bluuebunny/crossref_metadata_2025 huggingface dataset:
Structure and function of carboxypeptidase A alpha in supercooled water.
Authors: Thompson J S , Gehring H , Vallee B L
Published: May 2006

The spectral and enzymatic characteristics of chromophoric derivatives of carboxypeptidase A alpha (EC 3.4.17.1) have been examined at subzero temperatures in supercooled water-in-oil emulsions. Substrate and temperature dependencies of enzyme kinetics indicated the existence of a solution-like enzyme phase that greatly extends the temperature range (greater than 60 degrees C) over which the activity of this enzyme can be measured. The emulsion spectra were virtually identical to those of solutions over a wide range of temperatures. Subzero temperatures (less than -10 degrees C) may induce changes of enzyme conformation but not of geometry at the site of the metal atom, nor do they adversely affect activity at any of the temperatures studied. Both structure and function of carboxypeptidase A alpha can be examined in supercooled water under identical reaction conditions.
Examination of the conventional assumption of internal equilibrium within the states of a biochemical cycle.
Authors: Hill T L
Published: May 2006

If k is a representative "internal" rate constant between substates of a given biochemical state, and if alpha is representative of the rate constants of the biochemical cycle to which the state belongs, then cyclical activity at steady state pulls the substates out of internal equilibrium with each other by a factor of order 1 +/- O(alpha/k). For transients or steady isotonic contractions in muscle, the departures from internal equilibrium can be larger than this. The simplifying assumption that internal equilibrium is always maintained between the substates is justified at steady state, as a good approximation, if k/alpha greater than or equal to 100. In muscle contraction at maximum velocity, something like k/alpha greater than or equal to 500 is required. This problem is superficially similar to the question, in Eyring's rate theory, of the extent to which activated complex leads to products pulls the activated complex out of the assumed equilibrium with reactants.
I and i antigens of human peripheral blood lymphocytes cocap with receptors for concanavalin A.
Authors: Feizi T , Kapadia A , Yount W J
Published: May 2006

Surface immunofluorescence experiments using a human anti-i and two anti-I antisera have been performed on human peripheral blood lymphocytes. These are known to contain cold-reactive monoclonal IgM antibodies against the carbohydrate sequence: (formula: see text). A high proportion of B- and T-type lymphocytes express these I and i determinants. In the presence of anti-human immunoglobulin, the cold-reactive membrane-associated complexes of I-anti-I and i-anti-i become stabilized, and redistribution (with patching and capping) can be elicited at 37 degrees C. Dual fluorescence experiments have shown striking concordant staining of I or i (fluorescein) caps and patches with concanavalin A (rhodamine) reactive sites on normal and leukemic cells, suggesting that a proportion of I and i active structures of lymphocyte membranes are structurally associated or physiologically coupled with glycoproteins carrying oligosaccharides with branched mannosyl cores.
Identification of an adhesion factor for chondrocytes.
Authors: Hewitt A T , Kleinman H K , Pennypacker J P , Martin G R
Published: May 2006

The attachment of chondrocytes to collagen substrates is stimulated by serum but not by fibronectin. The active material in serum was partially purified and was shown to be a protein by its sensitivity to trypsin and heat and its chromatographic properties. This factor, which we have named chondronectin, is distinct from fibronectin and does not stimulate fibroblast attachment. Because material with similar attachment-enhancing activity is produced by chondrocytes and is extractable from cartilage, chondronectin may be a chondrocyte-specific attachment protein.
Fusion of liposomes with mitochondrial inner membranes
Authors: Schneider Heinz , Lemasters John J. , Höchli Matthias , Hackenbrock Charles R.
Published: May 2006

A procedure is outlined for the fusion of mixed phospholipid liposomes (small unilamellar vesicles) with the mitochondrial inner membrane, which enriches the membrane lipid bilayer 30-700% in a controlled fashion. Fusion was initiated by manipulation of the pH of a mixture of freshly sonicated liposomes and the functional inner membrane/matrix fraction of rat liver mitochondria. During the pH fusion procedure, liposomes became closely apposed with and sequestered by the inner membranes as revealed by freeze-fracture electron microscopy. After the pH fusion procedure, a number of ultrastructural, compositional, and functional characteristics were found to be proportionally related: the membrane surface area increased; the lateral density distribution of intramembrane particles (integral proteins) in the plane of the membrane decreased whereas the particles remained random; the membrane became more buoyant; the ratio of membrane lipid phosphorus to total membrane protein increased; the ratio of membrane lipid phosphorus to heme
a
of cytochrome
c
oxidase increased; and the rate of electron transfer between some interacting membrane oxidoreduction proteins decreased. These data reveal that liposomal phospholipid was incorporated into the membrane bilayer (not simply adsorbed to the membrane surface) and that integral membrane proteins diffused freely into the laterally expanding bilayer. Furthermore, the data suggest that the rate of electron transfer may be limited by the rate of lateral diffusion of oxidoreduction components in the bilayer of the mitochondrial inner membrane.
Intervening sequences divide the gene for the constant region of mouse immunoglobulin mu chains into segments, each encoding a domain.
Authors: Gough N M , Kemp D J , Tyler B M , Adams J M , Cory S
Published: May 2006

To elucidate the structure of the gene for the constant region of immunoglobulin mu chains, we have cloned a 9.9-kilobase-pair fragment of mouse DNA bearing a gene for the constant region of the mu chain (C mu gene) from an IgM-secreting mouse plasmacytoma. The sequence around this gene has apparently undergone somatic rearrangement; the gene occurs in an EcoRI restriction endonuclease fragment of a different size from that in embryo or liver DNA and no C mu-bearing fragment of embryo size remains in the plasmacytoma. The cloned sequence lacks a variable region gene; hence, if this C mu gene is active, its position within the clone indicates that the gene for the variable region of a heavy chain (VH gene) must be more than 3.7 kilobase pairs away. The C mu gene is divided by three intervening sequences into four coding segments, each of which encodes one of the domains (homology units) of the polypeptide. The nucleotide sequence coding for amino acids near the V-C junction is not present within the C mu clone or clones bearing homologous embryonic VH genes. This suggests that an immunoglobulin heavy chain, in common with light chains, is encoded not only by a V and C gene, but also by an independent joining region (JH) gene.
Isozymes of human phosphofructokinase: Identification and subunit structural characterization of a new system
Authors: Vora Shobhana , Seaman Carol , Durham Susan , Piomelli Sergio
Published: May 2006

The existence of a five-membered isozyme system for human phosphofructokinase (PFK; ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11) has been demonstrated. These multimolecular forms result from the random polymerization of two distinct subunits, M (muscle type) and L (liver type), to form all possible tetrameters—i.e., M
4
, M
3
L, M
2
L
2
, ML
3
, and L
4
. Partially purified muscle and liver PFKs were hybridized by dissociation at low pH and then recombination at neutrality. Three hybrid species were generated in addition to the two parental isozymes, to yield an entire five-membered set. The various species could be consistently and reproducibly separated from one another by DEAE-Sephadex chromatography at pH 8.0 with a concave elution gradient of salt. Under similar experimental conditions, erythrocyte PFK from hemolysates was also resolved into five species chromatographically indistinguishable from those produced in the above experiment. Immunological and kinetic studies of the isozymes provided corroborative evidence to support the proposed subunit structures. Erythrocyte PFK was found to have kinetic properties intermediate between those of muscle and liver PFK and was neutralized only 50% by an antiserum against muscle PFK that completely neutralized muscle PFK. These data demonstrate that muscle and liver PFKs are distinct homotetramers—i.e., M
4
and L
4
, respectively—whereas erythrocyte PFK is a heterogeneous mixture of all five isozymes. The structural heterogeneity of erythrocyte PFK provides a molecular genetic basis for the differential organ involvement observed in some inherited PFK deficiency states in which myopathy or hemolysis or both can occur.
Early nerve-muscle synapses in vitro release transmitter over postsynaptic membrane having low acetylcholine sensitivity.
Authors: Cohen S A
Published: May 2006

Functional nerve-muscle synapses form rapidly in cultures of embryonic chicken spinal cord and muscle cells. As early as 30 min after nerve processes first contact muscle fibers they are able to release stimulus-evoked neurotransmitter. This release was detected only after wave-form averaging because of the exceedingly low amplitude of the synaptic potentials. This small size was likely due to a postsynaptic effect, because the electrophysiologically assayed acetylcholine sensitivity of the synaptic muscle membrane was low and did not differ significantly from extrasynaptic levels. Transmitter release was elicited both from along the lengths of nerve processes and from active growth cones.
Nerve growth factor stimulates the development of substance P in sensory ganglia.
Authors: Kessler J A , Black I B
Published: May 2006

The development of the putative neurotransmitter substance P (SP) in rat dorsal root ganglion (DRG) was defined in vivo. The sixth cervical DRG of newborn rats contained 70 pg of SP, and the ganglionic content increased 5.5-fold during the first 5 weeks of life. Forelimb amputation partially prevented the normal developmental increase of SP in the sixth cervical DRG destined to innervate that limb. Conversely, treatment with nerve growth factor (NGF) increased both ganglionic SP and total ganglion protein. Moreover, NGF administration prevented the failure of SP development that followed amputation, suggesting that NGF may mediate the limb-DRG interaction. However, treatment with antiserum to NGF failed to significantly inhibit development of ganglion SP. Consequently, neonatal ganglia may remain responsive to NGF, without requiring the protein for survival. SP appears to be an excellent index of the maturation of neurons in dorsal root ganglia.
A theory of distortion of the reaction zone
Authors: Coon Nancy , Ma Shao-Mu , Redington Patrick K. , Brown Billings , Eyring Henry
Published: May 2006

A cylindrical detonating explosive behaves like a non-Newtonian viscous fluid emerging from a cylindrical pipe. The expression for non-Newtonian viscous flow has been applied to detonating explosives. The resultant fit to emerging flame fronts from detonation is excellent for a wide variety of detonating explosives both for transients (at various times) and for steady state. The quantities
k
′(λ/λ
1
)(2/
B
) and 1/
B
and the rate constant
k
′ increase abruptly at a time supporting a change in mechanism from deflagration to detonation in the transient during initiation. All of these parameters are temperature sensitive, indicating an abrupt temperature increase as a condition or indicator in the deflagration-to-detonation transition. Furthermore, a straight line results when log particle velocity
u
is plotted against log radial distance
r
, indicating that, for the explosives studied, the hyperbolic cosine wave front can be replaced by a parabolic wave front within the accuracy of the available experimental data.
Influence of intramolecular hydrogen bonding on the electronic structure of oxymorphone
Authors: Scheiner Steve , Kolb Vera M.
Published: May 2006

Approximate
ab initio
molecular orbital methods are used to examine the structural and electronic properties of oxymorphone. The most stable conformation of the molecule is found to include an intramolecular hydrogen bond between the C-14 hydroxyl group and the nitrogen atom in agreement with available experimental data. The total molecular electron density is transformed to a set of localized molecular orbitals, one of which corresponds to the lone electron pair on nitrogen. The hydrogen bond is shown to produce substantial bending and stretching of the lone pair when compared to its shape when such hydrogen bonding is precluded.
Coding of correlated sources with prescribed distortion by separated encoders
Authors: Wolfowitz J.
Published: May 2006

Let (

X
n
,Y
n

) = (
X
1
,
Y
1
),...,(

X
n
,Y
n

) be n independent, identically distributed pairs of chance variables. Each of two separated encoders encodes a different sequence of (

X
n
,Y
n

) to within given respective distortions
z
1
,
z
2
. Call a region in the plane of the respective rates (
R
1
,
R
2
)
M
-convex if it is convex, closed, and, to every point (
r
1
,
r
2
), contains all points such that each of their coordinates is larger than or equal to the corresponding coordinate of (
r
1
,
r
2
). This paper determines the (
M
-convex) region of achievable rates by giving its boundary. For
z
1
=
z
2
= 0, the region specializes to that of Slepian and Wolf. For
z
2
= ∞ and
R
2
=

H(Y
i
)

, the minimum
R
1
is that of Wyner and Ziv; for
z
2
= ∞ and general given
R
2
, the minimum
R
1
is that of my generalization of the theorem of Wyner and Ziv.
Artin relation for smooth representations
Authors: Dovermann Karl Heinz , Petrie Ted
Published: May 2006

Let
G
be a finite group. If
G
acts smoothly on a closed homotopy sphere
S
, we call
S
a smooth representation of
G
. The main result is: There is a function

h
G

such that for every smooth representation
S
of
G
, dimension

S
G

=

h
G

{dimension

S
H

ǀ
H
proper subgroup of
G
} if and only if
G
has prime power order and
G
is not cyclic. In other words, only for a noncyclic
p
-group
G
is dimension

S
G

a universal function of the dimensions of the fixed sets

S
H

as
H
ranges over proper subgroups of
G
. This result is compared with an old theorem of Artin's dealing with dimensions of fixed sets of orthogonal representations of
G
.
Recognition of local nucleotide conformation in contrast to sequence by a rRNA processing endonuclease.
Authors: Stahl D A , Meyhack B , Pace N R
Published: May 2006

RNase M5 of Bacillus subtilis cleaves twice in a double-helical region of a 179-nucleotide precursor of 5S rRNA to yield mature 5S rRNA (116 nucleotides) plus fragments (21 and 42 nucleotides) derived from both termini. Previous experiments had shown that the major recognition elements for the highly specific RNase M5 are in the mature domain of the precursor. However, one precursor residue, a G adjacent to the 5' cleavage site, significantly enhances the rate of its own cleavage as well as that of the 3' precursor fragment, so it must be an important component of the features recognized by the enzyme. This G residue is opposed in the helical substrate region to a C residue, which is at the 3' terminus of the mature domain, presenting the question of whether RNase M5 specifically contacts the cleavage site on the basis of nucleotide sequence (the G residue per se) or on the basis of more general aspects of helical conformation. We tested these alternatives by fabricating partially synthetic test substrates for RNase M5. Experiments were performed on 5' and 3' half-molecules derived from mature 5S rRNA. The 3'-terminal C was removed by periodate oxidation and beta elimination and replaced in a T4 RNA ligase condensation with each of the four mononucleoside bisphosphates. Artificial "precursor" segments containing each of the four nucleotides adjacent to the 5' cleavage site were added to the 5' terminus of the 5S rRNA half-molecule. We then annealed the modified half-molecules to yield test substrates containing all permutations of complementary in contrast to noncomplementary nucleotides at the cleavage site. The susceptibilities of these test substrates show that conformation, not sequence, is the important feature in the locale of the cleaved bonds.
Parathyroid hormone alters collagen synthesis and procollagen mRNA levels in fetal rat calvaria.
Authors: Kream B E , Rowe D W , Gworek S C , Raisz L G
Published: May 2006

Parathyroid hormone decreased the incorporation of [3H]proline into collagenase-digestible protein in cultured 21-day fetal rat calvaria but had little effect on the labeling of noncollagen protein. After 24 hr of culture, there was a 50% reduction in collaghen synthesis and a 40% decrease in the level of functional procollagen mRNA as measured in a reticulocyte lystate translation assay. The effect of parathyroid hormone on both parameters was detectable after 6 hr of treatment. In these cultures, there was also a substantial degradation or release of newly synthesized collagen from the calvaria, but parathyroid hormone had little effect on the release of collagen into the medium. These results suggest that parathyroid hormone inhibits collagen synthesis primarily by decreasing the steady-state level of procollagen mRNA.
Organization of actin gene sequences in the sea urchin: molecular cloning of an intron-containing DNA sequence coding for a cytoplasmic actin.
Authors: Durica D S , Schloss J A , Crain W R
Published: May 2006

Southern transfer and solution hybridization experiments, using as probe a DNA fragment that encodes for Drosophila actin, demonstrate cross hybridization to DNA from the sea urchin Strongylocentrotus purpuratus. Recombinant DNA clones that contained sea urchin genomic DNA fragments were constructed and screened for the presence of actin-encoding DNA sequences by colony hybridization with the Drosophila actin sequence. Two different putative actin-encoding clones were identified and were shown to specifically hybridize actin-encoding mRNA from a complex mRNA population. Southern blot hybridization experiments with both the Drosophila actin sequence and one of the cloned sea urchin sequences, in conjunction with solution hybridization data, suggest an actin gene copy number of 5-20 per haploid genome. DNA sequence analysis of one of the cloned sequences indicates that this fragment codes for a cytoplasmic form of actin and contains an intervening sequence of at least 200 nucleotides beginning immediately after amino acid 121 in the protein sequence.
Identification of the slow reacting substances from cat paws.
Authors: Houglum J , Pai J K , Atrache V , Sok D E , Sih C J
Published: May 2006

Perfusion of cat paws with compound 48/80 released two slow reacting substances (SRSs) which were isolated and characterized as 5-hydroxy-6-S-cysteinylglycyl-7,9,11,14-icosatetraenoic acid (SRS I) and 5-hydroxy-6-S-cysteinyl-7,9,11,14-icosatetraeonic acid (SRS II) on the basis of chemical degradations, amino acid analyses, spectroscopic and enzymic experiments, and comparison with synthetic samples. The smooth muscle-contractile activities of synthetic 5-hydroxy-6-gamma-glutamylcysteinylglycyl-7,9,11,14-icosatetraenoic acid, synthetic 5-hydroxy-6-S-cysteinyl-7,9,11,14-icosatetraenoic acid, and SRS II were not inactivated by arylsulfatase. On the other hand, the spasmogenic activities produced by synthetic 5-hydroxy-6-S-cysteinylglycyl-7,9,11,14-icosatetraenoic acid and SRS I were destroyed at the same rate by the arylsulfatase. This mode of inactivation was attributed to an aminopeptidase activity in the arylsulfatase preparation because 5-hydroxyl-6-S-cysteinyl-7,9,11,14-icosatetraenoic acid was isolated and identified as the reaction end product. Because the properties of SRS from cat paws closely resemble those of SRS generated by immunological stimulation of human tissues (SRS-A) and because all known SRS-A are inactivated by arylsulfatases, we contend that 5-hydroxy-6-S-cysteinylglycyl-7,9,11-14-icosatetraenoic acid (SRS I) corresponds to SRS-A.
Elongation of primed DNA templates by eukaryotic DNA polymerases.
Authors: Ikeda J E , Longiaru M , Horwitz M S , Hurwitz J
Published: May 2006

The combined action of DNA polymerase alpha and DNA polymerase beta leads to the synthesis of full-length linear DNA strands with phi X174 DNA templates containing an RNA primer. The reaction can be carried out in two stages. In the first stage, DNA polymerase alpha catalyzes the synthesis of a chain that averaged 230 deoxynucleotides long and was covalently linked to the RNA primer. In the second stage, DNA polymerase beta elongates the DNA strand covalently attached to the RNA primer to full length. With DNA primers, DNA polymerase alpha catalyzes only limited deoxynucleotide addition whereas DNA polymerase beta alone elongates DNA primed templates to full length. DNA polymerase beta can also stimulate the synthesis of adenovirus DNA in vitro in the presence of a cytosol extract from adenovirus-infected cells. In all of these systems, dNMP incorporation catalyzed by DNA polymerase beta was sensitive to N-ethylmaleimide; however, this polymerase activity was resistant to N-ethylmaleimide with poly(rA) x (dT) as the primer template.
White lines in L-edge x-ray absorption spectra and their implications for anomalous diffraction studies of biological materials.
Authors: Lye R C , Phillips J C , Kaplan D , Doniach S , Hodgson K O
Published: May 2006

We have measured high-resolution x-ray absorption spectra of lanthanide (Ln) and heavy transition metal complexes that display prominent narrow absorption peaks near the L2 and L3 absorption edges. The anomalous scattering factors (f' and f"), which are mathematically related to the absorption cross section, have correspondingly sharp changes in their magnitude within 5-10 eV of the absorption edge. Calculations of the magnitude of the change in f' and f" demonstrate that significant changes (on the order of 20 electrons in f') can be expected for these materials. These substantial changes in the anomalous scattering factors have applications to deriving structural information for macromolecules from x-ray diffraction studies. The magnitude of the changes indicate that the anomalous scattering technique is a powerful means of obtaining structural characteristics for macromolecules in single crystals, in solution, and in biological membranes.
Spontaneous production of human interferon.
Authors: Pickering L A , Kronenberg L H , Stewart W E
Published: May 2006

Several established lines of human lymphoblastoid cells were evaluated for abilities to produce interferons. Some cell lines were able to produce interferon when induced with either Newcastle disease virus or Sendai virus, whereas others failed to produce detectable interferon when so induced. However, several cell lines were able to spontaneously produce interferon without induction. Spontaneously produced interferon was liberated by cells only during logarithmic growth phase, reaching levels ranging from about 10 reference units/ml of growth medium for some cell lines to 1000 reference units/ml for others. The interferons produced by induced lymphoblastoid cells and the spontaneously produced interferons were all characterized as type I human leukocyte interferon by high levels of cross-species antiviral activities on bovine cells and by neutralizations by antiserum to human leukocyte interferon but not by antiserum to human fibroblast interferon. However, analysis by electrophoresis in sodium dodecyl sulfate/polyacrylamide gels revealed that spontaneously produced interferon was less size heterogeneous than human leukocyte interferon, migrating as a single band of activity with a peak at 20,000 daltons, whereas human leukocyte interferon contained peaks of major activity at 23,000 and 18,000 daltons and virus-induced Namalva lymphoblastoid cell interferon migrated predominantly as the 18,000-dalton form. Also, although neither virus-induced primary leukocyte interferon nor any of the virus-induced lymphoblastoid cell interferons were neutralized by antiserum to mouse interferon, all of the spontaneously produced interferons were neutralized by antiserum to mouse interferons. These data suggest significant structural similarities between the active cores of certain interferons from phylogenetically diverse animal species.
Human leukocyte interferon: structural and biological relatedness to adrenocorticotropic hormone and endorphins.
Authors: Blalock J E , Smith E M
Published: May 2006

Anti-alpha-corticotropin [anti-ACTH alpha (1-13)](also alpha-melanotropin) and anti-gamma-endorphin antisera neutralized human leukocyte interferon activity but not fibroblast interferon activity. Human leukocyte interferon was not neutralized by anti-human lutenizing hormone (lutropin) or follicle-stimulating hormone (follitropin) antisra. Conversely, antisera to human leukocyte interferon neutralized ACTH activity. The neturalization of human leukocyte interferon by anti-human leukocyte interferon serum was partially blocked by ACTH. These studies show strong antigenic relatedness among human leukocyte interferon, ACTH, and endorphins, implying that there are underlying structural similarities. Structural relatedness is shown by pepsin cleavage of ACTH activity from human leukocyte interferon. The implication for the natural functions of human leukocyte interferon are discussed.
Absolute requirement of glucocorticoid for expression of the casein gene in the presence of prolactin.
Authors: Ganguly R , Ganguly N , Mehta N M , Banerjee M R
Published: May 2006

Second thoracic mammary glands of immature BALB/c female mice were stimulated to pregnancy-like lobuloalveolar (LA) development aftr 6 days of incubation in a corticosteroid-free step I culture medium containing insulin, prolactin, estradiol, progesterone, and growth hormone. A low basal level (0.0009%) of casein mRNA (mRNAcsn) sequences was detectable in the LA glands by a specific cDNA probe. Subsequent incubation of the LA glands for 3 days in medium containing insulin and prolactin or insulin and cortisol failed to elicit mRNAcsn above the basal level, indicating that neither prolactin nor cortisol alone can support casein gene expression. However, an increase in mRNAcsn levels was observed when the 3-day incubation with insulin and cortisol or insulin and prolactin was followed by 3 days of culture in presence of insulin, prolactin, and cortisol. When a 3-day incubation with insulin and prolactin was followed by 3 days in insulin and cortisol medium, mRNAcsn levels in the gland remained similar to the basal level. However, a 20-fold increase in the mRNAcsn levels ensued when the LA glands were sequentially incubated for 3 days in insulin and cortisol and then for another 3 days in insulin and prolactin medium. After a preincubation in insulin and cortisol medium, the LA glands retained residual cortisol during subsequent incubation in insulin and prolactin medium, and the mRNAcsn levels in these glands were related to the level of residual cortisol present. When mRNAcsn and the residual cortisol level reached a minimum, addition of fresh cortisol to the medium caused a 20-fold increase in the mRNAcsn levels. This indicates that cortisol is a limiting factor in insulin and prolactin medium and its presence is absolutely required for casein gene expression.
Induction of mitotic recombination in yeast by starvation for thymine nucleotides.
Authors: Kunz B A , Barclay B J , Game J C , Little J G , Haynes R H
Published: May 2006

The biosynthesis of thymine nucleotides in Saccharomyces cerevisiae can be inhibited either by genetic lesions in the structural gene for thymidylate synthetase (TMP1) or by drugs that prevent the methylation of dUMP to dTMP. This methylation can be blocked by folate antagonists. We find that 5-fluoro-dUMP (FdUMP) is also an effective inhibitor in vivo. Inhibition of dTMP biosynthesis by these three different routes causes thymineless death. In addition to being cytotoxic, we find that FdUMP is highly recombinagenic in yeast but does not induce nuclear gene mutations. Provision of exogenous dTMP eliminates this induced mitotic recombination and cell killing. Similar results were obtained when a thymineless condition was provoked in cells by antifolate drugs or by dTMP deprivation in strains auxotrophic for this nucleotide. These findings show that, in contrast to the situation in prokaryotes, starvation for thymine nucleotides in yeast induces genetic recombination but is not mutagenic.
Constitutive uncoupling of pathways of gene expression that control growth and differentiation in myeloid leukemia: a model for the origin and progression of malignancy.
Authors: Sachs L
Published: May 2006

Chemical carcinogens and tumor promoters have pleiotropic effects. Tumor initiators can produce a variety of mutations and tumor promotres can regulate a variety of physiological molecles that control growth and differentiation. The appropriate mutation and the regulation of the appropriate molecules to induce cell growth can initiate and promote the sequence of changes required for transformation of normal cells into malignant cells. After this sequence of changes, some tumors can still be induced to revert with a high frequency from a malignant phenotype to a nonmalignant phenotype. Results obtained from analysis of regulation of growth and differentiation in normal and leukemic myeloid cells, the phenotypic reversion of malignancy by induction of normal differentiation in myeloid leukemia, and the blocks in differentiation-defective leukemic cell mutants have been used to propose a general model for the origin and progression of malignancy. The model states that malignancy originates by changing specific pathways of gene expresion required for growth from inducible to constitutive in cells that can still be induced to differentiate normally by the physiological inducer of differentiation. The malignant cells, unlike the normal cells, then no longer require the physiological inducer for growth. This changes the requirements for growth and uncouples growth from differentiation. Constitutive expression of other specific pathways can uncouple other controls, which then causes blocks in differentiation and the further progression of malignancy. The existence of specific constitutive pathways of gene expression that uncouple controls in malignant cells can also exlain the expresion of fetal proteins, hormones, and some other specialized products of normal development in various types of tumors.
Wavelength regulation in rhodopsin: effects of dipoles and amino acid side chains.
Authors: Hays T R , Lin S H , Eyring H
Published: May 2006

The effects of dipoles and aromatic amino acid side-chain models on the absorption and optical activity of the rhodopsin chromophore were calculated by using perturbation theory, and the results were compared with those of a Pariser-Parr-Pople calculation for the unperturbed system. The interaction was assumed to result from purely electrostatic interactions. It was concluded that the side chains of phenylalanine and tryptophan should have no important effects. However, the charge separation in tyrosine is sufficient to cause substantial electrostatic perturbation; in fact, the effect of tyrosine is large enough to approximately many of the spectral properties of rhodopsin quantitatively. This is encouraging because the use of aromatic amino acid side-chain analogs probably provides a better physical model than the use of isolated full charges, except in the case of the counterion to the protonated Schiff base.
Autonomously replicating sequences in Saccharomyces cerevisiae.
Authors: Chan C S , Tye B K
Published: May 2006

A method is presented for isolating DNA segments capable of autonomous replication from Saccharomyces cerevisiae chromosomal DNA based on the differential transforming ability of autonomously replicating plasmids and nonreplicating plasmids. DNA plasmids that are capable of self-replication in yeast transform yeast spheroplasts at about 1000-fold higher frequency than nonreplicating plasmids. We have cloned from total yeast DNA a number of DNA segments that permit the pBR322 plasmid carrying the yeast LEU2 gene to replicate autonomously. These plasmid clones are characterized by their ability to transform Leu- spheroplasts to Leu+ at a high frequency and their ability to replicate autonomously. Analysis of the insert DNAs carried in some of these self-replicating plasmids divides them into two categories: those that are unique in the yeast genome and those that are repetitive.
Cytoskeletal constraint of the beta-adrenergic receptor in frog erythrocyte membranes.
Authors: Cherksey B D , Zadunaisky J A , Murphy R B
Published: May 2006

A fluorescence receptor binding assay, based upon the high-affinity beta-adrenergic receptor antagonist propranolol, is utilized to probe the microenvironment of the antagonist-receptor complex in the frog (Rana catesbeiana) erythrocyte membrane. The technique of steady-state fluorescence depolarization is applied to the propranolol-receptor complex, allowing quantitation of the rotational relaxation time of the complex. It is found that the complex is dynamically constrained at 20 degrees C. However, in the temperature range 6-10 degrees C a sharp reversible release of constraint is observed. It is further demonstrated that the addition of drugs that are known to specifically disrupt the cytoskeleton (colchicine, vincristine, and vinblastine) causes a similar but irreversible release of constraint at 20 degrees C. Cytochalasin B has a much smaller influence on the rotational mobility of the propranolol-receptor complex than do the other drugs that disrupt the cytoskeleton. Amphotericin B is without effect on the rotational constraint of the complex. Binding of the antagonist [3H]dihydroalprenolol is not influenced by colchicine. A model is proposed which postulates that cytoskeletal elements are linked to the antagonist-receptor complex. Antagonist binding does not result in cytoskeletal release, whereas agonist binding is postulated to lead to dissociation of the agonist-receptor complex from the cytoskeleton, thereby activating adenylate cyclase.
Protein p3 is linked to the DNA of phage phi 29 through a phosphoester bond between serine and 5'-dAMP.
Authors: Hermoso J M , Salas M
Published: May 2006

To investigate the role of protein p3 in bacteriophage phi 29 initiation of replication, we have studied the nature of the covalent linkage between protein p3 and phi 29 DNA. The protein-DNA compound was digested with micrococcal nuclease and pronase resulting in a nucleotidyl-peptide that was further digested by alkaline phosphatase and snake venom phosphodiesterase yielding 5'-dAMP. The DNA-protein linkage is sensitive to alkali. Treatment of the nucleotidyl-peptide with 0.1 M NaOH at 37 degrees C for 3 hr after phosphatase digestion released 5'-dAMP. Hydrolysis of the nucleotidyl-peptide with 5.8 M HCl at 110 degrees C for 90 min yielded O-phosphoserine. These results, together with the sensitivity of the DNA-protein linkage to snake venom phosphodiesterase and its resistance to hydroxylamine, indicate that protein p3 is covalently linked to phi 29 DNA through a phosphoester bond between L-serine and 5'-dAMP, namely a O,5'-deoxyadenylyl-L-serine bond.
Specific photoaffinity labeling induced by energy transfer: application to irreversible inhibition of acetylcholinesterase.
Authors: Goeldner M P , Hirth C G
Published: May 2006

p-Dimethylaminobenzene diazonium fluoroborate belongs to a class of potential photoaffinity labeling reagents which, by irradiation, produces a highly reactive electrophilic species. In addition, it can be photodecomposed by photoexcited tryptophan derivatives (e.g., N-acetyltryptophanamide and tryptophan residues belonging to acetylcholinesterase) by an energy transfer reaction. This substance is a competitive inhibitor of acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7) and is able to inactivate the enzyme either by photoaffinity labeling after irradiation at 410 nm or by an energy transfer reaction after irradiation at 295 nm. The efficiency of this method is demonstrated by an increase of the rate of enzyme inactivation as well as by a decrease of nonselective labeling with a radioactive inhibitor p-[methyl-3H]-dimethylaminobenzene diazonium fluoroborate.
Recombination between higher plant DNA and the Ti plasmid of
Agrobacterium tumefaciens
Authors: Thomashow M. F. , Nutter R. , Postle K. , Chilton M.-D. , Blattner F. R. , Powell A. , Gordon M. P. , Nester E. W.
Published: May 2006

The Ti plasmid sequences (T-DNA) from the octopine-producing crown gall tumor A6S/2 were isolated by molecular cloning, using the bacteriophage λ vector Charon 4A. Analysis of the cloned DNA segments indicates that the Ti plasmid sequences are covalently joined to plant nuclear DNA. These data demonstrate that genetic recombination between a eukaryote and a prokaryote can occur as a natural phenomenon.
The inverted repeat as a recognizable structural feature in supercoiled DNA molecules.
Authors: Lilley D M
Published: May 2006

The single-strand-specific endonuclease S1 from Aspergillus oryzae introduces highly selective cleavages into supercoiled covalently closed circular DNA molecules, but not into their previously linearized counterparts. The cleavage sites are inverted repeats of unit length between 9 and 13 base pairs, separated by a nonrepetitious 2-6 base pairs. Such regions may adopt hairpin or similar structures stabilized by the negative superhelix density and may constitute recognition sites for cellular proteins.
Nucleotide sequence of a cyanobacterial
nifH
gene coding for nitrogenase reductase
Authors: Mevarech Moshe , Rice Douglas , Haselkorn Robert
Published: May 2006

The nucleotide sequence of
nifH
, the structural gene for nitrogenase reductase (component II or Fe protein of nitrogenase) from the cyanobacterium
Anabaena
7120 has been determined. Also reported are 194 bases of the 5′-flanking sequence and 170 bases of the 3′-flanking sequence. The predicted amino acid sequence was compared with that determined for the complete nitrogenase reductase of
Clostridium pasteurianum
and the cysteine-containing peptides of the protein from
Azotobacter vinelandii
. Amino acid sequences around five cysteines, located in the NH
2
-terminal two-thirds of the protein, are highly conserved in all three species. Codon usage in the first gene from a cyanobacterium to be sequenced shows striking asymmetries for eight amino acids.
Purification of the regulatory component of adenylate cyclase.
Authors: Northup J K , Sternweis P C , Smigel M D , Schleifer L S , Ross E M , Gilman A G
Published: May 2006

The regulatory component (G/F) of adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] from rabbit liver plasma membranes has been purified essentially to homogeneity. The purification was accomplished by three chromatographic procedures in sodium cholate-containing solutions, followed by three steps in Lubrol-containing solutions. The specific activity of G/F was enriched 2000-fold from extracts of membranes to 3-4 mumol x min-1 x mg-1 (reconstituted adenylate cyclase activity). Purified G/F reconstitutes guanine nucleotide-, fluoride-, and hormone-stimulated adenylate cyclase activity in the adenylate cyclase-deficient variant of S49 murine lymphoma cells. G/F also recouples hormonal stimulation of the enzyme in the uncoupled variant of S49. Preparations of pure G/F contain three polypeptides with approximate molecular weights of 52,000, 45,000, and 35,000. The active G/F protein behaves as a multisubunit complex of these polypeptides. Treatment of G/F with [32P]NAD+ and cholera toxin covalently labels the molecular weight 52,000 and 45,000 polypeptides with 32P.
Isolation of yeast genes with mRNA levels controlled by phosphate concentration.
Authors: Kramer R A , Andersen N
Published: May 2006

A library of DNA from the yeast, Saccharomyces cerevisiae, was constructed in phage lambda Charon 4 vector and then screened by differential plaque filter hybridization for genes induced by phosphate starvation. Two EcoRI fragments of 7.9 and 5.0 kilobase pairs that contained such genes were isolated. These cloned fragments may each carry one of the several copies of the genes for the repressible acid phosphatase of yeast. The fragments were use to examine mRNA levels of these genes in regulatory mutants of acid phosphatase.
Altered distributions of the cytoskeletal proteins vinculin and alpha-actinin in cultured fibroblasts transformed by Rous sarcoma virus.
Authors: David-Pfeuty T , Singer S J
Published: May 2006

It was recently shown by combined immunofluorescence and interference reflection microscopy that a protein named vinculin, along with alpha-actinin, is concentrated at focal adhesion plaques inside cultured normal fibroblasts [Geiger, B. (1979) Cell 18, 193-205]. These plaques are the discrete, isolated sites of strong adhesions formed between the ventral surfaces of the cells and the substrata on which they are grown. We show that after transformation of fibroblasts by Rous sarcoma virus a majority of the cells have many fewer focal adhesion plaques and now exhibit a cluster of small patches that are immunolabelled for both vinculin and alpha-actinin. Such a cluster (rosette) is located near the ventral surface of the cell, usually partly under the nucleus. The significance that these altered distributions of vinculin and alpha-actinin may have for the rounding up and loss of adherence of transformed cells is discussed.
Sequence-specific recombination of plasmid ColE1.
Authors: Warren G J , Clark A J
Published: May 2006

The conjugal transmission of plasmid ColE1 and its derivatives can lead to recA-independent recombination. The recombination is not observed during vegetative growth and takes place specifically between the sequences that are thought to represent the transfer origin of ColE1. This provides genetic evidence for breakage and reunion events during the transmission of ColE1.
Nucleic acid spot hybridization: rapid quantitative screening of lymphoid cell lines for Epstein-Barr viral DNA.
Authors: Brandsma J , Miller G
Published: May 2006

A simple nucleic acid hybridization method to screen numerous samples of eukaryotic cells rapidly for their Epstein-Barr virus (EBV) DNA content is described. Whole cells are spotted on nitrocellulose filters and their DNA is denatured and fixed to the filter. The resultant DNA spots are hybridized to nick-translated EBV DNA and the extent of hybridization is monitored by autoradiography and scintillation counting. Statistical analysis of serial dilutions of cells permits their viral genome content to be estimated quantitatively by reference to a known standard, such as Raji cells or an artificial mixture of pure viral DNA and uninfected lymphocytes. The sensitivity of the method is between 5 and 50 pg of viral DNA. With this method we are able to select subclones that are high produces of EBV DNA and to identify the optimal time for harvest of EBV DNA from cultured cells. Spot hybridization should permit any cell population or fluid to be screened for the presence of a DNA sequence for which a radioisotopically labeled probe is available.
Distinct cholecystokinin receptors in brain and pancreas.
Authors: Innis R B , Snyder S H
Published: May 2006

125I-Labeled (Bolton-Hunter) cholecystokinin triacontatriapeptide (CCK-33) binds saturably and reversibly to distinct receptors in brain and pancreatic membranes. The peptide specificity of pancreatic CCK binding is the same as that for pancreatic amylase release. In brain, gastrin and pentagastrin display nanomolar affinity for binding sites, whereas in pancreas these two peptides are virtually inactive. Though these differences indicate that brain and pancreas possess distinct CCK receptors, the two tissues show some similarities. Both pancreas and brain receptors show greater sensitivity to sulfated than to desulfated COOH-terminal octapeptide of CCK and display dissociation constants of 0.3-9.5 nM. The pancreas possesses about 300 times more binding sites than does brain. CCK binding in both brain and pancreas is enhanced by divalent cations and reduced by monovalent cations. Receptor binding in both tissues is regulated in a selective fashion by guanyl nucleotides.
Organization of unc gene cluster of Escherichia coli coding for proton-translocating ATPase of oxidative phosphorylation.
Authors: Kanazawa H , Tamura F , Mabuchi K , Miki T , Futai M
Published: May 2006

The proton-translocating ATPase (F1-F0) of oxidative phosphorylation (ATP phosphohydrolase, EC 3.6.1.3) is coded for by a set of structural genes comprising the unc operon in Escherichia coli. We have analyzed several new transducing phages and plasmids carrying various lengths of the DNA segments of the unc operon by complementation assay using 14 new unc- mutants and representatives of previously described strains which were made available to us. Transducing phages carrying parts of the unc gene cluster were isolated: lambda uncA-9 and lambda glmS phages converted only some of the unc- mutants to the Unc+, as determined by complementation assays. A new hybrid plasmid (pMCR533) carrying part of the unc operon was constructed by inserting the HindIII fragment of lambda asn-5 DNA (a phage carrying the entire unc operon) into the unique HindIII site of pBR322. This plasmid transformed eight unc- strains to Unc+, including uncB402 and uncA401, but did not complement uncD11 or four other strains. Two minichromosomes which carry the E. coli replication origin were also tested: plasmid pNH05 transformed the uncB402 but not the uncA401 strain to Unc+, whereas plasmid pMCF1 transformed none of the mutants tested. Analysis of the DNAs from these transducing phages and plasmids with restriction endonucleases suggested that all of the structural genes for the F1-F0 complex are localized within a DNA segment of approximately 4.5 megadaltons containing two EcoRI sites. The approximate locations of the unc- mutations were mapped on this DNA segment.
Deoxycorticosterone biosynthesis in human kidney: potential for formation of a potent mineralocorticosteroid in its site of action.
Authors: Winkel C A , Simpson E R , Milewich L , MacDonald P C
Published: May 2006

The extra-adrenal formation of deoxycorticosterone (DOC) from plasma progesterone has been demonstrated in humans. In those studies it was shown that in some persons the volume of plasma cleared of progesterone by DOC formation was great, namely, 75 liter/24 hr. Because steroid 231-hydroxylase activity [steroid 21-monooxygenase; steroid, hydrogen-donor:oxygen oxidoreductase (21-hydroxylating), EC 1.14.99.10] could not be demonstrated in homogenates or microsome-enriched preparations of human lung or liver tissue, we speculated that the 21-hydroxylation of plasma progesterone might take place in the kidney. Employing whole tissue homogenates and microsome-enriched preparations of human kidney tissue, we demonstrated the formation of [3H]DOC from kidney tissue, we demonstrated the formation of [3H]DOC from [3H]progesterone. The rate of formation of DOC from progesterone in microsomal preparations from kidney tissues of adult humans varied from 0 to 803 pmol per mg of microsomal protein per hr. The value computed for the apparent Km of the enzyme for progesterone was 0.140 microM. On the basis of these findings, we conclude that steroid 21-hydroxylase activity is present in human kidney tissue and that the kidney may be an important site of DOC formation as well as a site of DOC action.