bluuebunny/crossref_metadata_2025 huggingface dataset:
Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor.
Authors: Greene L A , Tischler A S
Published: May 2006

A single cell clonal line which responds reversibly to nerve growth factor (NGF) has been established from a transplantable rat adrenal pheochromocytoma. This line, designated PC12, has a homogeneous and near-diploid chromosome number of 40. By 1 week's exposure to NGF, PC12 cells cease to multiply and begin to extend branching varicose processes similar to those produced by sympathetic neurons in primary cell culture. By several weeks of exposure to NGF, the PC12 processes reach 500-1000 mum in length. Removal of NGF is followed by degeneration of processes within 24 hr and by resumption of cell multiplication within 72 hr. PC12 cells grown with or without NGF contain dense core chromaffin-like granules up to 350 nm in diameter. The NGF-treated cells also contain small vesicles which accumulate in process varicosities and endings. PC12 cells synthesize and store the catecholamine neurotransmitters dopamine and norepinephrine. The levels (per mg of protein) of catecholamines and of the their synthetic enzymes in PC12 cells are comparable to or higher than those found in rat adrenals. NGF-treatment of PC12 cells results in no change in the levels of catecholamines or of their synthetic enzymes when expressed on a per cell basis, but does result in a 4- to 6-fold decrease in levels when expressed on a per mg of protein basis. PC12 cells do not synthesize epinephrine and cannot be induced to do so by treatment with dexamethasone. The PC12 cell line should be a useful model system for neurobiological and neurochemical studies.
Modulation of the immune response by a synthetic adjuvant and analogs.
Authors: Chedid L , Audibert F , Lefrancier P , Choay J , Lederer E
Published: May 2006

N-Acetylmuramyl-L-alanyl-D-isoglutamine increases the humoral immune response of mice when given in aqueous media instead of the usual water-in-oil emulsions. Moreover, this compound is adjuvant active even by the oral route. In view of studying the relation between chemical structure and biological activity, several synthetic analogs were tested. The immune response could be modulated according to chemical modifications, and the synthetic analog with D- in place of L-alanine was shown to inhibit the immune response.
Role of major histocompatibility complex gene products in delayed-type hypersensitivity.
Authors: Miller J F , Vadas M A , Whitelaw A , Gamble J
Published: May 2006

Sensitized thymus-derived (T) lymphocytes can transfer delayed-type hypersensitivity (DTH) to naive mice only if there is identity at the major histocompatibility complex (MHC). The MHC region responsible differs according to the antigen used for sensitization. For transfer of DTH to fowl gamma globulin identity at I-A is necessary; for dinitrofluorobenzene, however, identity at either K, D, or I region is sufficient. T cells of one genotype, sensitized in a chimeric environment, transferred DTH to both parental strains even though these were MHC incompatible. However T cells from F1 hybrid mice, sensitized not in the F1 but in one parental strain, transferred DTH only to that parental strain, not to the other, in contrast to F1 T cells sensitized in the F1 which could transfer DTH to both parental strains. Macrophages pulsed with antigen in vitro could be used to sensitize syngeneic or semi-allogeneic mice for the transfer of DTH. Transfer was, however, successful only in the strain syngeneic to that from which the macrophages were derived. Evidence is also presented that genetically low-responder mice can be made to exhibit DTH provided they are pretreated with cyclophosphamide two days before sensitization. When considered in toto these results strongly argue in favor of the notion that there are receptors on activated T cells which recognize antigenic determinants and MHC gene products. The implications of these findings are discussed in relation to the role of macrophages in antigen presentation and to the possible parallel evolution of MHC gene products and of T cell receptors for antigen.
Short-lived methylated messenger RNA in mouse kidney.
Authors: Ouellette A J , Reed S L , Malt R A
Published: May 2006

In experiments originally designed to examine selective turnover of methylated "caps" in renal mRNA, we observed that [3H]methyl label decayed from mRNA containing poly(A) with a half-life of 1-2 hr. (Caps are blocked, methylated mRNA sequences of the general structure m7GpppNm p(1 or 2)Np.). To distinguish between metabolism of short-lived mRNA and discriminate turnover of "caps", we compared residual [3H]methyl label in 5' and 3'mRNA fragments prepared from mRNA isolated during the decay period. Hydrolysis of mRNA at 0 degrees with dilute KOH before oligo(dT)-cellulose selection produced 5' mRNA fragments enriched with an alkali-resistant oligonucleotide with a -5 charge; the 3' mRNA fraction was correspondingly reduced in oligonucleotide content. Since methyl label disappeared at the same rate from both fractions, we conclude that mouse kidney contains short-lived mRNA and that the "caps" of these labile mRNAs turn over with the rest of the mRNA molecule.
Detection of different types of damage in alkylated DNA by means of human corrective endonuclease (correndonuclease).
Authors: Duker N J , Teebor G W
Published: May 2006

Corrective endonuclease (correndunclease) activity of HeLa cells was assayed with alkylated DNA. Double-stranded, covalently closed DNA from phage PM II was treated with methyl methanesulfonate, N-methyl-N-nitrosourea, beta-propiolactone, or diepoxybutane to introduce alkylated bases and alkali-labile sites into the DNA. The damaged DNA was incubated with an extract of HeLa cells that catalyzes the formation of breaks at apurinic sites in double-stranded DNA. Methylated DNA was broken at every alkali-labile site by the HeLa correndonuclease, which indicated that these sites are similar to the apurinic sites produced by heating at acid pH. DNA alkylated with beta-propiolactone or diepoxybutane containing the same number of alkali-labile sites was broken to a far lesser extent. This indicates the presence of a second type of alkali-labile damage that is correndonuclease-insensitive.
ATP binding to a protease-resistant core of actin.
Authors: Jacobson G R , Rosenbusch J P
Published: May 2006

Actin can be cleaved by trypsin or chymotrypsin into a large, autonomous fragment with approximately 80% of the mass of the undegraded polypeptide. The protease-resistant cores obtained with either enzyme are very similar. Although the fragment does not bind calcium ions and fails to polymerize to the filamentous form of actin or to stimulate myosin adenosine triphosphatase (ATP phosphohydrolase, EC 3.6.1.3) activity, it retains the full capacity to bind ATP. This observation suggests that it represents an independent functional unit. Cleavage of globular actin with either trypsin or chymotrypsin occurs with half-times of 3 min, while that of filamentous actin proceeds with reaction half-times of 20 min for trypsin and nearly 2 hr for chymotrypsin. Denaturation and renaturation of the trypsin-resistant core shows that approximately 20% of the molecules refold to functional forms which indicates that the fragment can be considered as an independent unit of folding as well.
Chromosome-mediated gene transfer between closely realted strains of cultured mouse cells.
Authors: Degnen G E , Miller I L , Eisenstadt J M , Adelberg E A
Published: May 2006

Gene transfer between two closely related mouse cell lines has been carried out, using as the vector a cell-free preparation of metaphase chromosomes and nuclei. Distinction between gene transferents and revertants of the recipient mutant phenotype was achieved by the use of a donor strain carrying a mutationally altered (8-azaguanine-resistant) hypoxanthine-guanine phosphoribosyltransferase (HPRTase; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8). The transferred HPRTase gene is initially unstable; in nonselective medium, it is lost at a rate of about 0.1 per cell per generation. Stabilization occurs as a rare event, with a frequency on the order of 1 X 10(-5) per cell per generation. The unstable state can be maintained for at least 200 generations through serial passages of the transferent in selective medium. Under the conditions of cultivation used in these experiments, the unstable HPRTase-positive cells are eventually replaced by the stable HPRTase-positive cells in the population.
An equation of state describing hydrophobic interactions
Authors: Gill S. J. , Wadsö I.
Published: May 2006

Several thermodynamic properties for the process of dissolution of pure hydrocarbons into water are found to be linearly related to the number of hydrogens on the hydrocarbon molecule. From the correlations found for the Gibbs energy change, enthalpy change, and heat capacity change, along with the use of an average minimum solubility temperature, an equation of state for the hydrophobic effect is derived. The entropy change upon dissolution per hydrocarbon hydrogen atom is close to -
R
ln 2. A model based upon a “tetrahedrally” localized water molecule with one corner defined by a carbon-hydrogen group and the other three corners defined by water molecules is used to estimate the observed entropy and heat capacity changes.
Mechanism of nuclear DNA replication in radicles of germinating cotton
Authors: Clay Willard F. , Bartels Paul G. , Katterman Frank R. H.
Published: May 2006

The mechanism of nuclear DNA replication in radicles of germinating cotton (
Gossypium barbadense
) was investigated. Pulse-labeling with [
3
H]thymidine indicates that replication intermediates are of small molecular weight (4-10S) and behave as if single-stranded. Prolonged labeling indicates that intermediates are of discrete size, suggesting a mechanism of discontinuous replication. Electron microscopy of nuclear DNA demonstrates a complex architecture which may include “eye forms” and “forks” similar to those reported in prokaryotes and animal systems. The possible universality of DNA replication mechanisms is discussed.
Control of growth of benzo(a)pyrene-transformed 3T3 cells.
Authors: Holley R W , Baldwin J H , Kiernan J A , Messmer T O
Published: May 2006

The growth controls observed in benzo[a]pyrene-transformed 3T3 cells (BP3T3) are compared with those of virus-transformed and normal 3T3 cells. Superficially, the chemically transformed BP3T3 cells have the same behavior as virus-transformed SV3T3 cells. Both grow to high cell density in culture medium with 10% serum, both form colonies in Methocel, and both are tumorigenic. Closer examination, however, has disclosed that BP3T3 cells exhibit "normal" growth controls at low serum concentrations. In contrast to the behavior of SV3T3 cells, the initiation of DNA synthesis in BP3T3 cells is still dependent on a serum factor. If BP3T3 cells are grown in medium with 0.2% serum, the cells become quiescent, with growth arrested in the Gu or G0 phase of the cell cycle. The addition of serum or the fibroblast growth factor (FGF) to such quiescent cells leads to the initiation of DNA synthesis and the resumption of growth. As with normal 3T3 cells, if the growth rate of BP3T3 cells is limited by a suboptimal concentration of serum, the growth rate of the cells is increased by the addition of FGF. Also, BP3T3 cells show density-dependent regulation of growth, if the medium contains a low concentration of serum. BP3T3 cells, therefore, have the behavior of "transformed" cells when cultured in medium with 10% serum, but behave as "normal" cells in medium with low serum. In comparison with normal 3T3 cells, the difference in growth behavior of BP3T3 cells appears to be due to a substantial decrease in the cells' requirement for a serum growth factor of the FGF type. Exploration of possible causes of this substantial decrease indicates that the primary cause is a lower rate of depletion of the serum growth factor from the culture medium by BP3T3 cells. The decrease in rate of depletion is sufficient to account for the uncontrolled growth of BP3T3 cells in medium with 10% serum. It is suggested that a decreased rate of depletion of a growth factor may contribute to tumorigenicity of cells in vivo.
Synthesis and characterization of "face-to-face" porphyrins.
Authors: Collman J P , Elliott C M , Halbert T R , Tovrog B S
Published: May 2006

The syntheses of four binary porphyrins, two of which are constrained to a "face-to-face" conformation, and their Co2+ and Cu2+ derivatives are described. Electron spin resonance indicates that the intermetallic separation in the binuclear "face-to-face" porphyrins is about 6.5-6.8 A. Electronic spectra and proton magnetic resonance spectra support the postulated "face-to-face" conformations. A hypothesis that related compounds may serve as multielectron redox catalysts for O2 and N2 is presented.
Sensory transduction in Escherichia coli: a requirement for methionine in sensory adaptation.
Authors: Springer M S , Goy M F , Adler J
Published: May 2006

Chemotaxis of E. coli is a behavioral response to a change in the concentration of a stimulatory compound. The response is transient; thus, E. coli undergoes sensory adaptation. In this communication, we show that L-methionine is required by E. coli for adaptation to increases in the concentration of chemical attractants, but is not required for the maintenance of the adapted state. When the concentration of the attractant is lowered to its initial level, cells regain their sensitivity to the attractant. This process of deadaptation does not require methionine. We suggest that the methylation of a membrane protein, a reaction previously shown to be involved in chemotaxis [Kort, E.N., Goy, M.F., Larsen, S.H. & Adler J. (1975) Proc. Natl. Acad. Sci. USA 72, 3939-3943] underlies these phenomena.
Initiation of genetic exchanges in λ phage-prophage crosses
Authors: Lin Pin-Fang , Bardwell Eva , Howard-Flanders Paul
Published: May 2006

When
Escherichia coli
K-12 (λ) lysogens were infected with λ phages, genetic exchanges between phage and prophage occurred at low frequencies (less than 0.1% between the markers
P3
and
P80
), but at frequencies above 1% if the infecting phages were first treated with the photosensitizing agent 4,5′,8-trimethylpsoralen and 360 nm light. Exchanges were induced by psoralen damage at about the same frequency in wild-type lysogens and in those carrying
recB
-
,
recC
-
,
recF
-
, or
lexA
-
, but at an intermediate frequency in a quadruple mutant carrying
recB
-
recC
-
recF
-
sbcB
-
. Few if any exchanges were induced in lysogens carrying
uvrA
-
,
uvrB
-
, or
recA
-
. The increase in the frequency of recombination was presumably due to the psoralen damage in the phage DNA molecules and the action of host cell repair and recombination enzymes. The production of crosslinks in the phage DNA by psoralen and 360 nm light was measured by sedimentation in alkali. It showed second-order kinetics indicative of a two-photon reaction. In contrast, first-order kinetics had been reported for monoadduct formation. Second-order kinetics, similar to those for crosslink production, were found for genetic exchanges in homoimmune crosses. Presumably, crosslinks, rather than monoadducts, cause most of the exchanges. Because the
uvrA
-
gene product (UV-endonuclease) was required, it is likely that recombination was initiated by DNA molecules cut at crosslinks. This system has been used to show that after the crosslinked phage duplex has been cut, one or more of the subsequent steps—homologous pairing, cutting, and joining—require the
recA
+
gene product.
Activation of cardiac adenylate cyclase: horminal modification of the magnesium ion requirement.
Authors: Alvarez R , Bruno J J
Published: May 2006

Histamine and epinephrine stimulate the activity of guinea pig heart adenylate cyclase [ATP pyrophosphate-lyase (cyclizing) EC 4.6.1.1], in part, by decreasing the requirement for Mg2+ as an activator. This effect may represent an increase in affinity for Mg2+ and/or a decrease in sensitivity of the enzyme towards inhibition by free ATP. Both of these inotropic hormones also increase maximum velocity. Pretreatment of the membrane-bound enzyme with EDTA, to remove available divalent cations, almost eliminates persistent stimulation by guanyl-5'-yl imidodiphosphate [Gpp(NH)p]. Addition of Mg2+ to the preincubation medium restores the capacity of Gpp(NH)p to acutely activate the enzyme. These results indicate that Mg2+ interacts with the nucleotide (GTP) regulatory site. Persistent stimulation of the enzyme by either Gpp(NH)p or fluoride ion also involves a decrease in the requirement for Mg2+ and an increase in maximum velocity.
Sequences homologous to ribosomal insertions occur in the
Drosophila
genome outside the nucleolus organizer
Authors: Dawid Igor B. , Botchan Peter
Published: May 2006

Many repeating units of
Drosophila melanogaster
rDNA contain a DNA sequence within the gene for 28S rRNA that does not code for rRNA. This sequence has been called the ribosomal
insertion
[Wellauer, P. K. & Dawid, I. B. (1977)
Cell
10, 193-212]. We report here that members of the same sequence family occur outside the ribosomal locus. “Non-rDNA insertion DNA” was separated from rDNA by density gradient centrifugation, and sequences homologous to the ribosomal insertion were detected by hybridization with restriction endonuclease fragments derived from a cloned rDNA repeating unit. Pure insertion sequences from cloned rDNA separated from main band DNA and behaved like a component with high G + C content. Non-rDNA components hybridizing to the insertion also separated from main band DNA but less so than pure insertion sequences, suggesting that non-rDNA insertion sequences are linked to DNA of different nucleotide composition. Restriction endonuclease analysis of non-rDNA insertion DNA showed many fragments of different sizes. The patterns obtained were similar in embryonic, larval, pupal, and adult DNA and DNA from cultured cells (Schneider cell line 3). Non-rDNA insertion sequences account for about 0.2% of the genome or about 400 kbases of DNA per haploid complement.
Elongation factor Tu resistant to kirromycin in an
Esherichia coli
mutant altered in both
tuf
genes
Authors: Fischer Eckhard , Wolf Heinz , Hantke Klaus , Parmeggiani Andrea
Published: May 2006

A mutant of
Escherichia coli
is described that displays kirromycin resistance in a cell-free system by virtue of an altered elongation factor Tu (EF-Tu). In poly(U)-directed poly(Phe) synthesis the kirromycin resistance of the crystallized enzyme ranged between a factor of 80 and 700, depending on temperature. Similarly, kirromycin-induced EF-Tu GTPase activity uncoupled from ribosomes and aminoacyl-tRNA required correspondingly higher concentrations of the antibiotic. Resistance of EF-Tu to kirromycin is a consequence of a modified enzyme structure as indicated by its altered fingerprint pattern.

P1 transduction experiments showed that the kirromycin-resistant EF-Tu is coded by an altered
tuf
B gene (
tuf
B1). The known existence of two genes coding for EF-Tu would interfere with the recognition of a mutant altered in only one of those genes, if the mutation were recessive. Because kirromycin blocks EF-Tu release from the ribosome, kirromycin sensitivity is dominant, as shown by the failure of a mixed EF-Tu population to express resistance
in vitro
. Therefore, phenotypic expression of kirromycin resistance
in vivo
appears to be only possible if the EF-Tu mutant lacks an active
tuf
A gene, a property likely to be inherited from the parental D22 strain. The observations that introduction of a
tuf
A
+
region makes the resistant strain sensitive to the antibiotic and that transduction of
tuf
B1 into a recipient other than
E. coli
D22 yields kirromycin-sensitive progeny support these conclusions.
Comparative rates of transfer of lipid-linked oligosaccharides to endogenous glycoprotein acceptors in vitro.
Authors: Turco S J , Stetson B , Robbins P W
Published: May 2006

We have shown previously that particulate preparations of Nil 8 fibroblasts catalyze the synthesis of two oligosaccharide-lipids: one composed of N-acetylglucosamine and mannose residues and the other containing, in addition to mannose and N-acetylglucosamine, one or two glucose residues. These two oligosaccharide-lipids were purified and added to fresh microsomal preparations. In comparative studies, we find that the glucose-containing lipid-linked oligosaccharide is transferred much more rapidly to endogenous protein acceptors than the glucose-free compound. With materials of comparable specific activities, as much as 41% of the glucose-containing oligosaccharide was transferred to protein as compared to 5% for the glucose-free compound. These results suggest that the attachment of glucose to mannosyl lipid-linked oligosaccharide serves an important role in the transfer of these compounds from lipid carrier to protein acceptor.
Local control of neurite development by nerve growth factor
Authors: Campenot Robert B.
Published: May 2006

A three-chamber culture system was devised in which neurites growing from small clusters of somas of sympathetic neurons penetrated a virtually fluid-impermeable barrier; thus the local fluid environment of the distal portions of the neurites could be controlled independently of the local fluid environment of the somas and proximal portions of the neurites. Neurites regularly penetrated the barriers if a high concentration of nerve growth factor was present on both sides, but never penetrated into chambers to which no nerve growth factor had been added.

After neurites crossed the barrier, local removal of nerve growth factor from the distal portions of the neurites caused the growth of these portions to stop, and they eventually appeared to degenerate even though nerve growth factor was continuously present in the chamber that contained their somas and proximal portions. In contrast, local nerve growth factor was not required at the somas and proximal portions of the neurites; many neurons survived its withdrawal provided their somas were associated with neurite bundles that crossed into a chamber containing nerve growth factor.

These results show that the growth, and probably the survival, of neurites depends upon nerve growth factor in their local environment, regardless of the nerve growth factor concentrations to which other portions of the neuron are exposed. This is entirely consistent with the notion that nerve growth factor released by sympathetic target tissues promotes the establishment and maintenance of appropriate neuron-target connections during development.
How does selection reconcile individual advantage with the good of the group?
Authors: Leigh Egbert G.
Published: May 2006

An individual's advantage often conflicts with the good of its group, as when an allele spreads by meiotic drive through a population whose death rate it increases, or when an asexual genotype derives immediate advantage at the expense of future adaptability. We show how selection within populations may reconcile individual and group advantage, as in the evolution of “honest meioses” resistant to segregation distortion, and the avoidance of the “cost of sex.” Selection between species, whose evolutionary importance we document, presumably favors the survival and multiplication of species whose genetic systems or social organizations favor the evolution of mechanisms reconciling individual with group advantage: in other words, species with genetic or social systems where a gene's long-term selective advantage most nearly matches its contribution to the good of the species.
Precursor role of arachidonic acid in release of slow reacting substance from rat basophilic leukemia cells.
Authors: Jakschik B A , Falkenhein S , Parker C W
Published: May 2006

The release of slow reacting substance (SRS) from rat basophilic leukemia cells (RBL-1) by the ionophore A23187 (5-10 mug/ml) was stimulated 5-fold by arachidonate and inhibited 78% by 5,8,11,14-eicosatetraynoate (an inhibitor of both fatty acid cyclooxygenase and lipoxygenase). Linoleic acid and linolenic acid both inhibited SRS formation, whereas indomethacin (a cyclooxygenase inhibitor) had no effect. Radiolabel from [14C]- or [3H]arachidonate was incorporated into SRS as indicated by comigration of radioactivity and bioreactivity in several chromatographic systems after purification to apparent radiochemical homogeneity. The radiolabeled SRS was clearly separated chromatographically from other known arachidonate metabolites. Thus, SRS appears to be a previously undescribed product of arachidonic acid metabolism, probably formed through the lipoxygenase pathway. The ability to prepare purified, biosynthetically labeled, SRS should be of considerable help in further studies of its structure, biologic function, and catabolism.
Human leukocytic pyrogen: purification and development of a radioimmunoassay.
Authors: Dinarello C A , Renfer L , Wolff S M
Published: May 2006

Leukocytic pyrogen is a small endogenous protein that mediates fever. Because of the limitations of bioassays, circulating leukocytic pyrogen has not been demonstrated during fever in humans. The pyrogen was produced in vitro after phagocytosis of staphylococci by blood monocytes. Antibody against the pyrogen was obtained from rabbits immunized with leukocytic pyrogen and the antiserum was purified by solid-phase immunoadsorbants. Purified antibody to the pyrogen was attached to activated Sepharose 4B and used in conjunction with gel filtration to purify the pyrogen. The pyrogen was labeled with 125I and further purified by gel filtration and ion-exchange chromatography. The final preparation of 125I-labeled pyrogen demonstrated a homogeneous band during isoelectric focusing and other separation procedures. With antibody to pyrogen attached to Sepharose, less than 0.1 of a rabbit pyrogenic dose of human leukocytic pyrogen inhibited the binding of 125I-labeled pyrogen to this immunoadsorbant, and this inhibition was not affected by the presence of human serum. Thus, a radioimmunoassay for human leukocytic pyrogen has been developed that may be used to detect circulating pyrogen during fever in humans.
Sedimentation properties of the scrapie agent.
Authors: Prusiner S B , Hadlow W J , Eklund C M , Race R E
Published: May 2006

The sedimentation behavior of the scrapie agent in homogenates of spleen from infected mice has been determined. Approximately 90% of the scrapie agent was sedimented at an omega2t value of 3 X 10(10) rad2/sec in a fixed-angle rotor. Sedimentation of the agent was not substantially affected by sonication or by treatment with the detergent sodium deoxycholate. The sedimentation profiles of the scrapie agent were similar to those observed for free polyribosomes, but differed from those exhibited by five other subcellular markers. Comparative studies showed that the sedimentation profiles of subcellular markers in spleen suspensions from mice infected with scrapie did not differ from uninoculated controls. These studies suggest that the scrapie agent is a discrete infectious particle which should be separable from cellular membranes.
On a sharpened form of the Schauder fixed-point theorem
Authors: Browder Felix E.
Published: May 2006

If
K
is a compact convex subset of a locally convex topological vector space
X
, we consider a continuous mapping
f
of
K
into
X
. A fixed-point theorem is proved for such a map
f
under the assumption that for a given continuous realvalued function
p
on
K
×
X
with
p
(
x,y
) convex in
y
and for each point
x
in
K
not fixed by
f
, there exists a point
y
in the inward set

I
K

(
x
) generated by
K
at
x
with
p(x,y - f(x))
less than
p(x,x - f(x))
. For
X
a Banach space, in particular, this yields a sharp extension and a drastic simplification of the fixed point theory of weakly inward (and weakly outward) mappings. The result comes close in the domain of mappings of compact convex sets to the thrust of fixed point conditions of the Leray-Schauder type for compact maps of sets with interior in
X
.
Complete amino acid sequence of rabbit muscle glycogen phosphorylase.
Authors: Titani K , Koide A , Hermann J , Ericsson L H , Kumar S , Wade R D , Walsh K A , Neurath H , Fischer E H
Published: May 2006

The sequence of the 841 amino acid residues in each subunit (molecular weight 97,412) of rabbit muscle glycogen phosphorylase b (1,4-alpha-D-glucan:orthophosphate alpha-glucosyltransferase; EC 2.4.1.1) has been determined. The general strategy was based on limited proteolysis of native phosphorylase b by subtilisin BPN', yielding two large segments (light and heavy) which were fragmented by cleavage at methyonyl-, asparaginyl-glycine, and aspartyl-proline bonds. Analysis of two cyanogen bromide fragments (CB14 and CB17) isolated from the intact molecule yielded the overlap between the light and heavy fragments and the remainder of the sequence. The residues involved in the covalent and allosteric control of the enzyme, and in the binding of the cofactor pyridoxal 5'-phosphate, were identified as serine-14, tyrosine-155, and lysine-679, respectively.
Role of nucleotides in tubulin polymerization: effect of guanylyl 5'-methylenediphosphonate.
Authors: Sandoval I V , MacDonald E , Jameson J L , Cuatrecasas P
Published: May 2006

Incubation of 48,000 X g rat brain supernatants for 30 min at 37 degrees with 1-2 mM guanylyl 5'-methylenediphosphonate [Gmp(CH2)pp] results in polymerization of 95-98% of the tubulin present. This is considerably more than the 50% polymerization that can be achieved with the natural nucleotide, GTP, under optimal conditions. Gmp(CH2)pp is also much more effective than GTP in inducing polymerization of purified tubulin. Assembly of microtubules with Gmp(CH2)pp occurs at tubulin concentrations one-third of those possible with GTP. Furthermore, with Gmp(CH2)pp, microtubule assembly does not require the high molecular weight basic proteins needed with GTP. Polymerization of tubulin by Gmp(CH2)pp is neither prevented nor reversed by concentrations of calcium (2 mM) that can either prevent microtubule assembly or disrupt already formed microtubules if the nucleotide used is GTP or guanylyl imidodiphosphate. When Ca2+ is added before or after microtubule assembly, electron microscopy of the Gmp(CH2)pp preparations reveals normal microtubules turning into twisted ribbons. Low temperature (4 degrees) can both prevent and disrupt the tubulin assembled Gmp(CH2)pp although disruption proceeds much more slowly when GTP is used.
Isopeptide linkage between N-alpha-monomethylalanine and lysine in ribosomal protein S11 from Escherichia coli.
Authors: Chen R , Chen-Schmeisser U
Published: May 2006

Protein S11 from the Escherichia coli ribosome has a unique NH2-terminal structure not previously observed among ribosomal proteins. Owing to the formation of an isopeptide bond between a secondary amino acid (N-alpha-monomethylalanine) and the epsilon-amino group of the NH2-terminal lysine residue, a "branching point" is formed. Therefore, two amino acids are seen when the NH2 terminus of the protein is determined.
31
P nuclear magnetic resonance studies of HeLa cells
Authors: Evans Frederick E. , Kaplan Nathan O.
Published: May 2006

A survey of phosphorus compounds present in HeLa cells and their acid extracts has been carried out by
31
P nuclear magnetic resonance spectroscopy at 40 MHz. The proton decoupled
31
P spectrum of the neutralized extract had resolution adequate to enable the identification of the main phosphate compounds. The spectral intensities were converted to concentrations. The lower detection limit with extensive signal averaging was 0.02 μmol for the extract. The composition, listed in order of decreasing concentration, was: inorganic phosphate, ATP, phosphorylcholine, creatine phosphate, UTP, NAD
+
, glucose 6-phosphate, β-D-fructose 1,6-bisphosphate, α-D-fructose 1,6-bisphosphate, ADP, α-glycerophosphorylcholine, and α-glycerophosphorylethanolamine. UTP made up ⅕ of the total nucleotide triphosphate content. The composition was compared to the
31
P spectrum of an extract from a human astrocytoma grown in athymic mice. The signal from P-containing macromolecules such as nucleic acids was not detected in the intact HeLa cell spectrum because of broad lines. Effects of the glycolysis inhibitor iodoacetic acid could be clearly shown in spectra of both the intact cell and the extract as buildup of fructose 1,6-bisphosphate at the expense of ATP, UTP, and creatine phosphate.
Candida lipolytica mutants defective in an acyl-coenzyme A synthetase: isolation and fatty acid metabolism.
Authors: Kamiryo T , Mishina M , Tashiro S I , Numa S
Published: May 2006

Mutant strains of Candida lipolytica defective in an acyl-CoA synthetase [acid:CoA ligase (AMP-forming); EC 6.2.1.3]were isolated. The mutant strains apparently exhibited no acyl-CoA synthetase activity in vitro and were, in contrast to the wild-type strain, incapable of growing in the presence of exogenous fatty acid when cellular synthesis de novo of fatty acid was blocked. However, the mutant strains grew on either fatty acid or n-alkane as a sole carbon source at rates comparable to that observed for the wild-type strain. Analysis of the fatty acid composition of the lipids from the mutant cells grown on odd-chain-length fatty acid as well as [14C]oleic acid incorporation studies have shown that the mutant cells, unlike the wild-type cells, cannot incorporate exogenous fatty acid as a whole into cellular lipids, but utilize the fatty acid that is synthesized de novo from acetyl-CoA produced by degradation of exogenous fatty acid. This finding indicates the presence of at least two acyl-CoA synthetases that activate long-chain fatty acid. One, designated acyl-CoA synthetase I, which is absent in the mutant strains, is responsible for the production of acyl-CoA to be utilized for the synthesis of cellular lipids. The other acyl-CoA synthetase provides actyl-CoA that is exclusively degraded via beta-oxidation to yield acetyl-CoA.
Cell-free synthesis of the precursor polypeptide for avian myeloblastosis virus DNA polymerase.
Authors: Paterson B M , Marciani D J , Papas T S
Published: May 2006

High molecular weight RNA (35S) isolated from avian myeloblastosis virus directs the cell-free synthesis of two prominent polypeptides of 180,000 and 76,000 molecular weight. The latter polypeptide has previously been identified as the precursor to the group-specific antigens of the virus ("gag" proteins) [Vogt, V. M., Eisenman, R. & Diggelmann, H. (1975) J. Mol. Biol. 96, 471-493]. Two-dimensional tryptic peptide analyses of the [35S]methionine-labeled peptides demonstrate that the 180,000-dalton product is a polyprotein that can account for all the peptides of the avian myeloblastosis virus DNA polymerase (DNA nucleotidyltransferase, EC 2.7.7.7) and those of the gag viral proteins. This is direct confirmation of the genomic order of the viral structural genes, placing the polymerase gene adjacent to the 5'-proximal gag gene of the virus. Furthermore, our findings suggest that the primary polymerase gene product is the beta subunit of the enzyme. These results are discussed in relation to the proposed structural gene map for the avian retraviruses and suggest a model for the in vivo processing of the viral polymerase.
Sensory transduction in
Escherichia coli
: Role of a protein methylation reaction in sensory adaptation
Authors: Goy Michael F. , Springer Martin S. , Adler Julius
Published: May 2006

The behavioral response of
Escherichia coli
to the addition of a stimulatory compound is transient; thus the organism undergoes sensory adaptation. When the compound is removed,
E. coli
undergoes the inverse process, called deadaptation, and very rapidly regains its sensitivity to the stimulus. In this communication we demonstrate that the previously reported methylation of several cytoplasmic membrane proteins is correlated with, and very likely controls, the state of adaptation of the cell.

In the absence of an added stimulus these proteins are methylated to a basal level. When the bacteria are stimulated by the addition of an attractant, the extent of methylation increases over a period of several minutes to a new level, which is maintained as long as the attractant is present. The magnitude of the increase in methylation is a function of the size of the stimulus and is directly proportional to the duration of the behavioral response. Upon removal of the attractant the level of methylation very rapidly falls to the basal value. Previously we have shown that adaptation requires methionine, but maintenance of the adapted state and de-adaptation do not [Springer, M. S., Goy, M. F. & Adler, J. (1975)
Proc. Natl. Acad. Sci. USA
74, 183-187]; here we demonstrate that methylation requires methionine but maintenance of an attractant-induced level of methylation and the demethylation that occurs following removal of the attractant do not. These results strongly indicate a role for protein methylation in sensory adaptation.
Transmembrane interactions and the mechanism of capping of surface receptors by their specific ligands.
Authors: Bourguignon L Y , Singer S J
Published: May 2006

The mechanism of capping of cell surface receptors has been examined by a double fluorescence staining procedure that permitted simultaneous observations of the distribution of a surface-bound ligand together with intracellular actin or myosin. At an early stage in the capping of the T-25 antigen or the H2 histocompatibility antigens on mouse splenic T lymphocytes, or of concanavalin A receptors on HeLa cells, when the specific receptors in question were collected into patches that were distributed over the entire cell surface, the intracellular membrane-associated actin or myosin was also accumulated into patches that were located directly under the receptor patches. These and other results have led us to propose a general molecular mechanism for the process of capping, in which actin and myosin are directly involved. It is suggested that membrane-associated actin is directly or indirectly bound to an integral protein or class of proteins, X, in the plasma membranes of eukaryotic cells. When any receptor in the membrane is aggregated by an external multivalent ligand, the aggregate binds effectively to X, whereas unaggregated receptors do not bind to X. The receptor aggregates, linked to actin (and myosin) through X, are then actively collected into a cap by an analogue of the actin--myosin sliding filament mechanism of muscle contraction.
Translation of
Drosophila melanogaster
sequences in
Escherichia coli
Authors: Rambach Alain , Hogness David S.
Published: May 2006

Thirty-seven independently cloned segments of
Drosophila melanogaster
DNA (Dm segments) were individually tested for their ability to promote the synthesis of new polypeptides in
Escherichia coli
K-12. The cloning vector was the pSC101 plasmid and the test system consisted of
E. coli
K-12 minicells that contained the hybrid pDm plasmids. Each of four pDm plasmids produced a new polypeptide, and one, pDm107, was selected for detailed mapping of the sequences required for the translation of its 38,000-dalton polypeptide, the Dm107 protein. Mapping was accomplished by constructing (
i
) deletion derivatives of pDm107 and (
ii
) new plasmids consisting of fragments of the Dm107 segment inserted into other vectors, and then testing these hybrids for their ability to promote the synthesis of the Dm107 protein, or truncated versions of this protein, in minicells. The 1000 base pairs of sequences that are translated to yield the Dm107 protein were thereby mapped at the center of the 18,000-base pair Dm107 segment, which consists of nonrepetitive sequences located at the base of the right arm of chromosome 2. The four polypeptides produced by the four pDm plasmids require sequences of 4000 base pairs for their translation, and the total amount of DNA in the 37 cloned Dm segments that were tested is approximately 400,000 base pairs. Because no new polypeptides were detected with the remaining 33 pDm plasmids, the fraction of
D. melanogaster
sequences that can be efficiently translated in
E. coli
K-12 is estimated to be 1 × 10
-2
.
Idiotypic-anti-idiotypic regulation of antibody synthesis in rabbits.
Authors: Cazenave P A
Published: May 2006

Anti-ribonuclease antibodies (Ab1) of one rabbit were used to prepare in two other rabbits anti-idiotypic antibodies (Ab2) against them. Anti-idiotypic antibodies (Ab3) were prepared against Ab2 of one rabbit in two other rabbits that had the same phenotype as the former three for the allotypic a and b systems. After a rest period of 4 months, the latter two rabbits were immunized against ribonuclease. The antibodies produced (Ab1') crossreacted with the two anti-idiotypic sera (Ab2) against Ab1, showing an idiotypic similarity between anti-ribonuclease Ab1' and anti-ribonuclease Ab1. No such crossreaction was observed between the anti-idiotypic sera (Ab2) against Ab1 and any of the 12 unrelated antisera against ribonuclease that were tested. These results suggest (i) that rabbits, at least with the same allotypes, possess a closely related idiotypic repertoire; and (ii) that the immune system is a network of variable (V)-domains.
Dissociated ciliary ganglion neurons in vitro: survival and synapse formation.
Authors: Nishi R , Berg D K
Published: May 2006

Normally, about half of the ciliary ganglion neurons in 8-day-old chick embryos die before day 14 in ovo. However, when dissociated ciliary ganglion neurons were prepared from either 8- or 14-day-old embryos and grown in cell culture with skeletal myotubes, essentially all of the neurons survived for at least 3 weeks. Many of the neurons formed functional synapses on myotubes under these conditions; some neuromuscular synapses could be detected as early as 20 hr after addition of the ganglion cells to muscle cultures. In contrast, most neurons from 8-day embryos survived for only a few days when grown alone on either polyornithine- or collagen-coated dishes. These results suggest that neurons destined to die in ovo can be rescued when grown in cell culture with myotubes and that under these conditions the neurons develop and express differentiated properties.
Behavior-genetic analysis of Phormia regina: conditioning, reliable individual differences, and selection.
Authors: McGuire T R , Hirsch J
Published: May 2006

Using proboscis extension (unconditioned response) to sucrose (unconditioned stimulus), individual blowflies (Phormia regina) were classically conditioned to saline and to water (conditioned stimuli) with sensitization controls, thus providing unique, independently replicated evidence both of learning in Diptera and of reliably measured individual differences. Directional and stabilizing selection have bred high and low performance lines markedly different from an unselected control line as a step in the analysis of behavior-genetic correlates. This replicates and extends previous selection analysis with improved conditioning technique. Also, some unwarranted claims of learning in Diptera are discussed.
Time-resolved resonance Raman spectroscopy of bacteriorhodopsin on the millisecond timescale.
Authors: Terner J , Campion A , El-Sayed M A
Published: May 2006

A simple technique is described that uses a continuous wave laser with electromechanical modulation to obtain time-resolved Raman spectra of transient species on the millisecond timescale. The time behavior of the vibrational bands of the intermediates involved in the proton pumping of bacteriorhodopsin is determined. From these results, along with resonance enhancement and power dependence studies, the bands that appear in the continuous wave Raman spectrum of bacteriorhodopsin can be assigned to three intermediates in the photochemical cycle of bacteriorhodopsin, bR570, bL550, and bM412. The Raman spectra of bR570 and bM412 are compared with published spectra of model Schiff bases of all-trans and 13-cis retinal.
Hydrophobicity, hydrophilicity, and the radial and orientational distributions of residues in native proteins.
Authors: Rackovsky S , Scheraga H A
Published: May 2006

The radial distributions of the Calpha and side-chain atoms in a sample of 13 native proteins have been examined. It is found that there are substantial differences in behavior between different atoms of the same amino acid. In particular, the Calpha atoms of polar residues show no particular preference for being far from the center of mass. In light of these results, a new criterion for hydrophobicity and hydrophilicity is proposed--namely, the orientational preference of the side chain. The distribution of this property is shown, and it is suggested that this provides a basis for incorporating hydrophobic interactions into a protein folding algorithm.
Activation of factor IX by the reaction product of tissue factor and factor VII: additional pathway for initiating blood coagulation.
Authors: Osterud B , Rapaport S I
Published: May 2006

A study was carried out on mechanisms, independent of activated Factor XI, capable of activating Factor IX. The reaction product of tissue factor and Factor VII functioned as a potent Factor IX activator in the assay system used. Activated Factor IX itself activated Factor X; thrombin failed to activate Factor IX. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis confirmed that the reaction product of tissue factor and Factor VII activated Factor IX, with replacement of the band corresponding to native factor IX [molecular weight (Mr) 55,000] by bands corresponding to the heavy chain (Mr 27,000) and light chain (Mr 17,000) of activated Factor IX. When either Factor VII or calcium ions were left out of incubation mixtures, the band of native Factor IX persisted unchanged. Contact of blood with tissue factor represents a second mechanism, bypassing activated Factor XI, for the activation of Factor IX during hemostasis. It may help to explain the discrepancy between the mild bleeding of hereditary Factor XI deficiency and the severe bleeding of hereditary Factor IX deficiency.
Evidence for control of serotonin secretion from human platelets by hydroxyl ion transport and osmotic lysis.
Authors: Pollard H B , Tack-Goldman K , Pazoles C J , Creutz C E , Shulman N R
Published: May 2006

Serotonin secretion from human platelets, stimulated either by thrombin or the calcium ionophore A23187, was found to be inhibited by anion transport blocking drugs such as 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS), pyridoxal phosphate, probenecid, and suramin. These drugs have previously been shown to inhibit ATP-evoked release of epinephrine from isolated chromaffin granules by blocking chloride uptake and subsequent osmotic lysis. However, in contrast to granule release, platelet secretion was insensitive to chloride and, instead, was dependent on OH-. Platelet release was suppressed by low pH, and inhibition by the transport blocking drugs was competitive only with respect to OH-. Serotonin release from platelets was also suppressed by increasing extracellular osmotic strength, and the relationship between suppression and external osmotic strength was quantitatively similar to that observed in the case of chromaffin granules. We conclude that platelet exocytosis could occur when serotonergic granules are closely juxtaposed to the plasma membrane, thus exposing the granule anion transport site to the more alkaline medium. Secretion of serotonin could occur as a consequence of OH- transport and osmotic lysis of the granule-plasma membrane complex, analogous to the chemiosmotic mechanism of chloride-dependent epinephrine release from isolated chromaffin granules.
Phage T4-modified RNA polymerase transcribes T4 late genes in vitro.
Authors: Rabussay D , Geiduschek E P
Published: May 2006

Initiation of T4 late RNA synthesis has been achieved in an in vitro system prepared from Escherichia coli cells infected with wild-type or maturation-defective mutant T4 phage. The system uses a cellophane membrane as a mechanical support for concentrated cell lysates and for added streptolydigin-resistant RNA polymerases. Transcriptional activity and selectivity of added RNA polymerases are tested while endogenous RNA polymerase activity is inhibited by streptolydigin. T4-modified RNA polymerase is required for substantial stimulation of T4 late RNA synthesis.