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Biomedical subjects

E H McConkey

Publications and source records attributed to E H McConkey.

At least 37 records · Page 2Linked to original sources
PubMed1979

Human heterozygosity: a new estimate.

Several hundred polypeptides from four human diploid fibroblast cell lines were compared by high-resolution two-dimensional polyacrylamide gel electrophoresis and double-label autoradiography under conditions where allelic products that differ by a single charged amino acid would be distinguished. The average heterozygosity represented by this set of gene products appears to be less than 1% for changes involving charged amino acids.

Black People↗
PubMed1978

Message sequences are not adjacent to poly(A) in heterogeneous nuclear RNA of Friend leukemic cells.

Hybridization of labeled RNA with excess amounts of unlabeled complementary DNA (cDNA) was used to investigate the location of cytoplasmic mRNA sequences in heterogenous nuclear RNA (hnRNA) of noninduced Friend leukemic cells. A heterologous hybrid between hnRNA and cytoplasmic cDNA was formed. Two homologous hybrids were also formed, one between poly(A)-containing mRNA and cytoplasmic cDNA, and the other between poly-(A)-containing hnRNA and nuclear cDNA. All hybrids were selected on hydroxyapatite columns after RNase treatment. The hybrids were further investigated for the presence of poly(A). No poly(A) was found in the heterologous hybrid, while both homologous hybrids contained poly(A). From these results we conclude that there exists a spacer nucleotide sequence between the poly(A) and the message sequences in hnRNA.

Cell Line↗
PubMed1977

Phosphorylation of ribosomal protein S6 in suspension cultured HeLa cells.

HeLa cell ribosomal protein S6, and the increase in its phosphorylation level that occurs after resuspending cells in fresh medium plus serum, were studied using two-dimensional gel electrophoresis. The maximum level of S6 phosphorylation occurs about 2 h after adding fresh medium and seum to cells that have been allowed to grow to high density; this results in an almost complete shift of the spot representing S6 in two-dimensional polacrylamide gels to a new location. Mixing experiments showed that the differences in the level of phosphorylation occur in vivo and are not an artifact of in vitro sample preparation. This method of stimulating S6 phosphorylation provides a convenient system for studying the functional significance of the phenomenon. Only one other ribosomal protein was detectably phosphorylated using [32P]-labeling and autoradiography of dried two-dimensional gels. The level of phosphorylation of this protein, L14, does not change after serum stimulation.

Electrophoresis, Polyacrylamide Gel↗
PubMed1976

Exchange and stability of HeLa ribosomal proteins in vivo.

The relative stabilities of individual HeLa ribosomal proteins and their capacity for exchange between ribosome-bound and -free states in the cytoplasm were examined. Most ribosomal proteins on cytoplasmic ribosomes were found to have uniform, high stability as measured by comparing the short term (12-hour) to steady state (3-day) labeling ratios determined for each ribosomal protein. This would be expected if the proteins in ribosomes either were all stable or were all degraded as a unit. The data do not rule out the possibility that individual proteins have different stabilities prior to their assembly into ribosomes. Four proteins labeled atypically. One large subunit protein (L5) had a lower than average ratio. We interpret this low ratio as being due to a large free pool of this protein. Three proteins (L10, L28, S2) had higher than average ratios, interpreted as being due to reduced protein stability. Two of these proteins (L10, L28) with high ratios were also found to exchange in vivo. The exchangeable proteins may be subject to increased degradation during the time that they spend in the exchangeable free pool. The third protein (S2) with an atypically high ratio is thought to be degraded or altered while on the ribosome, or slowly lost as ribosomes age, because exchange of this protein was not detected. These interpretations and some alternate interpretations are explained. The exchange of three large subunit proteins (L10, L19, L28) was detected by labeling of protein after ribosome synthesis had been inhibited with actinomycin D. Autoradiography of two-dimensional polyacrylamide gels showed labeling of these spots.

Amino Acids↗
PubMed1976

Relative stoichiometry in ribosomal proteins in HeLa cell nucleoli.

Total protein was released from isolated HeLa cell nucleoli by guanidine hydrochloride, purified by cesium chloride density gradient centrifugation, and analyzed by two-dimensional polyacrylamide gel electrophoresis. Conditions of electrophoresis restricted attention to proteins that are positively charged at pH 8.6. Most of the major nucleolar protein spots co-electrophoresed with ribosomal proteins; the majority of ribosomal proteins from both the large and small ribosomal subunits were represented. Several proteins found in association with polysomes but not on ribosomal subunits and several proteins unique to the nucleolus were also identified in these nucleolar protein patterns. In order to determine whether the ribosomal proteins found in the nucleolus represented sizable pools of ribosomal proteins, or merely ribosomal proteins contained in the preribosomal particles, [35S]methionine-labeled nucleoli were mixed with [3H]methionine-labeled polysomes. From analysis of isotopic ratios in individual protein spots it was possible to determine the stoidchiometry of individual ribosomal proteins in the nucleolus relative to their complement on cytoplasmic ribosomes. All but a few proteins exhibited relative nucleolar stoichiometry values of approximately one, indicating that there are not significant pools of most ribosomal proteins in isolated nucleoli.

Cell Nucleolus↗
PubMed1976

Proteins of Friend leukemia cells. Comparison of hemoglobin-synthesizing and noninduced populations.

Proteins of Friend leukemia cells induced to form large amounts of hemoglobin by dimethylsulfoxide treatment were compared with proteins from noninduced cells by high resolution two-dimensional polyacrylamide gel electrophoresis. Approximately 98% of more than 500 proteins separated by this technique were qualitatively and quantitatively the same in both cell populations. Changes representing more than 50% of the control cell amount were detected in six non-histone chromosomal proteins, two nucleoplasmic proteins, and three cytoplasmic proteins. It is concluded that dimethylsulfoxide induces an extremely specific pattern of erythroid differentiation in these cells, which should be susceptible to detailed analysis. Comparison of protein patterns from Friend leukemia cells and HeLa cells revealed electrophoretic identity of approximately 20% of cytoplasmic proteins and 50% of non-histone chromosomal proteins.

Chromatin↗
PubMed1976

Non-histone chromosomal proteins from HeLa cells. A survey by high resolution, two-dimensional electrophoresis.

Analysis of HeLa non-histone chromosomal protein by two-dimensional polyacrylamide gel electrophoresis has revealed more than 450 components, most of which are rare (less than 10,000 copies/cell) and are not detectable in cytoplasm. Chromatin prepared from nuclei isolated by a nonaqueous technique contains essentially all of the proteins found in chromatin from aqueous nuclei, implying that cytoplasmic contamination is not significant. The complexity of chromatin is comparable to that of cytoplasm (500 proteins detected) and greater than that of nucleoplasm (300 proteins detected). Approximately 60 proteins occur in chromatin, nucleoplasm, and cytoplasm; one of these appears to be actin.

Animals↗
PubMed1975

Evidence for heterogeneity of ribosomes within the HeLa cell.

The proteins of several subsets of ribosomal subunits isolated from HeLa cells were separated by high resolution electrophoresis in polyacrylamide slab gels containing sodium dodecyl sulfate. Comparisons were made among native subunits, and subunits derived from single ribosomes, free polysomes, and membrane-bound polysomes. At least 10 heterogeneously distributed proteins were identified among the 40 S subunit proteins, compared wtih 17 to 20 proteins that occurred in all 40 S subunit classes. All 60 S subunits shared 34 separable proteins. One protein was missing only from membrane-bound 60 S subunits and another protein was present only in membrane-bound 60 S subunits. It is suggested that these two proteins regulate the binding of 60 S subunits to the endoplasmic reticulum.

Cell Fractionation↗
PubMed1975

Relationship between nuclear and cytoplasmic poly(adenylic acid).

The kinetics of accumulation of radioactive poly(A) in the nucleus and cytoplasm of mouse L cells have been determined using labeling conditions in which the cellular ATP pool is shown to have a nearly constant specific radioactivity. Most or all nuclear poly(A) accumulates with kinetics very similar to those of heterogeneous nuclear RNA, having a half-time of about 25 min. There is little or no lag before attainment of a maximal rate of accumulation of cytoplasmic poly(A). These data are consistent with a variety of models in which nuclear poly(A) does or does not all serve as a precursor of cytoplasmic poly(A). In either type of model there must be a class of poly(A) either synthesized in the cytoplasm or passing through a small nuclear pool separate from the main pool of nuclear poly(A).

Cell Nucleus↗
PubMed1974

Composition of mammalian ribosomal subunits: a re-evaluation.

A method for preparation of highly active mammalian ribosomal subunits is described, which yields 60S subunits containing no more than 33% protein. It is suggested that the composition of these subunits corresponds closely to that of Escherichia coli 50S subunits. Data on the composition of bacterial and mammalian ribosomal subunits recovered from CsCl are given. It is shown that the commonly employed assumptions about the relationship between the buoyant density of a particle in CsCl and its protein content are in error. The composition of ribosomal subunits cannot ordinarily be calculated from the buoyant density in CsCl.

Animals↗