosphorylating enzyme (protein kinase), in addition to its target phosphoproteins, is an integral component of the mRNP particles. This association extends through various stages in the formation and use of the mRNP particles. Examination of material from oocytes of an early developmental stage (early stage 1), when the level of accumulated mRNA is low, reveals an excess of protein particles free of RNA, sedimenting at 6-18 S, and containing protein kinase activity and mRNA-binding phosphoproteins. At stages of maximum rate of mRNA accumulation (stages 1 and 2), the phosphoproteins and kinase are found primarily in individual mRNP particles that sediment at 40-80 S. As ribosomes become abundant (stages 2 and 3), the mRNP particles tend to interact with ribosomal subunits, at least in vitro, to form blocked translation initiation complexes that sediment at 80-110 S. These results are compared with observation on stored mRNP in other developmental systems.

Animals↗
PubMed1987

Phosphorylation of a 60 kDa polypeptide from Xenopus oocytes blocks messenger RNA translation.

The stored mRNP particles of Xenopus oocytes contain protein kinase activity and two major phosphoproteins of 60 kDa (pp60) and 56 kDa (pp56). These proteins can be phospholabelled in the particles either in vivo or in vitro and then isolated by SDS-PAGE. On renaturing pp60 in the presence of globin mRNA, a stable RNA-protein complex is formed. The complex has a uniform density in Cs salt gradients, corresponding to the binding of about 10 protein molecules to each mRNA, probably at the poly(A) sequence. Compared with uncomplexed mRNA, the RNP complex is translated poorly both in vitro and in vivo. Translation of the complex can be regained after treatment with protein phosphatase. It is shown that dephosphorylation destabilizes the binding of protein to RNA, making the mRNA accessible for translation. Studies with native mRNP particles show that their translation also can be enhanced by dephosphorylation.

Animals↗
PubMed1987

Definitive characterization of human thymine glycol N-glycosylase activity.

An N-glycosylase activity that released cis-[3H]-5,6-dihydroxy-5,6-dihydrothymine (thymine glycol, TG) from chemically oxidized poly(dA-[3H]dT) was unambiguously characterized both in extracts of HeLa cells and in purified Escherichia coli endonuclease III. This was accomplished by use of microderivatization procedure that quantitatively converted cis-TG to 5-hydroxy-5-methylhydantoin (HMH). The reaction products were analyzed by high-pressure liquid chromatography before and after derivatization by using cis-[14C]TG and [14C]HMH, which had been independently synthesized, as reference compounds. This technique facilitated construction of a v/[E]t plot for the enzyme activity in HeLa cells, permitting estimation of its specific activity. The results obtained prove the existence of both human and bacterial N-glycosylase activities that effect removal of TG from DNA.

Carbon Radioisotopes↗
PubMed1987

Quantitative measurement of the diastereoisomers of cis thymidine glycol in gamma-irradiated DNA.

A technique for determining the relative content of each of the diastereoisomers of cis thymidine glycol (dTG) in DNA exposed to ionizing radiation has been developed. [3H]thymidine DNA was gamma-irradiated, digested to 2'-deoxyribonucleosides, authentic [14C] (+, -) cis dTG added to the digestate and the mixture resolved by HPLC. 3H fractions coeluting with [14C] (+, -) dTG were collected and acetylated. The acetoxy derivatives of (+) and (-) cis dTG were easily resolved by a second HPLC analysis and their absolute configuration determined by NMR and mass spectroscopies. We have constructed a dose-response curve for formation of each isomer in gamma-irradiated DNA and shown that they are formed in equal amounts. This technique may be used to determine the relative formation of cis dTG isomers in DNA resulting from other oxidative stresses and whether repair of these is influenced by their configuration.

Acetylation↗
PubMed1986

Further abrasion resistance and bond strength studies of fissure sealants.

As part of a laboratory evaluation, measurements of abrasion loss and tensile enamel--sealant bond strengths have been made on three visible light cured sealants (two filled, one unfilled), two chemically cured sealants (one filled and one unfilled) and an experimental sealant with alumina filler particles. A UV-cured filled sealant was used as a control. The unfilled sealants had abrasion loss approximately twice that of the filled commercial sealants. The experimental sealant, with filler content above 50%, had an abrasion loss approximately half that of the filled commercial sealants. The chemical cured sealants had bond strengths lower than the UV-cured control, while the bond strengths of the two visible light-cured filled sealants and the experimental sealant were not statistically different from either the unfilled visible light-cured sealant or from the UV-cured filled sealant.

Animals↗
PubMed1986

Laboratory studies of visible-light cured fissure sealants: setting times and depth of polymerization.

Measurements were made, using an acoustic transmission method, of the setting times of three visible-light curing sealants (two filled, one unfilled). Five commercially available visible-light polymerizing sources were tested with each material. Only three of the light sources gave setting times not significantly different from the manufacturers' recommended times for all three materials tested. The variation in microhardness with depth of sealant was measured to study the depth of polymerization of the sealants, all of which were adequate. The filled sealants had microhardness values greater than the unfilled sealant, at all depths studied.

Chemical Phenomena↗
PubMed1985

Quantitative determination of the 5-(hydroxymethyl)uracil moiety in the DNA of gamma-irradiated cells.

5-(Hydroxymethyl)uracil (HMUra) is a chemically stable derivative of thymine formed through the action of ionizing radiation which we previously identified in the DNA of gamma-irradiated HeLa cells [Teebor, G. W., Frenkel, K., & Goldstein, M. S. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 318-321]. In this report, we determine whether HMUra can be used as a marker of exposure of DNA to ionizing radiation. Dose-response curves for its formation in [3H]thymidine-labeled DNA were constructed by exposing the DNA to increasing amounts of gamma-radiation and measuring the HMUra content. DNA was irradiated both in solution and in intact cells. HMUra was identified as the 2'-deoxyribonucleoside 5-(hydroxymethyl)-2'-deoxyuridine (HMdU) by subjecting the irradiated DNA to enzymatic digestion and analyzing the mixture of 2'-deoxyribonucleosides by high-pressure liquid chromatography. The identity of the radiogenically formed HMdU was confirmed by acetylation and the structure of the acetyl derivative obtained by mass and nuclear magnetic resonance spectroscopies. At two different DNA concentrations in solution, the same number of thymidine moieties were converted to HMdU, indicating that within this range of concentration the formation of HMdU was mediated through the indirect action of ionizing radiation. Equal amounts of HMdU were formed in single- and double-stranded DNA at each radiation dose, indicating that DNA conformation did not affect HMdU formation. Surprisingly, the G value (number of HMdU molecules formed/100 eV) was higher in irradiated cellular DNA than in DNA irradiated in solution.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylation↗
PubMed1983

An assessment of new dental ultraviolet sources and u.v.-polymerized fissure sealants.

The performance of six dental ultraviolet (u.v.) sources was assessed to establish minimum output requirements and possible u.v. hazards. Exposure criteria for u.v.-cured materials (Nuva-seal P.A., Nuva-cote P.A., Nuva-fil P.A., Lee-seal, u.v.-Dentoseal, Uviobond, Visiobond) were assessed by determining setting time-intensity characteristics, u.v. transmission and microhardness as a function of u.v. intensity and depth. A minimum intensity of 10 mW cm-2 was required for adequate curing and both Nuva-seal P.A. and Nuva-cote P.A. set well in depth with realistic exposures. Total u.v. outputs, average intensities and spatial distributions were assessed for the u.v. sources, most of which were well above minimum requirements. Ultraviolet spectra and maximum permissible exposure measurements indicated that all of the sources were safe in normal use.

Chemical Phenomena↗
PubMed1983

Surface treatment studies aimed at streamlining fissure sealant application.

Three laboratory studies were carried out with the aim of improving or streamlining enamel surface treatment procedures in fissure sealing. In Study 1, the effect of reducing acid etch time from 60 s to 10 s was assessed by SEM and tensile bond strength measurements. Reduction in etch time produced a finer etch pattern, gave a reduced but still high, bond-strength, albeit with slightly poorer long-term adhesion. Study 2 showed that a conventional acid etch treatment could completely remove acquired pellicle, without prior prophylaxis. SEM studies and bond-strength measurements corroborated this view. In Study 3, compressed air and chemical drying agents were compared for drying speed and efficiency. Compressed air gave more rapid, thorough drying, and greater resultant bond strengths.

Acid Etching, Dental↗
PubMed1978

Microhardness studies on the setting characteristics of fissure sealants.

The hardness of five fissure sealants was measured using a Wallace Microhardness Tester and found to continue to increase long after the initial setting. The effect of the intensity of radiation and exposure time on the setting of two u.v. activated sealants (Nuva-seal and Alphaseal) was also investigated, with higher intensities producing both a more rapid set and greater final hardness. Alphaseal required longer exposure at a given intensity of u.v. radiation to achieve the same hardness as Nuva-seal, and displayed relatively slow setting in depth, which could give rise to incomplete polymerization during clinical use, resulting in early sealant loss. Thus special care must be taken to ensure that Alphaseal receives an adequate exposure to u.v. radiation, and sources which are suitable for setting Nuva-seal may not be sufficiently powerful when applying Alphaseal.

Acrylates↗