bluuebunny/crossref_metadata_2025 huggingface dataset:
Sugar Effects on Murine Sarcoma Virus Transformation
Authors: Hatanaka Masakazu
Published: May 2006

Transformation of NIH Swiss mouse fibroblasts by a murine sarcoma virus was influenced by the type of carbohydrate in the culture medium. Exposure to D-mannose and 2-deoxy-D-glucose caused a reduction in the number of transformed colonies
in vitro
and tumor formation
in vivo
. This is a new type of interference in the transformation process, manifested also by the absence of the pronounced enhancement of hexose entry into transformed cells, characteristic of a sarcoma virus infection. The uptake of D-glucose, D-mannose, and D-galactose, was repressed by normal mouse cells grown in the presence of D-mannose and D-glucose in the medium, while uptake of the same sugars was derepressed in cells grown in media containing D-xylose, L-xylose, or L-arabinose. D-Galactose had an intermediate effect on hexose uptake and permitted derepression. In every case, enhancement of the primary transport by sarcoma virus transformation was consistently beyond that observed at any time in the derepressed control cells, demonstrating the occurrence of another mechanism of enhancement of hexose uptake after sarcoma virus infection.
Virus-Like Particles in an Extranuclear Mutant of
Neurospora crassa
Authors: Küntzel Hans , Barath Zoltan , Ali Iqbal , Kind Jürgen , Althaus Hans-Hinrich
Published: May 2006

A particulate fraction not present in wild-type cells has been isolated from the cytoplasm and from lysed mitochondria of the respiratory-deficient extranuclear mutant “abnormal-1” of
Neurospora crassa
. The particles have a density between 1.13 and 1.2 g/ml. They appear in thin sections after OsO
4
fixation as virus-like polymorphic vesicles containing an electron-dense nucleoid of 120-170 nm in diameter. The central core is surrounded by one or two “unit membrane” envelopes of 100 Å thickness. The particles contain a single-stranded 33S RNA that is converted to 7-9S RNA by heat treatment in the presence of sodium dodecyl sulfate and that differs in its base composition from mitochondrial and cytoplasmic ribosomal RNA. They contain 7% phospholipids rich in phosphatidylethanolamine and two major proteins, a lipoprotein of molecular weight 15,000 and a glycoprotein of molecular weight 95,000. It is suggested that these virus-like particles originate within mitochondria.
Carbon-13 Nuclear Magnetic Resonance Spectra of Prostaglandins and Some Prostaglandin Analogs
Authors: Cooper Gary F. , Fried Josef
Published: May 2006

High-resolution pulsed Fourier-transform nuclear magnetic resonance spectroscopy at 22.63 MHz was used to observe the proton-decoupled natural-abundance
13
C nuclear magnetic resonance spectra of CDCl
3
solutions of the methyl esters of prostaglandins F
1α
, 15-epi-F
1α
, F
2α
, F
2β
, E
2
, A
2
, 13-dehydro-F
2α
, 13-dehydro-F
3α
, two intermediates on the synthetic pathway to 13-dehydro-PGF
3α
, and of 7-oxa-PGF
1α
. All resonances were assigned by chemical shift comparisons and single-frequency offresonance proton decoupling. With two exceptions, all the lines of the spectra are well-resolved single-carbon resonances. Those due to the cyclopentane and vinyl carbons are most sensitive to structural changes. Some of these effects can be rationalized in terms of the preferred conformations of the molecules.
A Comparative Evaluation of the Distribution of Concanavalin A-Binding Sites on the Surfaces of Normal, Virally-Transformed, and Protease-Treated Fibroblasts
Authors: Rosenblith J. Z. , Ukena T. E. , Yin H. H. , Berlin R. D. , Karnovsky M. J.
Published: May 2006

The topographical distributions of concanavalin A-binding sites on the surfaces of 3T3, proteasetreated 3T3, and simian virus 40-transformed 3T3 cultured mouse fibroblasts
appear
to be different, as shown by a shadow-cast replica technique using concanavalin A and a hemocyanin marker, or as shown previously on isolated membranes with concanavalin A coupled to ferritin. However, chemical fixation of cells before labeling with concanavalin A and hemocyanin, or labeling exclusively at 4°, allows one to distinguish between inherent concanavalin A-binding-site topography and potential rearrangement of sites induced by the action of the multivalent concanavalin A molecule itself. The
inherent
distribution of binding sites on 3T3, protease-treated 3T3, and transformed cells is actually the same on all cells, i.e., dispersed and random. Treatment of unfixed transformed or protease-treated 3T3 cells, but not normal 3T3 cells, with concanavalin A and hemocyanin at 37° (or at 4° with subsequent warming to 37°), however, results in clustering of binding sites, presumably due to crosslinking of neighboring lectin-binding sites by the quadrivalent concanavalin A. Thus, the underlying difference between concanavalin A-binding sites on normal as compared with transformed or protease-treated normal cells lies not in the inherent topography of binding sites, but rather in the susceptibility of the sites to aggregation by concanavalin A. The latter may reflect an increased mobility of lectin-binding sites on transformed or protease-treated cells.
Change in Cell-Surface Glycoproteins of Mouse Embryos Before Implantation
Authors: Pinsker M. Craig , Mintz Beatrice
Published: May 2006

The early biochemical ontogeny of the mammalian cell surface has been examined in mouse embryos. After
in vitro
incubation of embryos in D-[
3
H]- or D-[
14
C]glucosamine, the cell-surface glycopeptides were removed by trypsinization, digested with Pronase, and fractionated by gel filtration on Sephadex G-50; radioactivity was determined in the fractions by scintillation counting. Cochromatography of digests from the surfaces of embryos of two age groups yielded markedly different elution patterns. The labeled surface material from embryos in or near the blastocyst stage, when the outermost cells are differentiating into trophoblast, was substantially enriched, as compared with material from cleavage-stage embryos, in relatively more rapidly eluted components of presumed higher molecular weight. A similar change has previously been reported by others in cultured mammalian cell lines after viral transformation, in comparison with corresponding untransformed cells. This similarity between blastocysts and transformed cells suggests that the early biochemical differentiation of the surface, as the embryo passes into the blastocyst stage, may be responsible for the stage-specific capacity of the outermost cells to attach to the uterine wall in response to an implantation-initiating stimulus and to become invasive.
Membrane Changes and Adenosine Triphosphate Content in Normal and Malignant Transformed Cells
Authors: Vlodavsky Israel , Inbar Michael , Sachs Leo
Published: May 2006

Transformed fibroblasts had a low content of ATP when grown at a high cell density and a high content of ATP when grown at a low cell density. Concanavalin A agglutinated transformed cells with a low, but not those with a high, ATP content. Transformed cells with a high ATP content gained agglutinability after ATP depletion by inhibitors of the energy-generating systems, and those with a low ATP content lost their agglutinability after restoration of a high ATP content by glucose. Fixation of the surface membrane by formaldehyde, glutaraldehyde, or LaCl
3
, inhibited agglutination of cells with an ATP content that allows agglutination. Normal fibroblasts grown at a high or a low cell density were not agglutinated by concanavalin A. Depletion of the cellular ATP content of normal cells induced agglutination only in cells grown at a high, but not at a low, cell density. A similar number of concanavalin A molecules was bound to the surface membrane of agglutinating and nonagglutinating fibroblasts. It is suggested that a high content of ATP inhibits the movement of concanavalin A binding sites, and that a low content of ATP allows, in transformed cells, a new distribution of binding sites to form the clusters required for cell agglutination. Agglutinability of transformed cells is determined by ATP content, and in normal cells changes in the content of ATP are by themselves not sufficient to induce agglutination. Transformed cells, therefore, do not have a control, presumably for membrane stability, that exists in normal cells.
Specificity of Antibodies in Amphibian Larvae Possessing a Small Number of Lymphocytes
Authors: Haimovich Joseph , Du Pasquier Louis
Published: May 2006

Tadpoles of
Rana catesbiana
, possessing about two million lymphocytes, were immunized with 2,4-dinitrophenyl- and 2,4,6-trinitrophenyl-conjugated bacteria and proteins, and the antibodies produced were studied by inactivation of dinitrophenyl- and trinitrophenyl-conjugated bacteriophages. Crossreactions were determined by inhibition of inactivation of modified phages with the lysine derivatives of 2,4-dinitrophenyl-,2,4,6-trinitrophenyl-, and 4-mononitrophenyl-groups. Antibodies to the two hapten-conjugates differed as significantly from each other in tadpoles as in larger animals. The ability of the small number of lymphocytes in tadpoles to discriminate between structurally similar determinants, and the estimated large number of antibodies required to cover the whole spectrum of antigenic specificities, suggests that an antigen-reactive cell changes specificity during its life time.
Restoration of Hypoxanthine Phosphoribosyl Transferase Activity in Mouse 1R Cells After Fusion with Chick-Embryo Fibroblasts
Authors: Bakay Bohdan , Croce Carlo M. , Koprowski Hilary , Nyhan William L.
Published: May 2006

Fusion of the 1R mouse cell, which lacks activity of hypoxanthine phosphoribosyl transferase (EC 2.4.2.8), with chick-embryo fibroblasts yielded progeny cells that survived in hypoxanthine-aminopterin-thymidine selective medium. This property and the failure of the progeny to survive in 8-azaguanine indicated that hypoxanthine phosphoribosyl transferase activity was present. Electrophoretic analysis revealed that the enzyme was of mouse, not chick, origin. These observations are consistent with the operation of a regulator gene responsible for the absence of hypoxanthine phosphoribosyl-transferase activity in the 1R cell and its presence in the progeny.
Segregation During Ascidian Embryogenesis of Egg Cytoplasmic Information for Tissue-Specific Enzyme Development
Authors: Whittaker J. R.
Published: May 2006

Cleavage-arrested embryos of the ascidian
Ciona intestinalis
were able to differentiate two tissue-specific enzymes—muscle acetylcholinesterase (EC 3.1.1.7) and brain pigment cell tyrosinase (EC 1.10.3.1). Cytochalasin B, colchicine, Colcemid, and podophyllotoxin were used as cleavage inhibitors at early embryonic stages up to the 64-cell stage. Only certain cells in the cleavage-arrested embryos developed these histochemically detectable enzymes, and this ability followed the cell lineage patterns for the two tissues. This result implies the presence of specific positional information in the egg cytoplasm that is differentially segregated during cleavage. There were distinct and separate puromycin and actinomycin D sensitivity periods for the occurrence of each enzyme during development of both normal and cleavage-arrested embryos. The segregated information is apparently neither the enzyme proteins nor RNA templates for enzyme synthesis, but is probably concerned with activation of appropriate genes.
Mouse Lambda-Chain Sequences
Authors: Cesari Italo M. , Weigert Martin
Published: May 2006

Amino-acid sequences of the variable regions of three lambda chains produced by plasmacytomas of BALB/c mice are compared. Two are almost certainly identical and one differs from these by three amino acids. These findings extend our earlier conclusion on the relative uniformity of sequences in this type of immunoglobulin light chain. With amino-acid sequence data on two additional lambda chains, eight mouse lambda chains studied to date are indistinguishable and four probably differ from these by one, two, or three amino acids.
Discrimination between Messenger Ribonucleic Acids by a Mammalian Translation Initiation Factor
Authors: Nudel U. , Lebleu B. , Revel M.
Published: May 2006

A factor from rabbit-reticulocyte ribosome, which stimulates by 5-fold initiation of hemoglobin mRNA translation in Krebs ascites cell extracts, was purified to homogeneity. The factor, a protein of about 65,000 molecular weight in sodium dodecyl sulfate, discriminates between various mRNAs: it stimulates translation of α globin mRNA and of tobacco mosaic virus RNA, but has only a small effect on β globin mRNA and no effect at all on Mengo virus RNA translation.
RNA Polymerase from
Bacillus amyloliquefaciens
Infected with ϕ29 Bacteriophage
Authors: Holland Michael , Whiteley H. R.
Published: May 2006

DNA-dependent RNA polymerase was purified from uninfected and ϕ29-infected
Bacillus amyloliquefaciens
. Differences were observed in the specific activities, template specificities, stability, and sedimentation properties of the two enzymes. A polypeptide of 30,000 molecular weight was found in association with the polymerase of high specific activity from phage-infected cells and was absent from polymerase isolated from uninfected cells. The change in polymerase properties and the appearance of the polypeptide occurred early in phage infection.
Biological Activity of 1α-Hydroxycholecalciferol, A Synthetic Analog of the Hormonal Form of Vitamin D
3
Authors: Haussler Mark R. , Zerwekh Joseph E. , Hesse Robert H. , Rizzardo E. , Pechet Maurice M.
Published: May 2006

1,25-Dihydroxycholecalciferol, the apparent active hormonal form of cholecalciferol (vitamin D
2
), is formed from cholecalciferol by specific and sequential hydroxylations of the sterol at carbons 25 and 1. Recently, 1α-hydroxycholecalciferol was synthesized and we report on its biological activity in rachitic chicks. 1α-Hydroxycholecalciferol is identical in potency to 1,25-dihydroxycholecalciferol in stimulation of intestinal calcium absorption; either sterol elicits a near maximal effect at a dose of 0.3-0.6 nmol. The time-course of action of 1α-hydroxycholecalciferol also parallels that of the active metabolite 1,25-dihydroxycholecalciferol with a maximal increase in calcium transport occurring 5-10 hr after administration of sterol to vitamin D-deficient chicks. 6.5 nmol of 1α-hydroxycholecalciferol causes a doubling in calcium absorption in only 2-3 hr, which is the most rapid physiologic response yet detected for a vitamin D-sterol. 1α-Hydroxycholecalciferol is active also in enhancing bone calcium resorption and, like 1,25-dihydroxycholecalciferol, is at least 10 times as active as cholecalciferol in mobilizing bone calcium and raising plasma calcium concentration. It is concluded that 1α-hydroxycholecalciferol represents a synthetic analog of 1,25-dihydroxycholecalciferol that can be used both to study the mechanism of action of this hormone and as a therapeutic agent in the treatment of patients with certain metabolic bone diseases.
Adhesion of Cultured Fibroblasts to Insoluble Analogues of Cell-Surface Carbohydrates
Authors: Chipowsky Stefan , Lee Y. C. , Roseman Saul
Published: May 2006

Animal cells are coated with complex carbohydrates. Insoluble analogues of these substances were prepared by coupling monosaccharides to Sephadex beads (crosslinked dextran), and the interactions between these derivatives and three established cell lines were studied. A virally transformed fibroblast, simian virus 40-transformed 3T3 cells, adhered to the beads derivatized with D-galactose, but did not adhere to the corresponding beads derivatized with D-glucose or
N
-acetyl-D-glucosamine. Cells that adhered to the galactose-beads appeared to initiate a nucleation process in that they became more adhesive towards the cells in suspension, leading to the formation of large aggregates containing both cells and galactose-beads. The results suggest that specific carbohydrates are involved in the processes of cell recognition or cell adhesion, or both.
Molecular Structures of Acetylcholinesterase from Electric Organ Tissue of the Electric Eel
Authors: Dudai Yadin , Herzberg Max , Silman Israel
Published: May 2006

Three purified molecular forms of acetylcholinesterase (EC 3.1.1.7) with sedimentation coefficients of 18 S, 14 S, and 11 S were studied by analytical ultracentrifugation and electron microscopy. The three species have molecular weights of (1.1 ± 0.1) × 10
6
, (7.5 ± 1.5) × 10
5
, and (3.35 ± 0.25) × 10
5
, respectively. Electron micrographs reveal that the 18S and 14S forms are asymmetric, composed of a head, containing a large number of subunits, and an elongated tail. The 11S form of acetylcholinesterase is apparently a tetrameric structure devoid of the tail. Maleylation of 18S and 14S acetylcholinesterases abolishes their tendency to aggregate at low ionic strength.
Independent Segregation of the
H-2
Locus and the Locus for Responsiveness to Histamine-Sensitizing Factor
Authors: Ovary Zoltan , Vris Thomas W. , Szalay Csaba De , Vaz Nelson M. , Iritani Chika A.
Published: May 2006

The inheritance of responsiveness to histamine-sensitizing factor of pertussis bacilli was examined in (SWR/J x CBA/J)F
1
x CBA/J backcross mice. We found that about half of the backcross mice inherited this responsiveness in both sexes. There was no linkage between the inheritance of responsiveness to histamine-sensitizing factor and the

H-2
q

locus from the sensitive SW/J progenitor.
The Catalytic Mechanism of Carbonic Anhydrase
Authors: Lindskog Sven , Coleman Joseph E.
Published: May 2006

It is shown that an “inverse” relationship between the pH dependencies of the rates of hydration of CO
2
and dehydration of HCO
3
-
by carbonic anhydrase (EC 4.2.1.1) is a direct consequence of the thermodynamic equilibrium between CO
2
and HCO
3
-
and independent of any assumptions about the catalytic mechanism. It is further shown that proposed mechanisms for carbonic anhydrase involving HCO
3
-
as the substrate in the dehydration reaction and a proton transfer reaction, EH
+
⇌ E + H
+
, as an obligatory step during catalysis obey the rule of microscopic reversibility. This includes mechanisms in which the proton dissociation is from a zinc-coordinated water molecule. Such mechanisms can be in accord with the observed rapid turnover rates of the enzyme, since rapid proton exchange can occur with the buffer components, EH
+
+ B ⇌ E + BH
+
. Mechanisms in which H
2
CO
3
is the substrate in dehydration avoid the proton-transfer step, but require that H
2
CO
3
combines with enzyme more rapidly than in a diffusion-controlled reaction. Physico-chemical evidence for and against a zinc-hydroxide mechanism is discussed.
Isolation and Characterization of Lipopolysaccharide Protein from
Escherichia coli
Authors: Wu Ming-Chi , Heath Edward C.
Published: May 2006

Extraction of the cell envelope of
E. coli
with 1% sodium dodecyl sulfate yielded a lipopolysaccharide protein that was purified to homogeneity by conventional techniques. Analysis of the pure protein indicated that it is a complex lipopolysaccharide protein with the following molar ratios of constitutents: 3-deoxyoctulosonate, 1.0; glucosamine, 1.3; neutral sugar (glucose + heptose), 1.0; organic phosphate, 2.3; and amino acid, 21. On a quantitative basis, all of the 3-deoxyoctulosonate present in the cell envelope preparation was solubilized in hot sodium dodecyl sulfate and subsequently accounted for in its entirely in the isolated protein component. After incubation of the cell-wall particulate fraction of cells grown on [
3
H]glucosamine with UDP-[
14
C]galactose under conditions designed to measure galactosyl transferase activity, the isolated lipopolysaccharide protein contained all of the [
14
C]galactose that was incorporated during the incubation. We concluded that the lipopolysaccharide of this organism occurs in the outer cellenvelope membrane exclusively in the form of lipopolysaccharide protein.
Maximal Concanavalin A-Specific Agglutinability Without Loss of Density-Dependent Growth Control
Authors: Glynn Russell D. , Thrash Cornelia R. , Cunningham Dennis D.
Published: May 2006

Density-inhibited 3T3 mouse fibroblasts were treated for 10 min with nine concentrations of Pronase ranging from 0.25 to 40 μg/ml. Concanavalin A-specific agglutinability increased from control values of about 25% to plateau values of about 80%. None of these Pronase concentrations brought about an increase in cell number within 84 hr. Pronase-treated cells remained responsive to growth stimulation by serum and cortisol. Therefore, the protease-mediated surface change measured by increased concanavalin A-specific agglutinability is not an event sufficient by itself to bring about cell division.
Synergism Between Subpopulations of Thymus-Derived Cells Mediating the Proliferative and Effector Phases of the Mixed Lymphocyte Reaction
Authors: Cohen Lewis , Howe Michael L.
Published: May 2006

The mixed lymphocyte reaction is characterized by proliferation and generation of specifically cytotoxic effector lymphocytes. These two phases of the mixed lymphocyte reaction have been shown to be mediated by distinct subpopulations of thymus-derived cells. The current investigation demonstrates that combinations of cells from thymus and lymph nodes of CBA mice exhibit synergism with respect to proliferation and effector cell production in response to Balb/c alloantigens. That is, the magnitude of the proliferative and effector phases of the mixed lymphocyte reaction exhibited by combinations of thymus cells and lymph node cells was greater than that given by equal numbers of the two cell types cultured separately. This cooperative interaction between lymphoid cell subpopulations in the mixed lymphocyte reaction parallels that which is responsible for the graftversus-host reaction. It is proposed, therefore, that the mixed lymphocyte reaction provides an
in vitro
model for the study of
in vivo
thymus cell interactions occurring in the graft-versus-host reaction.
The Enduring Behavioral Changes in Rats with Experimental Phenylketonuria
Authors: Andersen Arnold E. , Rowe Vernon , Guroff Gordon
Published: May 2006

The biochemical features of phenylketonuria have been reproduced in developing rat pups by administering to them a combination of
p
-chloro-DL-phenylalanine plus L-phenylalanine for the first 21 days after birth. During the treatment period, the experimental animals show delayed eye opening and decreased brain weight compared with controls given saline. Neuropathological examination of developing animals reveals deficient myelination and some inhibition of cerebellar maturation. When tested as adults, after a long recovery period, animals with phenylketonuria are hyperactive in activity wheels. Adult rats are deficient in reversing a position choice and demonstrate impaired performance in a Y-maze. Rats treated with
p
-chloro-DL-phenylalanine plus L-phenylalanine during the vulnerable period of rapid brain development thus have enduring behavioral changes that persist throughout life.
Antibodies to Thymocytes in Sera of Patients with Schizophrenia
Authors: Luria E. A. , Domashneva I. V.
Published: May 2006

Sera from healthy people and from patients with schizophrenia were heated at 56° for 30 min and then kept at 4°. These sera were then tested for cytotoxicity and by the indirect immunofluorescence method with thymocytes, bone-marrow cells, and lymph-node cells of adult C
3
H mice. The cytotoxicity index for mouse thymocytes of sera from 60 healthy donors ranged from 0.00 to 0.4, with an average of 0.08. That of sera from 35 patients with schizophrenia ranged from 0.29 to 0.91, with an average of 0.71. The cytotoxicity index for mouse bone-marrow cells of sera from healthy donors ranged from 0.03 to 0.23, whereas that of sera from patients with schizophrenia ranged from 0.00 to 0.14. Sera of patients at dilutions of 1:8-1:32 displayed immunofluorescence of almost 100% of thymocytes and 58-78% of lymphocytes, whereas sera of healthy donors displayed none. We thus conclude that the sera of patients with schizophrenia contain antibodies against thymic antigens localized on thymocytes and thymus-derived lymphocytes.
D
-Nor Steroids: Anomalous Conformation and Reactivity of C/D-
cis
D
-Norandrostanes
Authors: Meinwald J. , Taggi A. J. , Luhan P. A. , McPhail A. T.
Published: May 2006

The C/D-
cis
D
-norandrostanes 1a, 1b, 1c, 2a, 2b, and 2c were prepared and their reactivity studied. Whereas carbonium ion reactions of 1b and 1c are readily interpreted on the basis of their assigned stereochemistry, reactions of 2b and 2c suggest that these compounds react
via
conformations in which the 16β-substituents are
pseudoequatorial
, requiring a ring C boat conformation as shown in formula 3b. X-ray crystallographic studies on 1a and 2a establish that while 1a has the expected conformation, 2a exists preferentially with ring C in the boat form. Molecular models suggest that this is the result of severe nonbonded interactions in the alternative conformation (3a) in which a chair C ring is retained.
Germ Line Basis for Antibody Diversity: Immunoglobulin V
H
- and C
H
-gene Frequencies Measured by DNA·RNA Hybridization
Authors: Premkumar E. , Shoyab M. , Williamson A. R.
Published: May 2006

The reiteration frequency for mouse immunoglobulin V
H
-genes and C
H
-genes has been directly estimated by hybridization of purified MOPC 315 α-chain mRNA with a vast excess of mouse DNA. A biphasic C
o
t curve resulted. The low C
o
t transition (C
o
t
1/2
about 1.5) was interpreted as hybridization to V
H
-genes and the high C
o
t transition (C
o
t
1/2
about 10
3
) as hybridization to C
H
-genes. These values correspond to about 5000 V
H
-genes and less than 8 C
H
-genes. This germ-line content of V
H
-genes is sufficient to account for antibody diversity given a comparable set of V
L
-genes. Minimal-gene models are invalidated and there is no need to invoke somatic generators of diversity.
Enzymatic Synthesis of Solid Phase-Bound DNA Sequences Corresponding to Specific Mammalian Genes
Authors: Venetianer Pal , Leder Philip
Published: May 2006

With oligo(dT)-cellulose as primer, RNA-dependent DNA polymerase catalyzes the synthesis of cellulose-bound DNA that is complementary to mouse globin mRNA. The resulting cellulose-bound or solid phase complementary DNA hybridizes specifically with globin mRNA and permits the recovery of intact globin mRNA. This simple technique for the synthesis of solid phase-bound complementary DNA provides an additional and convenient method for the purification of specific genetic sequences.
Kinetic Proofreading: A New Mechanism for Reducing Errors in Biosynthetic Processes Requiring High Specificity
Authors: Hopfield J. J.
Published: May 2006

The specificity with which the genetic code is read in protein synthesis, and with which other highly specific biosynthetic reactions take place, can be increased above the level available from free energy differences in intermediates or kinetic barriers by a process defined here as kinetic proofreading. A simple kinetic pathway is described which results in this proofreading when the reaction is strongly but nonspecifically driven, e.g., by phosphate hydrolysis. Protein synthesis, amino acid recognition, and DNA replication, all exhibit the features of this model. In each case, known reactions which otherwise appear to be useless or deleterious complications are seen to be essential to the proofreading function.
A Sequential Test for Two Binomial Populations
Authors: Robbins Herbert
Published: May 2006

A sequential procedure is proposed for deciding which of two sequences of Bernoulli trials has the greater success probability. The procedure covers cases in which the two sequences are being observed at different rates, whether by design or otherwise, as well as the case of pairwise observation. The error probability of the procedure depends approximately only on the odds ratio of the two probabilities, a desirable property in deciding which of two medical treatments has the greater probability of success.
A Manyfold Increase in Sister Chromatid Exchanges in Bloom's Syndrome Lymphocytes
Authors: Chaganti R. S. K. , Schonberg S. , German James
Published: May 2006

Dividing cells from persons with Bloom's syndrome, an autosomal recessive disorder of growth, exhibit increased numbers of chromatid breaks and rearrangements. A highly characteristic feature of the chromosome instability in this syndrome is the tendency for exchanges to occur between
chromatids of homologous chromosomes
at homologous sites. In the present experiments, a cytogenetic technique by which the sister chromatids of a metaphase chromosome are stained differentially has been used to demonstrate a striking and possibly specific, but hitherto unrecognized, increase in the frequency with which
sister chromatids
also exchange segments. The cells were grown in bromodeoxyuridine and stained with 33258 Hoechst and Giemsa. Whereas phytohemagglutinin-stimulated lymphocytes from normal controls had a mean of 6.9 sister chromatid exchanges per metaphase (range 1-14), those from persons with Bloom's syndrome had a mean of 89.0 (range 45-162). Normal frequencies of sister chromatid exchanges were found in cells heterozygous for the Bloom's syndrome gene, and also in cells either homozygous or heterozygous for the genes of the Louis-Bar (ataxia telangiectasia) syndrome and Fanconi's anemia, two other rare disorders characterized by chromosome instability.

In a differentially stained chromatid interchange configuration discovered during the study, it was possible to determine the new distribution of both sister and non-sister-but-homologous chromatids that had resulted from numerous exchanges. By following shifts in the pattern of staining from chromatid to chromatid, visual evidence was obtained that the quadriradial configurations long recognized as characteristic of Bloom's syndrome represent exchanges between homologous chromosomes, apparently at homologous points.

We postulate that the increase in the frequency of exchanges between nonsister-but-homologous chromatids and those between sister chromatids in Bloom's syndrome represents aspects of one and the same disturbance. A study of this phenomenon in relation to the clinical features of Bloom's syndrome may be helpful eventually in understanding the biological significance of chromatid exchange in somatic cells.
Participation of N
1
-Oxide derivatives of Adenine Nucleotides in the Phosphotransferase Activity of Liver Mitochondria
Authors: Jebeleanu G. , Ty N. G. , Mantsch H. H. , Bârzu O. , Niac G. , Abrudan I.
Published: May 2006

The modified adenine nucleotides ATP-NO, ADP-NO, and AMP-NO were tested as potential substrates and/or inhibitors of mitochondrial phosphotransferases. ADP-NO is not recognized by the translocase system located in the inner mitochondrial membrane; however, it is rapidly phosphorylated to ATP-NO in the outer compartment of mitochondria, by way of the nucleosidediphosphate kinase (EC 2.7.4.6) reaction, provided there is sufficient ATP in the mitochondria. AMP-NO is not phosphorylated by liver mitochondria to the corresponding nucleoside diphosphate; it cannot serve as substrate for adenylate kinase (EC 2.7.4.3). ATP-NO and ADP-NO, however, are substrates of this enzyme. The apparent equilibrium constant for the reaction, ADP-NO + ADP ⇌ ATP-NO + AMP, of 0.908 at pH 7.4 and 5 mM Mg
2+
is significantly higher than that of the reaction with natural nucleotides.

Although adenosine N
1
-oxide is easily phosphorylated to AMP-NO by adenosine kinase [Schnebli
et al.
(1967)
J. Biol. Chem.
242, 1997-2004], the formation of corresponding nucleoside triphosphate
in vivo
seems also to be limited by adenylate kinase; adenosine N
1
-oxide cannot replace adenosine in restoring the normal ATP level in ethionine-treated rats.
The Effect of Bicarbonate on Photosynthetic Oxygen Evolution in Flashing Light in Chloroplast Fragments
Authors: Stemler Alan , Babcock Gerald T. , Govindjee
Published: May 2006

The ability of bicarbonate ion (HCO
3
-
) to stimulate photosynthetic oxygen evolution in maize chloroplast fragments exposed to continuous light depends on light intensity. Stimulation by HCO
3
-
is less at low intensities. In HCO
3
-
-depleted chloroplasts exposed to brief saturating light flashes, period 4 oscillations (in O
2
yield per flash) are damped within three cycles. Readdition of HCO
3
-
to these preparations restores the oscillatory pattern to higher flash numbers, indicating that HCO
3
-
reduces the probability of “misses” in the photosystem II reaction center. The rate of the dark relaxation reaction S

n

′
→ S

n
+1

(where S refers to the oxidation state of the oxygen-evolving mechanism and
n
= 0, 1, or 2), after a photoact in the photosystem II reaction center, is retarded in HCO
3
-
-depleted chloroplasts compared to the rate for this reaction in depleted chloroplasts to which HCO
3
-
has been resupplied. However, the final oxygen-evolving reaction after the accumulation of four positive charges appears to be independent of HCO
3
-
. Bicarbonate has no effect on the dark deactivation of the higher oxidation states (S
2
and S
3
) of the positive charge-accumulating system. We propose two alternate ways in which the kinetic model of oxygen evolution developed by Kok
et al
. [(1970)
Photochem
.
Photobiol
. 11, 457-475] can be extended to include the action of HCO
3
-
.
Replacement of Ribosomal Protein S1 by Interference Factor iα in Ribosomal Binding of Phage MS2 RNA
Authors: Hermoso José M. , Szer Wlodzimierz
Published: May 2006

The MS2 RNA binding capacity of 30S ribosomal subunits, which is lost when protein S1 is removed, can be restored following incubation with interference factor iα and repelleting. Polyacrylamide-agarose gel electrophoresis shows that, under these conditions, a faster moving, non-RNA binding 30S species, which contains no S1, is converted to a slower moving RNA-binding one, having the same mobility as the 30S species that contains protein S1. Factor iα binds to single-stranded RNAs in a pattern that closely resembles the RNA binding pattern of initiation factor IF-3.
The Identification of the Eukaryotic Ribosomal Proteins Homologous with
Escherichia coli
Proteins L7 and L12
Authors: Stöffler Georg , Wool Ira G. , Lin Alan , Rak Karl-Heinz
Published: May 2006

Antibodies were raised against eukaryotic (rat liver) and prokaryotic (
E. coli
) ribosomal particles and ribosomal proteins. The antisera were characterized and used to determine the identity of the eukaryotic proteins homologous to
E. coli
L7 and L12. The large subunit of rat liver ribosomes contains two acidic proteins, L40 and L41; they migrate during two-dimensional polyacrylamide gel electrophoresis in a way that mimics the behavior of L7 and L12. Rat liver L40 and L41 were found to be immunologically related to
E. coli
L7/L12, hence the proteins are likely to share some structural homology.
Kinetics and Mechanism of Deoxyhemoglobin S Gelation: A New Approach to Understanding Sickle Cell Disease
Authors: Hofrichter James , Ross Philip D. , Eaton William A.
Published: May 2006

We report the results of a kinetic investigation on the gelation of purified deoxyhemoglobin S. Gelation was induced by raising the temperature and was monitored by measuring both the heat absorbed, with a microcalorimeter, and the appearance of linear birefringence, with a microspectrophotometer. The kinetics are unusual. Prior to the onset of gelation there is a delay period, followed by a sigmoidal progress curve. The delay time is formally dependent on approximately the 30th power of the deoxyhemoglobin S concentration; a decrease in concentration from 23 to 22 g/dl increases the delay time by a factor of four. It is also extremely temperature dependent; a 1°C temperature rise in the range 20-30°C almost halves the delay time. From these results we conclude that the initial rate is controlled by the nucleation of individual fibers. We present a kinetic model that accounts for the concentration, temperature, and time dependence of the initial phase of the gelation reaction. Extrapolation of our data to physiological conditions predicts that changes in intracellular hemoglobin concentration and oxygen saturation, realizable
in vivo
, produce enormous changes in the delay time. The range of delay times spans both the mean capillary transit and total circulation times. This result points to the delay time as an extremely important variable in determining the course of sickle cell disease, and suggests a new approach to therapy.
Effect of Initiation Factor 3 Binding on the 30S Ribosomal Subunits of
Escherichia coli
Authors: Pon Cynthia L. , Gualerzi Claudio
Published: May 2006

Under certain conditions, initiation factor 3 (IF-3) can cause the release of aminoacyl-tRNA bound to 30S ribosomal subunits of
E. coli
. It is shown that this IF-3-induced aminoacyl-tRNA release cannot be attributed to either nucleolytic attack or competition between IF-3 and aminoacyl-tRNA for the same ribosomal binding site. It was found that the 30S-aminoacyl-tRNA-codon complexes formed in the absence of IF-3 are intrinsically different from those prepared in the presence of IF-3. In the absence of IF-3, the ribosomal binding of aminoacyl-tRNA is a virtually irreversible process, since the bound aminoacyl-tRNA can neither be spontaneously released upon dilution nor exchanged for unbound aminoacyl-tRNA. In the presence of IF-3, the binding of one molecule of IF-3 per 30S ribosome renders the binding of aminoacyl-tRNA reversible upon dilution and promotes exchange between bound and unbound aminoacyl-tRNA. It is suggested that this difference is due to a conformational transition of the 30S ribosomal subunit induced by the binding of IF-3. The possible implications of this finding in relation to the mechanism of action of IF-3 and its functional role in the cell are discussed.
Thiostrepton Inhibition of Initiation Factor 1 Activity in Polypeptide Chain Initiation in
Escherichia coli
Authors: Sarkar Probir , Stringer Evan A. , Maitra Umadas
Published: May 2006

Thiostrepton, a peptide antibiotic, inhibits the GTP-dependent 70S initiation complex formation (as measured by binding of fMet-tRNA to ribosomes and concomitant hydrolysis of GTP) only when initiation factor 1 is present to permit catalytic recycling of initiation factor 2 in the initiation reaction. When initiation factor 1 is absent, the binding of fMet-tRNA and GTP hydrolysis occur stoichiometrically with respect to initiation factor 2, and thiostrepton has no effect on either reaction under these conditions. Detailed analysis of this inhibition process shows that thiostrepton prevents catalytic recycling of initiation factor 2 by blocking the action of initiation factor 1, which is required for the dissociation of initiation factor 2 from the 70S initiation complex. This dissociation is necessary for the catalytic reutilization of initiation factor 2 in the initiation reaction. The antibiotic does not directly inhibit GTP hydrolysis
per se
in initiation.

The inhibition of fMet-tRNA binding to ribosomes by thiostrepton is also dependent on the concentration of GTP; the inhibition is most pronounced at low concentrations of GTP, but at a high molar ratio of GTP to thiostrepton, the inhibition is completely abolished.
Cell-Free Translation of Messenger RNA of Simian Virus 40: Synthesis of the Major Capsid Protein
Authors: Prives Carol L. , Aviv Haim , Paterson Bruce M. , Roberts Bryan E. , Rozenblatt Shmuel , Revel Michel , Winocour Ernest
Published: May 2006

Extracts of wheat germ are capable of synthesizing the major capsid protein of simian virus 40. Poly(A)-containing RNA from BS-C-1 cells infected with simian virus 40 directed the synthesis of a novel polypeptide that migrates in polyacrylamide gels together with the major capsid polypeptide of simian virus 40, VP-1. The patterns of the major tryptic peptides of purified VP-1 and the novel polypeptide synthesized
in vitro
were identical after two-dimensional paper electrophoresis. The novel polypeptide was not synthesized in response to poly(A)-rich RNA from uninfected cells or from virus-infected cells treated with cytosine arabinoside. Messenger RNA from infected cells purified by selective hybridization to DNA of simian virus 40 directs the synthesis of a major polypeptide of electrophoretic mobility similar to that of VP-1 of simian virus 40. This approach should prove useful in identifying additional products specified by DNA tumor viruses.
Homology Between Burkitt Herpes Viral DNA and DNA in Continuous Lymphoblastoid Cells from Patients with Infectious Mononucleosis
Authors: Kieff Elliott , Levine Judith
Published: May 2006

At least 90% of the sequences of purified,
in vitro
labeled, DNA from Epstein-Barr virus (prepared from HR-1, Burkitt's lymphoblastoid cells) are homologous to the DNA of the herpes virus contained in cell lines derived from patients with infectious mononucleosis. The thermal stability of the homologous and heterologous hybrid DNA molecules could not be differentiated, indicating at least 97% matching of base pairs between DNA of Epstein-Barr virus and the herpes viral DNA contained in the lymphoblasts from patients with infectious mononucleosis.
Nucleotide Sequences of RNA Transcribed in Infected Cells and by
Escherichia coli
RNA Polymerase from a Segment of Simian Virus 40 DNA
Authors: Dhar R. , Zain S. , Weissman S. M. , Pan J. , Subramanian K.
Published: May 2006

The nucleotide sequence of 180 residues of an RNA transcript of DNA of simian virus 40 has been deduced. This sequence adjoins a preferred initiation site for
E. coli
RNA polymerase. Comparison of this sequence with that of complementary RNA of simian virus 40 from infected cells has shown that this site also adjoins the apparent 3′ terminus of some of the cytoplasmic complementary RNA of simian virus 40.
Chemotactic Activity in Dialyzable Transfer Factor
Authors: Gallin John I. , Kirkpatrick Charles H.
Published: May 2006

Dialyzable transfer factor from human leukocytes was found to be strongly chemotactic for granulocytes and weakly chemotactic for monocytes
in vitro
. Chemotactic properties were also demonstrated
in vivo
in rhesus monkey skin. Initial purification of the dialyzable transfer factor by Sephadex G-25 chromatography revealed multiple fractions containing material with 255-nm absorbance. The fractions containing chemotactic activity were also capable of transferring delayed hypersensitivity to rhesus monkeys. This chemotactic material has an apparent molecular weight of 5000 daltons or less and was not inactivated by goat antibody to components C3 or C5 of human complement; chemotactic activity was lost after storage for two weeks at 4° and the activity of two of three preparations was decreased by heating for 30 min at 56°. This previously undescribed chemotactic activity of dialyzable transfer factor may have significance in relation to cell-mediated immune responses.
Site of J Chain Attachment to Human Polymeric IgA
Authors: Mestecky Jiri , Schrohenloher Ralph E. , Kulhavy Rose , Wright Genesis P. , Tomana Milan
Published: May 2006

A fragment containing J chain was released from human polymeric myeloma IgA protein by cyanogen bromide cleavage. The identity of the fragment was determined by its electrophoretic mobility and antigenic determinants. After purification by gel filtrations and DEAE-Sephadex chromatography, this fraction appeared similar (with respect to its amino acid and carbohydrate compositions and its peptide maps) to the J chain isolated from this IgA protein; the molecular weight was 17,000 ± 100. Upon reduction and alkylation, with subsequent separation of peptides by gel filtration, three components were obtained: the largest component (molecular weight 13,400) corresponded to the N-terminal segment of J chain and contained a homoserine residue, the second corresponded to the C-terminal part of J chain with 13-18 amino acid residues, and the third corresponded to the C-terminal octapeptide of the α chain. The data indicate that J chain is attached to α chain(s) through the penultimate cysteine residue of the C-terminal octapeptide.
In Vitro
Activation of the Transcription of
ara
BAD Operon by
ara
C Activator
Authors: Lee Nancy , Wilcox Gary , Gielow William , Arnold John , Cleary Paul , Englesberg Ellis
Published: May 2006

The transcription of
ara
BAD operon requires
ara
C activator and cyclic AMP. D-Fucose inhibits
ara
mRNA synthesis. Our results indicate that the positive control by
ara
C activator is exerted at the level of transcription.
Persistence of
O
6-Ethylguanine in Rat-Brain DNA: Correlation with Nervous System-Specific Carcinogenesis by Ethylnitrosourea
Authors: Goth Regine , Rajewsky Manfred F.
Published: May 2006

The nervous system specificity of the carcinogenic effect of
N
-ethyl-
N
-nitrosourea in rats at the perinatal age is of particular interest because the formation of the ultimate reactant, an electrophilic ethyl cation, occurs nonenzymatically and, hence, is not tissue specific. Indeed, similar initial degrees of DNA ethylation were found in the DNA of target (brain) and nontarget tissue (liver), in terms of the molar fractions of
O
6-ethylguanine,
N
7-ethylguanine, and
N
3-ethyladenine 1 hr after a pulse of [
1
-
14
C]ethylnitrosourea. However, over a 240-hr period of observation, the elimination rate from DNA of
O
6-ethylguanine (a modified base likely to cause anomalous base pairing during DNA replication) was strikingly slower in brain (half-life, about 220 hr) as compared to liver (about 30 hr), and also much slower than the elimination rates from brain DNA of
N
7-ethylguanine (about 90 hr) and
N
3-ethyladenine (about 16 hr). The data suggest that the rate of elimination from DNA of
O
6-ethylguanine may be an important factor (with the requirement for DNA replication of the target cell) in neoplastic transformation by ethylnitrosourea.
A Microassay for Uroporphyrinogen I Synthase, One of Three Abnormal Enzyme Activities in Acute Intermittent Porphyria, and its Application to the Study of the Genetics of this Disease
Authors: Sassa S. , Granick S. , Bickers D. R. , Bradlow H. L. , Kappas A.
Published: May 2006

A new spectrofluorometric assay is described for quantitating uroporphyrinogen I synthase (EC 4.3.1.8) activity in volumes of human blood as small as 2 μl. By this sensitive assay the inheritance of the enzyme's activity has been studied and the genetic defect for acute intermittent porphyria has been confirmed to be autosomal dominant in nature. There is a 3-fold range of uroporphyrinogen I synthase activity in erythrocytes in the normal population, with a mean
V
max
± SD of 35.7 ± 8.4 nmol of uroporphyrinogen I formed per ml of erythrocytes per hr, at 37°. One-half this level of enzyme activity (18.0 ± 5.0) is found in erythrocytes from patients with clinically manifest acute intermittent porphyria; and in erythrocytes from those of their relatives, including prepubertal children, who have the latent gene defect for the disease. The

K
m

of erythrocyte enzyme of normal people is 12.3 ± 3.9 μM, whereas the

K
m

of the erythrocyte enzyme of patients with acute intermittent porphyria is 6.2 ± 3.9 μM, as determined on whole blood lysates. Three enzymic changes have now been identified in patients with acute intermittent porphyria; a high level of δ-aminolevulinate synthase activity; a low level of uroporphyrinogen I synthase activity; and a deficiency of steroid Δ
4
-5α reductase activity.