bluuebunny/crossref_metadata_2025 huggingface dataset:
Microtubule structure at low resolution by x-ray diffraction.
Authors: Mandelkow E , Thomas J , Cohen C
Published: May 2006

Analysis of x-ray diagrams of oriented hydrated cytoplasmic microtubules shows that the tubule wall extends from about 70 to 150 A radially. The central region of the wall appears homogeneous, but the outside surface is subdivided by vertical grooves separating the 13 protofilaments and by a steep 10-fold family of grooves. The inside surface is dominated by the 10-start grooves with no clear subdivision between the protofilaments.
Levels of translatable mRNAs for cell surface protein, collagen precursors, and two membrane proteins are altered in Rous sarcoma virus-transformed chick embryo fibroblasts.
Authors: Adams S L , Sobel M E , Howard B H , Olden K , Yamada K M , de Crombrugghe B , Pastan I
Published: May 2006

Transformation of chick embryo fibroblasts by Rous sarcoma virus results in decreased amounts of a major cell surface protein and of collagen. To determine the mechanism accounting for the decreased production of these proteins, we have measured the relative amounts of functional mRNAs for these and other transformation-sensitive proteins. Total cellular RNAs extracted from normal cells and from cells transformed by the Schmidt-Ruppin strain of Rous sarcoma virus were translated in a cell-free system derived from wheat germ. Analysis of the in vitro translation products of RNAs from normal and transformed chick embryo fibroblasts shows a 5-fold reduction in the translatable mRNA for cell surface protein and a 10-fold reduction in translatable mRNA for two collagen precursors. In addition, increases in functional mRNA are observed for myosin and for two membrane polypeptides with molecular weights of 95,000 and 78,000; the latter two proteins increase on transformation, but the increases are in large part secondary to the depletion of glucose from the medium of transformed cells. Our data suggest that some of the major cellular changes induced by oncogenic viruses are due to changes in the activity of specific cellular genes.
Sequential stages of human T lymphocyte differentiation
Authors: Touraine Jean-Louis , Hadden John W. , Good Robert A.
Published: May 2006

Induction of thymus-dependent lymphocyte (T cell) differentiation was performed
in vitro
with thymic factors as inducers. T cell precursors from human bone marrow first expressed surface differentiation antigens and then acquired the capacity to form rosettes with sheep erythrocytes. The latter marker could not be induced when cells with differentiation antigens had been eliminated. The proliferative responses to phytomitogens or to allogeneic stimuli appeared to be characteristics of later stages in differentiation that also can be induced or amplified by
in vitro
incubation of marrow cells or thymocytes with thymic factors. When phytomitogen-responsive cells from peripheral blood were inactivated
in vitro
, the allogeneic response was enhanced. Although these responses are acquired almost concomitantly, they are therefore envisioned to be characteristics of separate T cell subsets. After immunological reconstitution of patients, the T cell development
in vivo
involves a succession of differentiation events similar to that described above. Our experiments with mice, using similar methods, have also shown that graft-versus-host inducing capacity is a function of a cell population distinct from that which yields a proliferative response to
in vitro
stimulation by phytohemagglutinin. These results support our model of sequential differentiation of human prothymocytes into various subsets of mature T cells.
Formation of lung surfactant films from intact lamellar bodies.
Authors: Paul G W , Hassett R J , Reiss O K
Published: May 2006

Lamellar bodies, an intracellular source of lung alveolar surfactant, were isolated from rat lung homogenates and studied in the Langmuir-Adams surface balance. By layering intact lamellar bodies on the surface of a more dense sucrose subphase, we studied the factors affecting film formation from surface tension-vs-time data and determined surface tension-surface area isotherms by compression and expansion of the resulting films. We found that films with properties representative of the alveolar surfactant are formed in the presence of Ca2+ or Mg2+ alone, or either plus Na+; that film formation is incomplete with Na+ alone or on ion-free subphases; and that Ca2+-induced film formation is blocked by chelation with EGTA but is unaffected by diisopropylfluorophosphate. The results suggest that divalent cations induce film formation by interactions at sites within the lamellar bodies and may be responsible for the binding of membrane lipids to membrane proteins in lung surfactant.
An observation on the structure of production sets with indivisibilities
Authors: Scarf Herbert E.
Published: May 2006

A subset of the constraints of an integer programming problem is said to be binding if, when the remaining constraints are eliminated, the smaller problem has the same optimal solution as the original problem. It is shown that an integer programming problem with
n
variables has a set of binding constraints of cardinality less than or equal to

2
n
-1

. The bound is sharp.
Reconstitution of catecholamine-sensitive adenylate cyclase activity: interactions of solubilized components with receptor-replete membranes.
Authors: Ross E M , Gilman A G
Published: May 2006

Membranes of mouse L cells that contain adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] but lack beta-adrenergic receptors have been solubilized with Lubrol 12A9. Addition of such adenylate cyclase-containing extracts to beta-adrenergic receptor-replete membranes from adenylate cyclase-deficient S49 lymphoma cells results in the production of a catecholamine-sensitive adenylate cyclase system. The effects of beta-adrenergic agonists and antagonists on the reconstituted system reproduce those that are characteristic of the wild-type S49 lymphoma cell. The uncoupled variant of the S49lymphoma contains adenylate cyclase, but donor extracts from this clone fail to reconstitute the hormone-sensitive enzyme activity when added to adenylate cyclase-deficient membranes. Thus, the uncoupled and adenylate cyclase-deficient variants of the S49 cell are not complementary.
Terminus region of the chromosome in Escherichia coli inhibits replication forks.
Authors: Kuempel P L , Duerr S A , Seeley N R
Published: May 2006

Induction of prophage P2sig5 at 42 degrees caused replication of the bacterial chromosome in a dnaA mutant of Escherichia coli. The P2sig5 is integrated in this strain near the metG locus, which is at min 47 on the genetic map. The regions of the chromosome replicated after prophage induction have been determined by means of DNA-DNA hybridization with various DNAs obtained from Proteus mirabilis/E. coli F' merogenotes and from lambda specialized transducing phage. The replication was initiated at the prophage site and was bidirectional. Most of the replication occurred in a counterclockwise direction on the genetic map, and the replication quickly proceeded to the aroD locus (min 37). The replication forks were retarded between aroD and rac (min 31) loci, although the rac locus was finally replicated. A more severe inhibition of replication occurred between the rac and trp (min 27) loci. It is proposed that the replication terminus is near the rac locus and that the terminus inhibits replication forks.
Intracellular distribution of terminal deoxynucleotidyl transferase in rat bone marrow and thymus
Authors: Gregoire Kathleen E. , Goldschneider Irving , Barton Randall W. , Bollum F. J.
Published: May 2006

A subset of bone marrow cells that contains terminal deoxynucleotidyl transferase (DNA nucleotidylexotransferase; nucleosidetriphosphate:DNA deoxynucleotidylexotransferase, EC 2.7.7.31) can be identified in adult rats by immunofluorescence using affinity-column-purified antibody to homogeneous calf transferase. The transferase-positive cells comprise approximately 1.8% of bone marrow cells. Correcting the specific activity of terminal transferase in total bone marrow cells (0.21 units per 10
8
cells) for the percentage of transferase-positive bone marrow cells (1.8%) gives 11.7 units per 10
8
cells, a value approximately half that found for transferase-positive thymocytes.

Fluorescence appears to be restricted to the nucleus of transferase-positive bone marrow cells, in contrast to the predominantly cytoplasmic fluorescence of small thymocytes from adult rats. Some large thymocytes contain intranuclear transferase fluorescence patterns similar to those seen in bone marrow. These thymocytes are especially numerous in neonatal rat thymus, where they are localized in the subcapsular region of the cortex. Thymocytes with combined patterns of nuclear and cytoplasmic transferase are also present. In addition, Thy-1.1 antigen, which is present on thymic and prethymic cells but not on the majority of post-thymic cells in the rat, is also present on transferase-positive bone marrow cells.

The results suggest that the transferase-positive subset of bone marrow cells may contain the immediate progenitors of cortical thymocytes in the rat. The nuclear location of fluorescence may indicate the site of physiological activity of terminal transferase in thymocytes and their precursors.
Histopathogenesis of 7,12-diemthylbenz(a)anthracene-induced rat mammary tumors.
Authors: Haslam S Z , Bern H A
Published: May 2006

The histopathogenesis and growth behavior of mammary tumors and dysplasias induced by a single intragastric dose of 7,12-dimethylbenz[a]anthracene in 50-day-old virgin female Lewis rats were examined both in situ and after transplantation into gland-free mammary fat pads of syngeneic hosts. Terminal mammary ductules are indicated as a site of origin of both ovarian hormone-dependent mammary tumors and spontaneously gegressing mammary tumors, and terminal ductule hyperplasia appears to be an early stage in mammary tumor formation. The precancerous nature of hyperplastic alveolar nodules induced by dimethylbenzanthracene in rats has been further examined, and our studies indicate that these nodules are not significantly preneoplastic.
Bond angles in transition-metal tricarbonyl compounds: A test of the theory of hybrid bond orbitals
Authors: Pauling Linus
Published: May 2006

The theory of hybrid bond orbitals is used to calculate equations giving the value of the bond angle OC—M—CO in relation to the bond number of the metal—carbonyl bond for tricarbonyl groups in which the transition-metal atom is enneacovalent or octacovalent and the group has approximate trigonal symmetry. For cobalt and iron and their congeners the average experimental values lie within about 1° of the theoretical values for enneacovalence, which are 101.9° for Co(CO)
3
and 94.5° for Fe(CO)
3
. This agreement provides strong support for the theory. For Mn(CO)
3
and Cr(CO)
3
the experimental values indicate the average covalence to be about 8.4 and 7.7, respectively, in agreement with considerations based on the electroneutrality principle.
Structure of nascent replicative form DNA of coliphage M13.
Authors: Dasgupta S , Mitra S
Published: May 2006

Nascent replicative form type II (RFII) DNA of coliphage M13 synthesized in an Escherichia coli mutant deficient in the 5' leads to 3' exonuclease associated uith DNA polymerase I contains ribonucleotides that are retained in the covalently closed RFI DNA sealed in vitro by the joint action of T5 phage DNA polymerase and T4 phage DNA ligase. These RFI molecules are labile to alkali and RNase H, unlike the RFI produced either in vivo or from RFII with E. coli DNA polymerase I and E. coli DNA ligase. The ribonucleotides are located at one site and predominantly in one strand of the nascent RF DNA. Furthermore, these molecules contain multiple small gaps, randomly located, and one large gap in the intracistronic region.
Specific regulation by steroid hormones of the amount of type I cyclic AMP-dependent protein kinase holoenzyme.
Authors: Fuller D J , Byus C V , Russell D H
Published: May 2006

The total amounts of type I and type II cytoplasmic cyclic AMP-dependent protein kinase activities were measured in various tissues of intact rats and rats subjected to castration, hypophysectomy, or adrenalectomy. After castration, the total amount of type I activity decreased rapidly in classifically steroid-responsive tissues such as the ventral prostate and levator ani muscle and less rapidly in the liver. After hypophysectomy and adrenalectomy, type I activity in the liver decreased to the same extent as after castration. Type I activity could be maintained in the ventral prostate and levator ani muscle at control levels by the daily injection of dihydrotestosterone. Furthermore, after post-castration regression of the prostate for 3 days, three daily subcutaneous injections of dihydrotestosterone resulted in a complete restoration of type I activity toe the intact level. The amount of type II activity was not altered by any of the experimental ablations. This study provides evidence linking steroid action to the ability of steroid-responsive tissues to maintain a substantial activity of type I cyclic AMP-dependent protein kinase.
Electronic properties of some protein--methylglyoxal complexes.
Authors: Bone S , Lewis T J , Pethig R , Szent-Györgyi A
Published: May 2006

Steady-state conductivity measurements and dielectric measurements in the frequency range 10(-5) to 100 Hz are reported for samples of bovine serum albumin, casein, and lysozyme complexed with methylglyoxal. Compared with the untreated proteins, the brown complexed proteins exhibit an increased conductivity and free electron spin density, together with a low-frequency dielectric dispersion. These results can be taken as evidence that the interaction with methylglyoxal results in the proteins possessing an increased electronic activity associated with the creation of mobile electron holes within the valence band states of the protein molecules.
Determination of capsid size by satellite bacteriophage P4.
Authors: Shore D , Dehò G , Tsipis J , Goldstein R
Published: May 2006

Satellite bacteriophage P4 requires all morphogenic gene products provided by a helper phage, such as coliphage P2, to assemble its own capsid, which is one-third the volume of the larger helper capsid. We have isolated a satellite phage P4 sid (size determination) mutant that is unable to direct the assembly of the small wild-type-size P4 capsid. Instead, this mutant produces P4 plaque-forming units with large P2-size capsids which contain two or three copies of the P4 sid1 genome. P4 sid1 is evidently mutated in a protein that is specifically responsible for determining the precise size and symmetry of the structure into which the helper P2 gene products will assemble. In addition, we have found that the physical size of the genome does not appear to play an essential role in the proper assembly of the icosahedral capsid, since the majority of the P4 sid1 plaque-forming units do not contain a complete capsidful of DNA.
Blocking of primary in vitro antibody responses to thymus-independent and thymus-dependent antigens with antiserum specific for IgM or IgD.
Authors: Cambier J C , Ligler F S , Uhr J W , Kettman J R , Vitetta E S
Published: May 2006

The effect of anti-mu and anti-delta on the primary in vitro IgM response of murine splenocytes to thymus-dependent (trinitrophenylated erythrocytes) and thymus-independent (trinitrophenylated brucella) forms of trinitrophenyl was studied. The results indicate that either anti-mu or anti-delta can block the response of adult splenocytes to the thymus-dependent antigen. The thymus-dependent responses of neonatal splenocytes that bear a low concentration of IgD were also abrogated by treatment with anti-delta. In contrast, anti-mu, but not anti-delta, blocked the response of adult splenocytes to the thymus-independent antigen used. These results indicate that both IgM and IgD are receptors required for triggering cells by a thymus-dependent antigen but that only IgM receptors are required for triggering by the thymus-independent antigen used.
Structure of the orgin of DNA replication of bacteriophage fd.
Authors: Gray C P , Sommer R , Polke C , Beck E , Schaller H
Published: May 2006

An RNA-polymerase-protected DNA fragment of 125 nucleotides from the origin of single-strand to double-strand replication of bacteriophage fd (ori-DNA) was located on the physical map of the phage genome. A stretch of 187 base pairs of DNA including the ori-DNA was sequenced. This DNA segment contains regions with a highly asymmetric pyrimidine/purine distribution next to regions with 2-fold symmetry that form stable hairpin structures in the viral DNA strand.
Melatonin secretion decreases during the proestrous stage of the rat estrous cycle.
Authors: Ozaki Y , Wurtman R J , Alonso R , Lynch H J
Published: May 2006

Urine was collected from rats during 12 consecutive daily dark periods and assayed for melatonin and norepinephrine; the phase of the vaginal estrous cycle associated with each urine sample was determined from daily vaginal smears. The proestrous phase of the estrous cycle was consistently associated with significant reductions in the excretions of both compounds. The level of melatonin in any urine sample tended to vary as a function of its norepinephrine content; however, the slope of the curve relating these two compounds in metestrous-diestrous samples differed from that for proestrous-estrous specimens. This difference suggests that factors other than the catecholamine (e.g., gonadal hormones) also affect melatonin secretion. Oophorectomy elevated the melatonin concentration of serum but not that of the pineal; this rise was suppressed by the administration of estrogen plus progesterone. The fate of circulating melatonin (as indicated by the proportion of an exogenous dose excreted into the urine) was not affected by the state of the estrous cycle.
Utilization of a depsipeptide substrate for trapping acyl—enzyme intermediates of penicillin-sensitive D-alanine carboxypeptidases
Authors: Rasmussen James R. , Strominger Jack L.
Published: May 2006

The penicillin-sensitive D-alanine carboxypeptidases of
Bacillus subtilis, Escherichia coli
, and
Staphylococcus aureus
catalyzed the hydrolysis of the D-lactic acid residue from the depsipeptide diacetyl-L-lysyl-D-alanyl-D-lactic acid. The ester substrate was hydrolyzed faster than the peptide analogue, diacetyl-L-lysyl-D-alanyl-D-alanine, by the
B. subtilis
(15-fold) and
E. coli
(4-fold) carboxypeptidases, presumably because acylation (
k
2
), which is the rate-limiting step of the peptidase reaction, occurred more rapidly during cleavage of the ester bond than during cleavage of the amide bond. No rate acceleration was observed with the
S. aureus
carboxypeptidase for which deacylation (
k
3
) is already the rate-determining step with the peptide substrate. The efficiency of utilization of the depsipeptide (
V
max
/
K
m
) was greatly enhanced (19- to 147-fold) for all three enzymes. After incubation of the
B. subtilis
carboxypeptidase and [
14
C]diacetyl-L-lysyl-D-alanyl-D-lactic acid at pH 5.0 and lowering of the pH to 3.0, a radioactive acyl-enzyme intermediate containing 0.43 mol of substrate per mol of enzyme was isolated by Sephadex G-50 chromatography. After acetone precipitation, the acyl group of the denatured acyl-enzyme complex appeared to be bound to the protein by an ester bond. Acyl enzymes were also detected for the
S. aureus
and
E. coli
carboxypeptidases after sodium dodecyl sulfate/polyacrylamide gel electrophoresis and fluorography of enzyme incubated with [
14
C]depsipeptide and precipitated with acetone.
Social dynamics and the quantifying of social forces
Authors: Montroll Elliott W.
Published: May 2006

Social and industrial evolutionary processes are considered to be a sequence of replacements or substitutions: new ideas for old, new labor patterns for old, new technologies for old. The logistic equation has often been used to describe population growth processes and replacement processes. It sometimes suffers from contradicting observational data. It is shown here that the deviations are often associated with unusual intermittent events—wars, strikes, economic panics, etc.—and that in many cases a few years after the event it can be abstracted as an instantaneous δ function impulse. After the event, the evolutionary process continues along its normal course. A formula is derived to use the observational data to determine the strength of the impulse modeling an event.
The dimension of product spaces
Authors: Wage Michael L.
Published: May 2006

The covering dimension of a product space can exceed the sum of the dimensions of its factors. A separable metric space
X
and a paracompact space
Y
are constructed such that dim
X
+ dim
Y
= 0 < 1 = dim(
X
×
Y
). Related results are also discussed.
Brain-derived fibroblast growth factor: identity with a fragment of the basic protein of myelin.
Authors: Westall F C , Lennon V A , Gospodarowicz D
Published: May 2006

Fibroblast growth factors (FGF) isolated from bovine brain have been identified chemically and immunologically as components of the myelin basic protein. The intact bovine basic protein molecule (170 residues), prepared by the standard acid extraction procedure, lacked mitogenic activity (tested at concentrations up to 10 microgram/ml). However, the polypeptide FGF-2, identified as residues 44-153 of the basic protein, was maximally mitogenic for fibroblasts at 10 ng/ml and polypeptide 44-166 (FGF-1) was maximally active at 100 ng/ml. Pituitary-derived FGF is a potent a growth factor as FGF-2, but appears to be biochemically and immunologically distinct from brain-derived FGF. FGF released in the central or peripheral nervous system as a consequence of myelin damage and basic protein proteolysis could provide a physiological stimulus for wound healing and myelin repair.
DNA clones containing mouse immunoglobulin kappa chain genes isolated by in vitro packaging into phage lambda coats.
Authors: Lenhard-Schuller R , Hohn B , Brack C , Hirama M , Tonegawa S
Published: May 2006

Endonuclease EcoRI-digested DNAs from BALB/c mouse embryos and MOPC 321 (a kappa chain secretor) myeloma were fractionated by agarose gel electrophoresis, and the DNA fragments containing part or all of the MOPC 321 kappa chain structural gene sequences were visualized by the Southern gel blotting technique using as the hybridization probes pCRI plasmids containing all or part of the enzymatically synthesized cDNA transcripts of the MOPC 321 kappa chain mRNA. The clear differences observed in the hybridization patterns of the two DNAs are in agreement with our previously reported results obtained with endonuclease BamHI and confirms that the sequence arrangement of kappa chain genes is different in the embryo and myeloma cells. We have cloned most of the kappa-sequence-positive EcoRI DNA fragments in Charon 4A phage by using the highly efficient in vitro phage lambda DNA packaging method, and we have characterized the cloned mouse DNA sequences by agarose gel blotting and R-loop mapping in electron microscopy. These studies identified, among others, one EcoRI DNA fragment which contains both variable and constant immunoglobulin kappa-gene sequences and is present only in the myeloma DNA. The two sequences are separated by a 2.8-kbase intron. We tentatively conclude that the kappa gene sequences on this DNA fragment underwent somatic rearrangement.
Biosynthesis of beta-endorphin from beta-lipotropin and a larger molecular weight precursor in rat pars intermedia.
Authors: Crine P , Gianoulakis C , Seidah N G , Gossard F , Pezalla P D , Lis M , Chrétien M
Published: May 2006

When isolated rat pars intermedia cells were incubated for 10 min with radioactive amino acids, one major labeled protein with a molecular weight of 30,000 +/- 1500 was extracted. This protein was shown to contain in its sequence the antigenic determinants for corticotropin and beta-melanotropin by immunoprecipitation. When the radioactivity incorporated into this large molecular weight protein during the first 10 min was chased by a further incubation in presence of an excess of unlabeled amino acid, the initial protein was degraded into several smaller peptides including beta-endorphin and beta-lipotropin. Another 18,000-dalton peptide was also observed and was tentatively identified as a large molecular form of corticotropin. From the kinetics of the maturation of the initial precursor, it is concluded that the initial cleavage of the 30,000-dalton peptide gives rise to beta-lipotropin and the 18,000-dalton form of corticotropin. beta-Lipotropin is subsequently cleaved to form beta-endorphin.
Phosphate (oxygen)-water exchange reaction catalyzed by human prostatic acid phosphatase.
Authors: Van Etten R L , Risley J M
Published: May 2006

Conclusive evidence is presented that an acid phosphatase catalyzes phosphate (oxygen)-water exchange. Studies conducted with human prostatic acid phosphatase by two independent methods have established that, despite earlier reports to the contrary, the enzyme catalyzes an exchange reaction between oxygen atoms of phosphate ion and of water. Kinetic data were obtained both by chemical conversion to trimethyl phosphate followed by mass spectroscopy and by a totally independent method involving 31P isotope shift nuclear magnetic resonance spectroscopy. Analysis showed that the enzyme catalyzes the exchange in a random, noncoupled process. If any coupled exchange occurs, it must represent less than 10% of the total. By mass spectral analysis, catalytic rate constants kcat = 0.14 sec-1 (4 degrees) and 1.8 sec-1 (37.5 degrees) were obtained. By 31P nuclear magnetic resonance kcat = 1.6 sec-1 (31 degrees) was obtained. The energy of activation for the exchange reaction is approximately 13kcal mol-1. The kcat value for exchange is about 10-fold greater than that observed with Escherichia coli alkaline phosphatase.
Electron spin resonance studies on interaction of complement proteins with erythrocyte membranes.
Authors: Dahl C E , Levine R P
Published: May 2006

Sheep erythrocytes have been spin labeled with 5-, 12-, and 16-nitroxystearic acid in order to investigate complement-induced changes in the physical state of the lipid bilayer. Formation of osmotic lesions in the membrane causes an increase in the fluidity of the membrane which overcomes the decrease in membrane fluidity caused by the interaction of the complement proteins. A decrease in membrane fluidity is observed only when complement-lysed membranes are resealed or when complement proteins react with isosmolar ghosts that do not undergo osmotic lysis. The decrease in bulk fluidity of the membrane is first observed when C8 binds to the membranes bearing C5b67 and is enhanced upon the subsequent binding of C9. The decrease in membrane fluidity shown by the electron spin resonance spectra of spin-labeled fatty acids suggests that certain of the complement proteins penetrate the membrane and interact with hydrophobic regions of the lipid bilayer.
Partial purification and characterization of the intercellular bridge from cultured mouse cells
Authors: Krystal Gerald , Rattner J. B. , Hamkalo Barbara A.
Published: May 2006

At the completion of mitosis, the two daughter cells are connected by a channel of membrane-bound cytoplasm, the intercellular bridge. This structure contains parallel arrays of spindle microtubules which are associated, at the bridge midline, with a metallophilic band called the midbody. In an effort to characterize midbody components, intercellular bridges were partially purified. The purification consisted of brief sonication of telophase populations of mouse L929 cells in order to shear intercellular bridges from daughter cells, digestion of chromatin by an excess of micrococcal nuclease, and differential centrifugation to enrich for intercellular bridges. Electron microscopy of these preparations substantiated the identity of the bulk of material as intercellular bridges. After solubilization with sodium dodecyl sulfate, the protein components of these preparations were iodinated with Na
125
I and separated by two-dimensional gel electrophoresis. From these analyses, the major polypeptide components of intercellular bridges appear to be tubulin, varying amounts of plasma membrane proteins, and a polypeptide with an apparent molecular weight of 42,000. Time-course studies reveal that this polypeptide is first detectable in a pelletable form approximately 30 min after cells are released from metaphase block, reaches maximal spot intensity in late telophase, and is no longer detectable in G1 populations. We interpret these data to suggest that the 42,000-dalton polypeptide is a component of the midbody.
Translocation of a hydrocarbon fluorescent probe between Epstein-Barr virus and lymphoid cells: An assay for early events in viral infection
Authors: Rosenthal Ken S. , Yanovich Saul , Inbar Michael , Strominger Jack L.
Published: May 2006

Translocation of the hydrocarbon fluorescent probe diphenylhexatriene (DPH) between membranes was studied by fluorescence polarization (
P
) analysis. First, using a model system, the high
P
value (0.324) of DPH-labeled cholesterol/phosphatidylcholine liposomes and the low
P
value (0.157) of DPH-labeled phosphatidylcholine liposomes allowed detection of DPH translocation between interacting liposomes. This was monitored by the change in
P
in either direction. Early events during cell-virus interactions were similarly studied by monitoring DPH translocation. The
P
value of DPH-labeled Epstein-Barr Virus (EBV) was significantly higher (0.350-0.392) than the
P
value of DPH-labeled lymphoid cells (0.238-0.289). Hence, DPH translocation could be detected by changes in
P
following incubation of DPH-labeled EBV and nonlabeled cells. A marked decrease in
P
was observed after incubation of DPH-labeled EBV with either nonlabeled lymphoblastoid Raji cells or fresh human B lymphocytes. However, only a slight decrease in
P
was obtained when DPH-labeled EBV was incubated with either nonlabeled fresh human T lymphocytes or fresh T or B rabbit lymphocytes. Moreover, incubation of fresh human B lymphocytes with the purified C3 component of complement (a putative inhibitor for the EBV receptor) prior to the addition of DPH-labeled EBV abolished the observed decrease in the
P
value. Most of these experiments were carried out with both the P3HR-1 and the B95-8 strains of EBV. DPH translocation, as determined by fluorescence polarization analysis, is, therefore, measuring some early event during interaction of this enveloped virus and mammalian cells. The potential applicability of this technique to other viruses is illustrated by an experiment with Semliki Forest virus.
Isolation of malaria merozoites: release of Plasmodium chabaudi merozoites from schizonts bound to immobilized concanavalin A.
Authors: David P H , Hommel M , Benichou J C , Eisen H A , da Silva L H
Published: May 2006

The ability of concanavalin A to bind erythrocytes but not malarial parasites was used for the development of a method of merozoite isolation: cells from infected blood were allowed to bind to a column of concanavalin A linked to Sepharose beads and merozoites naturally released by maturation of the schizonts bound to the gel were collected. The principle of this method allows its application to several Plasmodium species. The kinetics of merozoite production and the quality of the preparations (purity, infectivity, and ultrastructural morphology) were investigated by using Plasmodium chabaudi.
Estimation of expected number of rare alleles of a locus and calculation of mutation rate
Authors: Rothman Edward D. , Adams Julian
Published: May 2006

An approach is described for the estimation of the number of rare variants in the population from the number in a sample drawn at random from the population. This quantity is used to derive an estimate of the mutation rate. The data required are the number of rare variants in the sample and the distribution of offspring within a population of well-defined size with little or no immigration. Application of this approach to data on 28 loci assayed in the Yanamamo, a tribe of South American Indians, yields an average mutation rate of 0.1 ∼ 0.2 × 10
-5
. Determination of this figure is subject to several assumptions concerning the nature of the rare variants and the structure of the population. Violation of these assumptions will generally result in the underestimate of the true mutation rate.
Spontaneous and antibody-dependent cell-mediated cytotoxicity by human T cell subpopulations.
Authors: Gupta S , Fernandes G , Nair M , Good R A
Published: May 2006

Human peripheral blood non-T cells, T cells and their subpopulations (Tmu, Tgamma, Tphi, Tgamma-depleted cells, and Tmu-depleted cells) were assayed for their capacity to mediated spontaneous lymphocyte-mediated cytotoxicity (SLMC) or natural killer activity against K562 tumor cell line and antibody-dependent cellular cytotoxicity (ADCC) against chicken erythrocytes coated with antibody. Non-T cells, unseparated T cells, Tgamma cells, and Tmu-depleted (Tgamma-enriched) cells were found to have both SLMC (NK activity) and ADCC. Tmu, Tphi, and Tgamma-depleted cells had minimal or no SLMC and ADCC activity. This study demonstrates that SLMC and ADCC activity in T cells is mediated by Tgamma cell subpopulations. These two cytotoxic reactions were either mediated by two distinct subsets of Tgamma cells or by a single effector cell using two different mechanisms.
Growth cone formation in cultures of sensory neurons.
Authors: Bray D , Thomas C , Shaw G
Published: May 2006

Three experimental situations have been found in which cultured sensory neurons from embryonic chicken will form growth cones from positions along the length of the neurite. If the neurons are dissected with a remaining short axonal stump and plated into serum-free medium, they can form a morphologically normal growth cone from the stump within 15 min, even in the presence of cycloheximide or puromycin. When neurites growing in culture media with low levels of serum are cut at any point with microneedles, growth cones are produced quickly from the amputated stump, usually within 20 min. Treatment of growing neurons with low concentrations of colchicine, Colcemid, or podophyllotoxin results in the progressive appearance of lateral filopodia and regions of flattened cytoplasm that closely resemble growth cones except for their preterminal positions. These observations show that the potential to form growth cones is distributed throughout the neuron and suggest that this normally repressed in some way by the neuronal microtubules.
Adenosine analogs inhibit adipocyte adenylate cyclase by a GTP-dependent process: basis for actions of adenosine and methylxanthines on cyclic AMP production and lipolysis.
Authors: Londos C , Cooper D M , Schlegel W , Rodbell M
Published: May 2006

Adenylate cyclase in purified membranes from rat adipocytes is inhibited by low concentrations of purine-modified adenosine analogs, particularly those modified in the N6 position. Such inhibition is antagonized competitively by methylxanthines, but not by other cyclic nucleotide phosphodiesterase inhibitors, and it is dependent on "inhibitory" concentrations of GTP in the assay medium. Ribose-modified adenosine analogs inhibit adenylate cyclase through a process that is neither dependent upon the GTP concentration nor antagonized by methylxanthines. These results explain the potent effects of adenosine and methylxanthines on fat cell metabolism and demonstrate the importance of GTP in mediating inhibition by agents that act at cell surface receptors.
Rapid intramolecular coupling of active sites in the pyruvate dehydrogenase complex of
Escherichia coli
: Mechanism for rate enhancement in a multimeric structure
Authors: Danson Michael J. , Fersht Alan R. , Perham Richard N.
Published: May 2006

In the absence of CoA and presence of pyruvate, the lipoic acid residues covalently bound to the lipoate acetyltransferase core component (acetyl-CoA:dihydrolipoate
S
-acetyltransferase, EC 2.3.1.12) of the pyruvate dehydrogenase multienzyme complex of
Escherichia coli
become reductively acetylated. A study of a series of reassembled complexes varying only in their content of pyruvate decarboxylase [pyruvate:lipoate-oxidoreductase (decarboxylating and acceptor-acetylating) EC 1.2.4.1] showed that the initial direct reductive acetylation of lipoic acid residues can be followed by extensive intramolecular transacetylation reaction between lipoic acid residues on neighboring polypeptide chains of the lipoate acetyltransferase core [Bates, D. L., Danson, M. J., Hale, G., Hooper, E. A. & Perham, R. N. (1977)
Nature
(
London
) 268, 313-316]. Pulsed-quenched-flow measurements of the rates of the acetylation reactions in the various complexes now demonstrate that the intramolecular transacetylation reactions are not rate-determining in the normal reaction mechanism of the enzyme. There is therefore the potential for rapid multiple coupling of active sites in the lipoate acetyltransferase core. The rate constant for the overall complex reaction, measured by stopped-flow fluorimetry, is found to be approximately twice that for the reductive acetylation reaction measured by pulsed-quenched flow. This result could mean that CoA is an allosteric stimulator of the reductive acetylation part of the overall reaction or that there are two active sites on each chain of the lipoate acetyltransferase component working in parallel. A system of rapid functional connection of active sites in a multienzyme complex ensures that sequential reactions can be successfully coupled even under conditions of low substrate concentrations for the different steps. The substantial rate enhancement thus achieved offers a plausible explanation for the unusual complexity of the quaternary structure of the enzyme.
Sequence of oxygen binding by hemoglobin
Authors: Asakura Toshio , Lau Pui-wah
Published: May 2006

A nitroxide spin-label probe was attached directly to a propionic acid group of heme in either the α or the β chain of hemoglobin. The electron paramagnetic resonance (EPR) spectrum of the spin label is altered by the spin-state change of the heme iron to which the spin label is attached. These hybrid hemoglobins showed normal optical and functional properties, indicating that the attachment of the spin label did not perturb the function of hemoglobin. Upon deoxygenation of α-heme-spin-labeled hemoglobin, EPR signals changed proportionally with oxygen saturation (determined by measuring absorption spectra). This result indicates that there is no binding preference between the α and β chains of hemoglobin. However, the cross plot for the fraction of the EPR changes vs. the fraction of oxygen saturation deviated significantly from the diagonal straight line in response to the addition of 2,3-diphosphoglycerate and inositol hexaphosphate. The deviation indicated that the EPR change precedes the optical change at low oxygen tension. This result implies that, in the presence of organic phosphate, oxygen binds preferentially to the α subunit of deoxyhemoglobin. This conclusion was supported by the result obtained with β-heme-spin-labeled hemoglobin: the direction of the deviation for β-heme-spin-labeled hemoglobin in the presence of diphosphoglycerate and inositol hexaphosphate was opposite to that obtained for α-heme-spin-labeled hemoglobin. However, the curve deviated even in the absence of organic phosphate. This deviation for β-heme-spin-labeled hemoglobin can be explained by the intersubunit interaction of hemoglobin. From these results, it was concluded that, in the absence of organic phosphate, oxygen combines with the α and β chains with equal probability whereas, in the presence of organic phosphate, oxygen binds preferentially to the α chains of hemoglobin.
Characterization of a nontrypsin cholecystokinin converting enzyme in mammalian brain
Authors: Straus Eugene , Malesci Alberto , Yalow Rosalyn S.
Published: May 2006

An enzyme has been partially purified from canine and porcine cerebral cortical extracts that differs from trypsin in that it manifests some degree of hormone specificity since it converts porcine cholecystokinin to smaller immunoreactive forms, i.e., the COOH-terminal dodecapeptide and octapeptide fragments, but fails to convert big gastrin (34 amino acids) to heptadecapeptide gastrin. This enzyme is distinguishable from trypsin not only in substrate specificity, but also in several physiochemical properties. It is not inhibited in the presence of concentrations of lima bean trypsin inhibitor sufficient to inhibit 1 mg of trypsin per ml of incubation mixture. It is inactivated when incubated with substrate at 45°C for 1 hr, whereas trypsin remains fully active when incubated under the same conditions at 55°C. The enzyme elutes in the void volume on Sephadex G-50 and G-75 gel filtration. On sucrose gradient centrifugation, the proteolytic activity associated with trypsin is recovered above albumin but that of the solubilized brain enzyme is recovered below gamma globulin. The enzyme is not detectable in splenic extracts, which do contain nonspecific proteases capable of completely degrading cholecystokinin. Further investigation is required to determine whether the enzyme in the gut that converts cholecystokinin to the bioactive and immunoactive COOH-terminal fragments resembles or is different from the brain converting enzyme.
A model for hydration of peptides and its application to the conformational analysis of terminally blocked amino acids and dipeptides
Authors: Némethy George , Hodes Zachary I. , Scheraga Harold A.
Published: May 2006

A theoretical model for peptide structure, which takes into account the effects of hydration in conformational energy calculations, is described. The free energy of hydration is composed of a term for “specific hydration,” representing solute-water hydrogen bonding, and a term for “non-specific hydration,” describing the interaction of the solute with water molecules in a first-neighbor shell. Minimum-energy conformations were computed for the hydrated
N
-acetyl-
N
′-methylamides of the 20 naturally occurring amino acids, and the results were compared with those computed in the absence of hydration. The relative energies of many conformations and the width of some low-energy regions of the (ø, Ψ) conformational maps are altered when the free energy of nonspecific hydration is included. The term for specific hydration causes large charges of the energy, but only in
some
regions of the maps. Observed vicinal coupling constants are approximated better by the computation when hydration is included. Conformational preferences of the individual residues in hydrated dipeptides are similar to those computed for the hydrated single residues, showing that intraresidue interactions predominate in dipeptides. This supports the concept of the importance of short-range interactions in proteins. Bend probabilities were computed and compared with observed frequencies of occurrence of bends in proteins of known structure. Computed values improve only for some of the dipeptides containing polar residues or glycine when hydration is included. For bends involving two nonpolar residues, computations omitting hydration give better results.
Epidermal growth factor: Biological activity requires persistent occupation of high-affinity cell surface receptors
Authors: Shechter Yoram , Hernaez Lydia , Cuatrecasas Pedro
Published: May 2006

The enhancement of DNA synthesis by epidermal growth factor (EGF) in cultured human fibroblasts is demonstrable 24 hr after incubation of the cells at 37°C with very low concentrations (0.83 nM) of the hormone for very short periods (30 min) followed by thorough washing of the cells to remove the free hormone in the medium. This effect must result from persistent, extraordinarily tight binding of the hormone to surface receptors, because the addition of specific, purified anti-EGF IgG as late as 8 hr after initial hormone exposure can completely reverse the biological effects of the hormone. This causes only a slight (but significant) increase in the rate of dissociation at 37°C of the cell-bound
125
I-labeled EGF at low occupancy. Together with the fact that in the presence or absence of antibody virtually all of the demonstrable cell-bound
125
I-labeled EGF can be shown to dissociate from the cell during a period as short as 2-3 hr, the data suggest the possibility that the biological effects of this hormone may be mediated by occupation of only a negligible fraction of very high affinity binding sites. Thus, the processes of hormone internalization, degradation, and “down regulation” may be irrelevant to the effects of the hormone on DNA synthesis. For this effect the crucial and limiting processes appear to be strictly related to the continuous and persistent occupation of cell surface receptors.
Interferon action: two distinct pathways for inhibition of protein synthesis by double-stranded RNA.
Authors: Farrell P J , Sen G C , Dubois M F , Ratner L , Slattery E , Lengyel P
Published: May 2006

Double-stranded RNA inhibits protein synthesis in at least two ways. It activates a protein kinase that blocks peptide chain initiation by phosphorylating the peptide chain initiation factor eIF-2 and also activates an endonuclease that inactivates different mRNAs at different rates. The protein kinase and the endonuclease have been partially purified from interferon-treated Ehrlich ascites tumor cells. The 2',5'-oligoadenylates [pppA(2'p5'A)n], found found earlier to be mediators in the activation of the endonuclease by double-stranded RNA, are not mediators in the activation of the protein kinase by double-stranded RNA.
A gene adjacent to satellite DNA in Drosophila melanogaster.
Authors: Carlson M , Brutlag D
Published: May 2006

Several copies of a sequence adjacent to 1.688 g/cm3 satellite DNA in the Drosophila melanogaster genome have been isolated by molecular cloning. This sequence, called the Dm142 gene, is homologous to a 1.6-kilobase RNA found in both D. melanogaster embryos and tissue culture cells. One cloned DNA segment includes two copies of the Dm142 gene and 1.688 g/cm3 satellite DNA sequences, which are located between and flanking both gene copies. The Dm142 gene is repeated many times in the D. melanogaster genome, and some copies are not flanked by 1,688 g/cm3 satellite DNA.
Protein factor that inhibits binding and promotes release of androgen-receptor complex from nuclear chromatin.
Authors: Shyr C I , Liao S
Published: May 2006

A protein factor that can inhibit binding of the androgen-receptor complex to the nuclear chromatin has been isolated from the cytosol of the ventral prostate of rats. The inhibition is reversible and not caused by an irreversible destruction of the receptor or the chromatin. The inhibitor also can promote, by a temperature-dependent process, the release of steroid-receptor complex already bound to chromatin. It is conceivable that such a protein factor plays a regulatory role in the nuclear-cytoplasmic recycling or chromatin binding of the steroid hormone receptor.