PubMed Health⌕ Search

Biomedical subjects

W T Garrard

Publications and source records attributed to W T Garrard.

At least 73 records · Page 4Linked to original sources
PubMed1979

Overall pathway of mononucleosome production.

Five electrophoretically distinguishable classes of mononucleosomes (MI, MII, ...MV) are produced upon treatment of mammalian nuclear chromatin with micrococcal nuclease. These five forms differ in their initial DNA lengths, relative mass proportions, stability, contents of histone H1, and presence of certain nonhistone proteins. A new "chromatin fingerprinting" technique has been developed in order to trace nuclease-mediated interconversions between these mononucleosomes and their polynucleosomal precursors. Application of this technique, together with earlier findings from this laboratory, has made possible the elucidation of the overall pathway of nuclease cleavage of chromatin which leads to the production and interconversion of these mononucleosomes, and has permitted reconstruction of the organization of these mononucleosomes in undigested chromatin...

Animals↗
PubMed1979

Histone molar ratios among different electrophoretic forms of mono- and dinucleosomes.

The relative molar ratios of each of the histone classes and protein A24 have been determined in nuclei, chromatin, and different electrophoretic forms of mono- and dinucleosomes of cultured mouse cells. For this purpose, [3H]lysine- and [14C]arginine-labeled cells were used for sample preparations, and stoichiometries were estimated from protein radioactivity profiles and known amino acid compositions following sodium dodecyl sulfate (SDS)-gel electrophoresis. The results demonstrate that upper limits of one and two histone H1 molecules exist per mono- and dinucleosome, respectively. However, isolated nuclei contain less than one copy of histone H1 per nucleosome. In addition, among the chromatin subfractions studied, histones H3, H2B, and H4 are essentially equimolar, while histone H2A is less than equimolar by 19 +/- 9%. This latter finding offers direct support to the proposal of Goldknopf (Goldknopf, I. L., French, M. F., Musso, R., and Busch, H. (1977) Proc. Natl. Acad. Sci. U. S. A. 74, 5492-5495) that protein A24 replaces histone H2A in the octamer protein core of the nucleosome, since about 10% of the total histone H2A of cultured mouse cells is in the form of protein A24 and is present in nucleosomes. From the results of the present study, it is concluded that electrophoretic fractionation of mono- and dinucleosomes is not due to variable molar ratios or amounts of the four smaller histone classes, but depends on part on DNA length, the number of associated histone H1 molecules, and non-histone chromosomal proteins.

Arginine↗
PubMed1977

Heterogeneity in nucleosome spacing.

The organization of spacer DNA connecting 160 base-pair cores of bovine thymus polynucleosomes has been studied by a combination of biochemical and electron microscopic techniques. The results reveal that a major fraction of chromatin consists of a spectrum of repeating units; these differ from each other by up to 20 base-pairs due to variation in spacer DNA length. Those polynucleosomes which have larger spacer DNA lengths are processed by micrococcal nuclease to mononucleosomes more rapidly than those organized with shorter spacers. Although a distribution of spacer DNA lengths exists, a significant proportion of spacers that are nearest neighbors share common DNA lengths. These findings imply that functional roles may be related to the manners in which spacers are organized along chromatin fibers.

Animals↗
PubMed1974

Partial purification of the template-active fraction of chromatin: a preliminary report.

A fraction of rat-liver chromatin that is transcriptionally active in vivo has been purified 6- to 7-fold over whole chromatin. This was accomplished by selectively shearing chromatin with DNase II followed by fractionating the released portion on the basis of its solubility properties in 2 mM MgCl(2). The resulting soluble material comprises 11% of the total chromatin DNA and is impoverished in histone and enriched in nonhistone protein. Compared with unsheared chromatin, this minor fraction exhibits marked differences in chromosomal protein species. DNA renaturation studies indicate that this fraction is composed of a specific subset of whole genomal DNA sequences. Furthermore, DNA.RNA hybridization experiments suggest that almost 60% of the nonrepetitious DNA sequences of this minor fraction could code for cellular RNA.

Animals↗
PubMed1971

Selective release of proteins from Spirillum itersonii by tris (hydroxymethyl) aminomethane and ethylenediaminetetraacetate.

Treatment of Spirillum itersonii with tris(hydroxymethyl)aminomethane (Tris)-ethylenediaminetetraacetate (EDTA) results in the quantitative release of alkaline phosphatase and ribonuclease into the surrounding medium. At the same time, about 90% of the total cellular soluble cytochrome c is liberated. This process occurs within 1 min of treatment at both 24 and 4 C. Release of these proteins by Tris-EDTA treatment is highly selective, since only 9% of the total cell protein is liberated, concomitantly with less than 5% ribonucleic acid, deoxyribonucleic acid, and malate dehydrogenase. Different sigmoidal curves are obtained for release of proteins as a function of EDTA concentration. The order of liberation with increasing EDTA is as follows: alkaline phosphatase, protein, soluble cytochrome c, and ribonuclease. Treatment of cells with Tris-EDTA under conditions which cause extensive loss of alkaline phosphatase, soluble cytochrome c, and ribonuclease results in cell death, with cessation of protein and ribonucleic acid synthesis. Cells treated with EDTA in phosphate buffer (in the absence of Tris) liberate a large portion of their soluble cytochrome c, but negligible amounts of alkaline phosphatase and ribonuclease. Addition of Tris to cells pretreated with phosphate-buffered EDTA releases high levels of alkaline phosphatase, but not ribonuclease. These results suggest that a common surface alteration is not solely responsible for release of periplasmic proteins. More likely, each protein of the periplasm is bound in an independent and specific manner.

Acrylates↗
PubMed1970

Nitrate reductase and soluble cytochrome c in Spirillum itersonii.

The addition of nitrate to cultures of Spirillum itersonii incubated under low aeration produced a diauxic growth pattern in which the second exponential phase was preceded by the appearance of nitrite in the medium. The organism also grew anaerobically in the presence of nitrate. Nitrate reductase activity could be demonstrated in cell-free extracts by use of reduced methyl viologen as the electron donor. The enzyme was located in the supernatant fraction after centrifugation of extracts for 2 hr at 40,000 x g, and it sedimented as a single peak when centrifuged in a sucrose gradient. Nitrate reductase activity was found in cells grown with low aeration without nitrate, but was increased about twofold by addition of nitrate. Enzyme activity was negligible in cells grown with high aeration. The proportion of soluble cytochrome c was increased two- to threefold in cells grown with nitrate. The specific activities of nitrate reductase and soluble cytochrome c rose when nitrate or nitrite was added to cell suspensions incubated with low aeration; nitrite was more effective than nitrate during the early stages of incubation. A nitrate reductase-negative mutant synthesized increased amounts of soluble cytochrome c in response to nitrate or to nitrite in the cell suspension system. It is concluded that enhanced synthesis of soluble cytochrome c does not require the presence of a functional nitrate reductase.

Cell-Free System↗