bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural gene identification and mapping by DNA-mRNA hybrid-arrested cell-free translation.
Authors: Paterson B M , Roberts B E , Kuff E L
Published: May 2006

We present a simple method for directly correlating structural gene sequences in DNA with their corresponding mRNAs. This is based upon the fact that mRNA hybridized with its complementary DNA will not direct the cell-free synthesis of a complete polypeptide. Full translational activity of the mRNA is recovered upon the heat melting of the hybrid. Utilizing the rabbit beta globin clone PbetaG1, we demonstrate the application of hybrid-arrested translation for the identification of structural gene sequences within recombinant DNA molecules. In addition, the method is used to locate and order precisely several adenovirus 2 polypeptides within the viral genome.
Food webs and the dimensionality of trophic niche space
Authors: Cohen Joel E.
Published: May 2006

If the trophic niche of a kind of organism is a connected region in niche space, then it is possible for trophic niche overlaps to be described in a one-dimensional niche space if and only if the trophic niche overlap graph is an interval graph. An analysis of 30 food webs, using the combinatorial theory of interval graphs, suggests that a niche space of dimension 1 suffices, with unexpectedly high frequency and perhaps always, to describe the trophic niche overlaps implied by real food webs in single habitats. Consequently, real food webs fall in a small subset of the set of mathematically possible food webs. That real food webs are compatible with one-dimensional trophic niche spaces, more often than can be explained by chance alone, has not been noticed previously.
Size-mediated onset of genetically determined maturation in the platyfish,
Xiphophorus maculatus
Authors: Sohn Joel Jay , Crews David
Published: May 2006

A single sex-lined gene that controls the maturation process had previously been identified in
Xiphophorus maculatus
. It was presumed that this gene controls the age of onset of maturation. We demonstrate that the gene is either activated or not inhibited from activity by the attainment of a critical weight rather than age.
Tritium suicide selection of mammalian cell mutants defective in the transport of neutral amino acids.
Authors: Finkelstein M C , Slayman C W , Adelberg E A
Published: May 2006

Mouse lymphocytic cells of the established line GF-14 were allowed to accumulate intracellular 3H-labeled aminoisobutyric acid (AIB), frozen, and stored over liquid N2. After internal radiation had reduced survival to 1 in 10(4), survivors were plated and tested for their ability to transport AIB. Out of 200 clones tested, two (designated GF-17 and GF-18) were found to have reductions to 13-35% of the parent in the rate of transport of AIB, L-alanine, L-proline, and L-serine; GF-18 also showed significant reductions in the rate of transport of L-glutamate and DL-cysteine. Little or no change was observed for 10 other amino acids or for thymidine. Kinetic analyses revealed that the mutants were not altered in Km for AIB uptake, but had Vmax values approximately 20% the value of the parent strain, GF-14, suggesting that either the number of AIB transport sites or the turnover rate of the sites has been reduced in the two mutants.
Role of high-molecular-weight kininogen in surface-binding and activation of coagulation Factor XI and prekallikrein.
Authors: Wiggins R C , Bouma B N , Cochrane C G , Griffin J H
Published: May 2006

In the contact phase of activation of the kinin-forming, intrinsic clotting, and fibrinolytic systems, high-molecular-weight kininogen acts as a cofactor for the activation of Factor XI, prekallikrein, and Hageman factor. One mechanism by which high-molecular-weight kininogen acts as a cofactor has been studied by using 125I-labeled Factor XI and prekallikrein in kaolin-activated normal human plasma and plasmas deficient in high-molecular-weight kininogen and Hageman factor. High-molecular-weight kininogen was found to be essential for normal binding and cleavage of both Factor XI and prekallikrein on the kaolin surface. Hageman factor was essential for cleavage but not for binding of Factor XI and prekallikrein to kaolin. In normal plasma 80% of the activated Factor XI remained surface-bound, whereas 80% of the kallikrein was not surface-bound. These findings are consistent with the hypothesis that, in the initial phase of contact activation, high-molecular-weight kininogen links both Factor XI and prekallikrein to the exposed surface where they are activated by surface-bound activated Hageman factor. Once activated, the Factor XI molecules remain localized at the site of activation, in contrast to the kallikrein molecules which are found largely in the surrounding plasma.
Sensory electrophysiology of bacteria: relationship of the membrane potential to motility and chemotaxis in Bacillus subtilis.
Authors: Miller J B , Koshland D E
Published: May 2006

The relationship of membrane potential to motility and chemotaxis of Bacillus subtilis has been tested by using the fluorescence of a cyanine dye as a probe of the potential. The dye fluorescence was found to be an indicator of membrane potential by correlation with triphenylmethylphosphonium ion distribution and with changes due to anaerobicity and ionophore addition. When the potential was sufficient for motility and constant over time, it was found that the absolute level of the potential did not affect the swimming behavior of the bacteria. Transient alteration of the membrane potential did, however, lead to changes in swimming behavior. Attractants were found to alter the swimming behavior of the bacteria without altering the membrane potential. Thus, change of the overall membrane potential of a normal B. subtilis is not required for chemotaxis, but such a change is sensed by the bacteria just as changing levels of attractants and repellents are sensed.
Location of pyridoxal phosphate in glycogen phosphorylase a.
Authors: Sygusch J , Madsen N B , Kasvinsky P J , Fletterick R J
Published: May 2006

The pyridoxal 5'-phosphate cofactor of glycogen phosphorylase a (1,4-alpha-D-glucan:orthophosphate alpha-glucosyltransferase, EC2.4.1.1.) has been positioned on the protomer with x-ray diffraction data, chemical markers, and sequence information. The electron density was computed from 3.0-A resolution phases calculated from four heavy-atom derivatives. The cofactor is buried inside the protomer adjacent to the glucose-binding site. The phosphoryl substrates Pi and glucose-1-P each bind at two sites on the protomer. At low concentrations, Pi and glucose-1-P bind in the same location as does the allosteric effector AMP, near the monomer-monomer interface and some 30 A from the glucose site. At high concentrations glucose-1-P also binds strongly at the glucose site, with its phosphate only 7.2 A from that of the cofactor. Inorganic phosphate can also bind at this site. Implications for the participation of the pyridoxal phosphate in the catalytic mechanism are discussed in the light of these structural findings as well as the wealth of indirect evidence in the literature.
Mechanism of action of nalidixic acid: Purification of
Escherichia coli nalA
gene product and its relationship to DNA gyrase and a novel nicking-closing enzyme
Authors: Sugino Akio , Peebles Craig L. , Kreuzer Kenneth N. , Cozzarelli Nicholas R.
Published: May 2006

A target protein for nalidixic and oxolinic acids in
Escherichia coli
, the
nalA
gene product (Pnal), was purified to homogeneity as judged by gel electrophoresis, using an
in vitro
complementation assay. It is a dimer of identical 110,000-dalton subunits. A polypeptide of this molecular weight is uniquely induced by a λ
nalA
transducing phage, thereby showing that the purified Pnal is a product of the
nalA
gene. Nalidixic and oxolinic acids inhibit DNA gyrase activity and induce formation of a relaxation complex analogue. Treatment of the complex with sodium dodecyl sulfate causes a doublestrand break in the DNA substrate and the resulting linear molecule seems covalently bound to protein. Complex formation, unlike the introduction of supertwists, does not require ATP or relaxed circular DNA and is insensitive to novobiocin. DNA gyrase from a strain with a
nalA
mutation conferring drug resistance (
nalA
r
) is 1/100 as sensitive to oxolinic and nalidixic acids with respect to inhibition of supertwisting and induction of the pre-linearization complex. Addition of Pnal restores drug sensitivity and stimulates DNA gyrase activity. DNA gyrase preparations and Pnal catalyze a third reaction sensitive to nalidixic and oxolinic acids, the ATP-independent relaxation of supertwister DNA. Relaxation by gyrase from
nalA
r
cells is drug resistant. The nicking-closing activity is distinct from
E. coli
ω protein in several properties, including the ability to relax positively supertwisted DNA. We postulate that the
nalA
gene product occurs in two molecular forms, as Pnal and as a gyrase component. Both forms catalyze nicking-closing, and inhibition of this activity by nalidixic and oxolinic acids may account for the inhibition of DNA synthesis by these drugs.
Structure of histone H1-DNA complex: Effect of histone H1 on DNA condensation
Authors: Hsiang Myrtle W. , Cole R. David
Published: May 2006

Large doughnut-shaped complexes were formed when histone H1 was mixed with DNA in amounts that extensively neutralized it. The doughnut shape is the most prevalent from observed by electron microscopy for linear double-stranded DNA independent of the molecular weight of the DNA in the range 1.2 × 10
6
to 25 × 10
6
at an H1/DNA input weight ratio of 1.3, at ionic strength 0.17. Doughnuts were not observed for single-stranded DNA-H1 complexes; instead, the complexes were globular. The circumference of the doughnutshaped particles indicates that much of the rigidity of duplex DNA in the complex has remained. Evidently, the condensation of the nucleohistone is constrained by the rigidity of duplex DNA and, under this constraint, surface contact with water is minimized by adopting a doughnut shape. Histone H5 causes a type of DNA condensation similar to that of H1 at comparable charge ratios. Core histones H2A-H2b, H3, and H4 complex with DNA to form globular aggregates of such small diameter that the duplex DNA in them must be much more tightly folded than is the case with the doughnut-shaped complexes. Because these histones are designed to fold DNA into nucleosomes 100 Å wide, they must destroy the rigidity of free duplex DNA, perhaps by forming kinks in the chain.
Flexibility of myosin rod, light meromyosin, and myosin subfragment-2 in solution.
Authors: Highsmith S , Kretzschmar K M , O'Konski C T , Morales M F
Published: May 2006

Myosin rod was prepared by papain proteolysis of myosin. The components of rod, light meromyosin (LMM) and subfragment-2 (S-2), were prepared by proteolysis of myosin and rod, respectively, using trypsin treated with tosylphenylalanine chloromethyl ketone. S-2, thus prepared, was of greater molecular weight than obtained previously, so that the combined molecular weights of LMM and S-2 were equal to that of rod, and S-2 contained virtually all of the region of the rod susceptible to trypsin. Electro-optical measurements were made on the three fragments in 2 mM sodium pyrophosphate, pH 9.3 at 3 degrees, over a large range of protein concentrations. Analysis of the relaxation of birefringence, at low protein concentration where there was no aggregation, showed that LMM (relaxation time 13.1 micros) behaves as a rigid cylinder. Rod (relaxation time 41.2 micros) and S-2 (relaxation time 6.0 micros) had relaxation rates that were too fast for rigid molecules of their dimensions, and therefore are not straight rods. This implies that myosin rod is flexible in the S-2 portion, presumably in the region susceptible to proteolysis. The implications of rod flexibility for the mechanism of muscle contraction are discussed.
Phylogenetic structure of the prokaryotic domain: The primary kingdoms
Authors: Woese Carl R. , Fox George E.
Published: May 2006

A phylogenetic analysis based upon ribosomal RNA sequence characterization reveals that living systems represent one of three aboriginal lines of descent: (
i
) the eubacteria, comprising all typical bacteria; (
ii
) the archaebacteria, containing methanogenic bacteria; and (
iii
) the urkaryotes, now represented in the cytoplasmic component of eukaryotic cells.
Simple Mendelian inheritance of the reiterated ribosomal DNA of yeast.
Authors: Petes T D , Botstein D
Published: May 2006

A diploid strain of yeast (Saccharomyces cerevisiae) was found to be heterozygous for two forms of the highly repetitious ribosomal DNA. These forms could be distinguished by the pattern of fragments produced after digestion with the site-specific restriction endonuclease EcoRI. The mode of inheritance of ribosomal DNA was determined by tetrad analysis. Of 14 tetrads analyzed, 12 clearly showed the ribosomal DNA forms segregating as a single Mendelian unit. The simplest interpretation of this result is that all of the approximately 100 copies of the ribosomal DNA genes of the yeast cell are located on one chromosome and that meiotic recombination within these genes is suppressed. Two of the 14 tetrads showed the segregation patterns expected as the result of mitotic recombination within the ribosomal DNA.
Defects of glycosyltransferase activities in human fibroblasts of Pk and p blood group phenotypes.
Authors: Kijimoto-Ochiai S , Naiki M , Makita A
Published: May 2006

We demonstrate that human fibroblasts of the rare Pk phenotype lack globoside, which was identified as the blood group P antigen, and that p cells possess neither globoside nor trihexosyl ceramide, which was identified as Pk antigen. Our investigations indicate also that these glycosphingolipid patterns are most likely caused by inherited preferential biosynthetic pathways in the abnormal phenotypes rather than by excess catabolism of the antigens. Evidence is presented that the fibroblasts of Pk phenotype lack beta-N-acetylgalactosaminyltransferase (globoside synthetase; UDP-N-acetylgalactosamine:trihexosylceramide beta-N-acetylgalactosaminyltransferase; EC 2.4.1.79) activity, and those of p are deficient in alpha-galactosyltransferase (trihexosylceramide synthetase; UDP galactose:lactosylceramide alpha-galactosyltransferase) and possibly also in globoside synthetase. The diminished globoside synthetase activity in p cells, however, is not caused by the defect in the gene coding for this enzyme. It appears, rather, to be caused by a failure in gene expression because one-third of Pk X p hybrids became able to express P antigenicity with a time lag of 3-4 days after cell fusion [Fellous, M., Gerbal, A., Nobillot, G. & Weils, J. (1977) Vox Sang. 32, 262-268].
Molybdenum cofactors from molybdoenzymes and
in vitro
reconstitution of nitrogenase and nitrate reductase
Authors: Pienkos Philip T. , Shah Vinod K. , Brill Winston J.
Published: May 2006

A molybdenum cofactor (Mo-co) from xanthine oxidase (xanthine:oxygen oxidoreductase, EC 1.2.3.2) can be isolated from the enzyme by a technique that has been used to isolate an iron-molybdenum cofactor (FeMo-co) from component I of nitrogenase.
N
-Methylformamide is used for the extraction of these molybdenum cofactors. Mo-co from xanthine oxidase activates nitrate reductase (NADPH:nitrate oxidoreductase, EC 1.6.6.2) in an extract from
Neurospora crassa
mutant strain Nit-1; however, FeMo-co is unable to activate nitrate reductase in strain Nit-1. Mo-co from xanthine oxidase is unable to activate nitrogenase in an extract of
Azotobacter vinelandii
mutant strain UW45. Inactive component I in this extract can be activated by FeMo-co. These results indicate that nitrate reductase and xanthine oxidase share a common molybdenum cofactor, but this cofactor is different from the molybdenum cofactor in nitrogenase.

A. vinelandii
synthesizes both Mo-co and FeMo-co. Mo-co is produced when the cells fix N
2
and also when they are repressed for nitrogenase synthesis by growth in a medium containing excess ammonium. However, FeMo-co is not produced when cells are grown in an ammonium-containing medium.

Partially purified preparations of component I from
A. vinelandii
and
Klebsiella pneumoniae
contain both FeMo-co and Mo-co. The presence of both FeMo-co and Mo-co activities in partially purified preparations of component I explains previous reports of activation of inactive nitrate reductase in strain Nit-1 by acid-treated component I of nitrogenase. The Mo-co can be separated from FeMo-co in these preparations by chromatography on Sephadex G-100 in
N
-methylformamide. Both FeMo-co and Mo-co are sensitive to oxygen.
Presence of protein A24 in rat liver nucleosomes.
Authors: Goldknopf I L , French M F , Musso R , Busch H
Published: May 2006

Two-dimensional gel profiles of the 0.2 M H2SO4-soluble proteins of monomer nucleosomal fractions were found to contain protein A24. Protein A24 is of interest because it is composed of histone 2A and "ubiquitin", apparently joined by an isopeptide linkage [Goldknopf, I.L. & Busch, H. (1977) Proc. Natl. Acad. Sci. USA 74, 864-868; Hunt, L.T. & Dayhoff, M.O. (1977) Biochem. Biophys. Res. Commun. 74, 650-655]. Monomer nucleosomal fractions were obtained by sucrose density gradient centrifugation of micrococcal nuclease digests of rat liver nuclei. As shown by their DNA size, the monomer fractions were highly purified. Proteins A24 and Bu, another protein of unknown characteristics, were found along with histones 1, 2A, 2B, 3, and 4 in the monomer fractions in relative amounts similar to those found in extracts from whole nuclei and chromatin. Other acid-soluble proteins found in the nuclear and chromatin extracts were essentially absent from the monomer fraction. Inasmuch as protein A24 and Bu were found in lesser amounts than the histones, it is suggested that they are associated with specialized subsets of nucleosomes.
Antigen-binding mutants of mouse myeloma cells.
Authors: Cook W D , Scharff M D
Published: May 2006

A cultured mouse myeloma cell line, S107, that secretes an IgA phosphocholine-binding immunoglobulin has been cloned in soft agar and overlaid with phosphocholine-hemocyanin. Spontaneous mutants that secrete immunoglobulin with a decreased ability to precipitate antigen were detected with this plate assay and occur at a very high frequency. From one such mutant, phenotypic revertants arise spontaneously with a frequency of 0.28-2.8%. This mutant and one of its revertants were studied, and they were found to differ from the parent and from each other serologically and in antigen binding. While it is not yet clear whether these findings bear any relationship to the normal generation of antibody diversity, they do indicate that it is possible to generate antigen binding diversity in somatic cells.
Selectivity of action of an antiherpetic agent, 9-(2-hydroxyethoxymethyl)guanine
Authors: Elion Gertrude B. , Furman Phillip A. , Fyfe James A. , Miranda Paulo de , Beauchamp Lilia , Schaeffer Howard J.
Published: May 2006

A guanine derivative with an acyclic side chain, 2-hydroxyethoxymethyl, at position 9 has potent antiviral activity [dose for 50% inhibition (ED
50
) = 0.1 μM] against herpes simplex virus type 1. This acyclic nucleoside analog, termed acycloguanosine, is converted to a monophosphate by a virus-specified pyrimidine deoxynucleoside (thymidine) kinase and is subsequently converted to acycloguanosine di- and triphosphates. In the uninfected host cell (Vero) these phosphorylations of acycloguanosine occur to a very limited extent. Acycloguanosine triphosphate inhibits herpes simplex virus DNA polymerase (DNA nucleotidyltransferase) 10-30 times more effectively than cellular (HeLa S3) DNA polymerase. These factors contribute to the drug's selectivity; inhibition of growth of the host cell requires a 3000-fold greater concentration of drug than does the inhibition of viral multiplication. There is, moreover, the strong possibility of chain termination of the viral DNA by incorporation of acycloguanosine.

The identity of the kinase that phosphorylates acycloguanosine was determined after separation of the cellular and virus-specified thymidine kinase activities by affinity chromatography, by reversal studies with thymidine, and by the lack of monophosphate formation in a temperature-sensitive, thymidine kinase-deficient mutant of the KOS strain of herpes simplex virus type 1 (tsA1).
Kinetic evidence for an intermediate in the deacetylation of monoacetyl-chymotrypsin.
Authors: Chibber B A , Tomich J M , Mertz E T , Viswanatha T
Published: May 2006

Mono[14C]acetyl-chymotrypsin was prepared by treating alpha-chymotrypsin with a 10-fold molar excess of p-nitrophenyl[14C]acetate at pH 5, and the acetylated enzyme was isolated free of excess reagents by gel filtration. Deacetylation at pH 6.0 was followed by observing the decrease in acid-precipitable radioactivity and provided a first-order rate constant of 0.02 +/- 0.008 min-1. Reactivation of the acetylated protein was followed by continuously monitoring the appearance of esterolytic activity towards alpha-N-acetyltyrosine ethyl ester. Reactivation at pH 6.0 occurred exponentially with a first-order rate constant of 0.2 +/- 0.015 min-1, the reactivated enzyme exhibiting an apparent catalytic contant (k' cat) of 1200 +/- 60 min-1, which decreased to a value of 945 +/- 15 min-1 by an apparent first-order process with a rate constant of 0.025 +/- 0.006 min-1. These results are interpreted in terms of a two-step deacetylation of monoacetyl-chymotrypsin involving an acetylated intermediate with esterase activity.
Calcium dependence of glucocorticoid-induced lymphocytolysis.
Authors: Kaiser N , Edelman I S
Published: May 2006

A potent glucocorticoid, triamcinolone acetonide (9alpha-fluoro-11beta, 16alpha,17alpha, 21-tetrahydroxypregna-1,4-diene-3,20-dione-16,17-acetonide) and a divalent cation ionophore (A23187) had similar effects in vitro on [3H]uridine uptake and on lysis of thymocytes of adrenalectomized rats. Removal of Ca2+ from the medium blunted the cytolytic action of triamcinolone acetonide and virtually eliminated that of A23187. In Ca2+-free media, treatment of the thymocytes for 15 hr with triamcinolone acetonide or A23187 followed by re-introduction of Ca2+ resulted in a rapid decrease in cell survival. Based on the time courses of the responses, triamcinolone acetonide and A23187 evoked proportionate increases in 45Ca uptake and lysis of the thymocytes. These findings implicate enhanced Ca2+ uptake in glucocorticoid-dependent lymphocytolysis.
Thymosin alpha1: isolation and sequence analysis of an immunologically active thymic polypeptide.
Authors: Goldstein A L , Low T L , McAdoo M , McClure J , Thurman G B , Rossio J , Lai C Y , Chang D , Wang S S , Harvey C , Ramel A H , Meienhofer J
Published: May 2006

The amino acid sequence of a biologically active polypeptide isolated from calf thymus, termed thymosin alpha1, has been determined. Thymosin alpha1 is a heat stable, highly acidic molecule composed of 28 amino acid residues. This peptide is one of several present in thymosin fraction 5 that may participate in the regulation, differentiation and function of thymus-dependent lymphocytes (T cells). A nomenclature for the family of polypeptides present in thymosin fraction 5 is suggested.
beta-Endorphin: behavioral and analgesic activity in cats.
Authors: Meglio M , Hosobuchi Y , Loh H H , Adams J E , Li C H
Published: May 2006

beta-Endorphin has been shown to possess potent behavioral and antinociceptive activities when administered intraventricularly in cats. On a molar basis, beta-endorphin is 72-96 times more potent than morphine and its actions are blocked by the specific opiate antagonist, naloxone.
Cell-cycle-related changes of 3':5'-cyclic GMP levels in Novikoff hepatoma cells.
Authors: Zeilig C E , Goldberg N D
Published: May 2006

Intracellular and extracellular levels of 3':5'-cyclic GMP and 3':5'-cyclic AMP were studied in synchronized Novikoff rat hepatoma cells. Intracellular levels of cyclic GMP increased spontaneously from 2-fold (without colcemid) to 10-fold (with colcemid), in proportion to the number of cells in mitosis. As cells entered mitosis, cellular cyclic AMP declined simultaneously with the rise in cyclic GMP. These reciprocal changes in cyclic nucleotide levels were reversed as cells passed out of metaphase and through anaphase. Maximum cyclic AMP and minimum cyclic GMP concentrations occurred during G-1. Less marked reciprocal fluctuations in both cyclic nucleotides were also found in S-phase and early G-2, where the ratio of cyclic AMP to cyclic GMP concentrations first fell and then increased. These changes in cyclic nucleotide ratios were closely correlated with major cell-cycle transitions at the boundaries between G-1/S-phase, S-phase/G-2, G-2/prophase, and metaphase/anaphase. Most, but not all, of the extracellular cyclic nucleotides were extruded when cells traversed mitosis. Colcemid or vinblastine completely prevented the appearance of extracellular cyclic AMP but augmented the appearance of extracellular cyclic GMP in parallel with the accumulation of mitotic cells. These results reflected changes in intracellular cyclic nucleotides and indicated that increased intracellular turnover of cyclic GMP and cyclic AMP occurred before and after metaphase, respectively. Elevated cyclic GMP levels during mitosis and S-phase are consistent with potential modulatory roles for this cyclic nucleotide in proliferation.
An extracellular inducer of asexual plasmodium formation in Physarum polycephalum.
Authors: Youngman P J , Adler P N , Shinnick T M , Holt C E
Published: May 2006

Asexual conversion of amoebae to plasmodia was studied in the Colonia isolate of the myxomycete, Physarum polycephalum. When a culture of Colonia amoebae is grown on a bacterial lawn, a period of amoebic growth precedes the appearance of cells committed to the plasmodial state. The onset of plasmodium production appears to be related to amoebic nutrition since cultures supplied with fewer bacteria display earlier differentiation. For a period of time after differentiation is initiated, conversion of amoebae to plasmodia is rapid and proceeds as an exponential function of time. A filter-transmissible substance, apparently released by differentiating cells, is implicated in the control of this rapid conversion.
Studies on the control of development: isolation of Bacillus subtilis mutants blocked early in sporulation and defective in synthesis of highly phosphorylated nucleotides.
Authors: Rhaese H J , Hoch J A , Groscurth R
Published: May 2006

To test our model on the mechanism of initiation of differentiation in Bacillus subtilis, we tested early blocked (stage 0) sporulation mutants for their ability to synthesize highly phosphorylated nucleotides. We also isolated early blocked asporogenous mutants with the aid of the intercalating drug tilorone. Among all mutants tested we found that the spo0F-bearing strain was unable to synthesize adenosine 3'(2')-triphosphate 5'-triphosphate, pppAppp. A revertant of this mutant regained the ability to both sporulate and synthesize pppAppp. Ribosomes of the asporogenous mutant isolated at T2 (2 hr after the end of logarithmic growth) of sporulation, in contrast to the wild type, do not synthesize adenosine 3'(2')-diphosphate 5'-diphosphate, ppApp, or adenosine 3'(2')-diphosphate 5'-triphosphate, pppApp, but synthesize guanosine 3'(2')-diphosphate 5'-diphosphate, ppGpp, and guanosine 3'(2')-diphosphate 5'-triphosphate, pppGpp. This behavior is characteristic of ribosomes from vegetative, not sporulating, cells. Ribosomes from the sporogenous revertant behave like those of the wild type. The results suggest that the spo0F mutation may be a mutation in the structural gene for pppAppp synthetase. The inability to synthesize pppAppp in this strain also prevents the formation of "sporulation-specific ribosomes," i.e., ribosomes that synthetize ppApp and pppApp. The present experiments suggest that the nucleotide pppAppp participates in the initiation of sporulation by triggering a sequencies of events required for the production of heat-resistant spores.
A protease inhibitor blocks SOS functions in Escherichia coli: antipain prevents lambda repressor inactivation, ultraviolet mutagenesis, and filamentous growth.
Authors: Meyn M S , Rossman T , Troll W
Published: May 2006

Inhibition of DNA synthesis in E. coli by treatment with carcinogenic and mutagenic agents results in the coordinate expression of a group of diverse functions (SOS functions) including lambda prophage induction, filamentous growth, and an error-prone DNA repair activity (SOS repair) believed to be responsible for ultraviolet mutagenesis. It has been proposed that this SOS induction proceeds via irreversible proteolytic inactivation of repressor(s) for SOS functions. To test this hypothesis, we investigated the effect of a protease inhibitor, antipain [(1-carboxy-2-phenylethyl)carbamoyl-L-arginyl-L-valylargininal], on SOS induction. We found that 0.5 mM antipain (which has no effect on cell growth, overall RNA and protein synthesis, or induction of beta-galactosidase) drastically decreases mutagenesis. Antipain also blocks expression of thermally induced mutator activity (another manifestation of SOS repair) and filamentous growth in a tif-1 mutant that expresses SOS functions at 42 degrees without inhibition of DNA synthesis or detectable DNA damage. Furthermore, antipain inhibits thermal induction of lambda prophage in the tif-1 mutant without affecting the kinetics of thermal induction of lambdacI857 prophage. This lambda mutant codes a temperature-sensitive repressor that is directly destroyed by heat and does not require the SOS induction pathway for inactivation at 42 degrees. From our results we conclude that antipain inhibits lambda prophage induction by blocking proteolytic inactivation of lambda repressor and that it inhibits the induction or expression of SOS repair and filamentous growth. Our results suggest a role for proteolytic cleavage in the regulation of SOS functions.
Biochemical catalysis involving coenzyme B-12: a rational stepwise mechanistic interpretation of vicinal interchange rearrangements.
Authors: Corey E J , Cooper N J , Green M L
Published: May 2006

A mechanism is proposed for the catalytic action of coenzyme B-12 which is consistent with current knowledge of organometallic reactions and with the experimental data now available from biochemical studies. A key feature of the proposal is an electrocyclic cleavage of the coenzyme that reduces cobalt and also leads to a 1,19-seco-corrin. The seco-corrin serves as a tridentate ligand about Co(I). This arrangement permits the metal to take part in the kinds of organometallic reactions that are ideal for coenzyme B-12 catalysis, including oxidative addition and its reverse, reductive elimination. It is further proposed that the rearrangement steps involve cobaltcarbene complexes.
Age incidence curves for cancer.
Authors: Watson G S
Published: May 2006

Age incidence curves for carcinomas tend to be linear on doubly logarithmic graphs. This fact has been used to support the somatic mutation hypothesis. It is shown that this should also be expected on the epigenetic hypothesis when the reversible cellular changes are programmed to occur randomly. Such changes could be programmed differently to obtain the age incidence curves observed for other cancers, e.g., leukemias and sarcomas.
Secondary structural complementarity between DNA and proteins.
Authors: Church G M , Sussman J L , Kim S H
Published: May 2006

A model for the complex between double-stranded DNA and a beta-ribbon (a two-stranded antiparallel beta-sheet) of proteins is proposed as one of the possible modes of structural recognition between DNA and proteins. In this model, the contact occurs on the narrow groove of DNA, and the symmetry elements as well as the repeat distances of DNA and the beta-ribbon coincide, thus providing favorable complementary contacts.
The amino acid sequence of beta-galactosidase of Escherichia coli.
Authors: Fowler A V , Zabin I
Published: May 2006

The amino acid sequence of beta-galactosidase was determined. The protein contains 1021 amino acid residues in a single polypeptide chain. The subunit molecular weight calculated from the sequence is 116,248. The sequence determination, carried out mainly by conventional methods, was aided by complementation tests, by the use of termination mutant strains, and by a new immunochemical method. The five residue sequence Thr-Pro-His-Pro-Ala appears twice within the polypeptide chain, but no other striking homologous features are evident.
Effect of polyploidy on phosphoglucose isomerase diversity in Festuca microstachys.
Authors: Adams W T , Allard R W
Published: May 2006

Studies of the inheritance of electrophoretic banding patterns in Festuca microstachys support the hypothesis that three closely related loci, one located in each of the three ancestral genomes, code the multiple phosphoglucose isomerase (glucosephosphate isomerase; D-glucose-6-phosphate ketol-isomerase, EC 5.3.1.9) variants found in this hexaploid species. The close relationship among the three loci is indicated by the observation that hybrid enzymes of intercistronic origin form when the loci in different genomes carry alleles coding homodimers with unlike migration rates. Homozygous individuals fixed for different alleles in different genomes produce hybrid enzymes and, when self-fertilized, they breed true for isozyme patterns normally found only in the heterozygotes of diploid species. Biochemical diversity due to this "fixed heterozygosity" is high in F. microstachys; although this species is more than 99% self-fertilized the proportion of individuals with at least one heterodimer exceeded 61% in all of the 16 natural populations studied and it exceeded 92% in 11 of the populations. This great biochemical diversity may contribute to the ability of F. microstachys to survive in the wide range of habitats in which it is found over western North America.
Structural identification of the major DNA adduct formed by aflatoxin B1 in vitro.
Authors: Essigmann J M , Croy R G , Nadzan A M , Busby W F , Reinhold V N , Büchi G , Wogan G N
Published: May 2006

The covalent binding of the hepatocarcinogen aflatoxin B1 by rat liver microsomes to calf thymus DNA resulted in a binding level equal to one aflatoxin residue per 60 DNA nucleotides. An aflatoxin derivative-guanine adduct was efficiently liberated from DNA with formic acid. Analytical reversed-phase high-pressure liquid chromatography of the DNA hydrolysate revealed that approximately 90% of the carcinogen bound to DNA could be accounted for as a single component. Preparative high-pressure liquid chromatography was used to isolate sufficient quantities of the adduct for structural analysis from large quantities (340 mg) of DNA. A combination of spectral and chemical data indicates that the major product of the interaction of metabolically activated aflatoxin B1 and DNA is 2,3-dihydro-2-(N7-guanyl)-3-hydroxyaflatoxin B1 with the guanine and hydroxyl functions possessing a trans configuration. The structural data support the hypothesis that the putative 2,3-oxide of aflatoxin B1 is quantitatively important as an intermediate in the binding of aflatoxin B1 to nucleic acids.
Interaction of filamin with f-actin in solution.
Authors: Wang K , Singer S J
Published: May 2006

Filamin is a major high-molecular-weight protein in smooth muscle which was recently identified and isolated [Wang, K., Ash, J. F. & Singer, S. J. (1975) Proc. Natl. Acad. Sci. U.S.A. 72, 4483-4486]. In the present studies, we shown that highly purified chicken gizzard filamin and muscle F-actin react in solution to form aggregates containing both proteins. Occasionally, these aggregates coagulate and contract into a dense gel in the absence of MgATP or CaATP. Immunofluorescence and electron microscopic studies suggest that the F-actin filaments are collected into fiber bundles and a crosslinked fiber meshwork by the binding of filamin molecules. These studies suggest that the function of filamin intact cells may be to regulate the ultrastructural state of F-actin filaments in a variety of dynamic cellular processes.
3′:5′-Cyclic AMP-dependent 3′:5′-cyclic GMP accumulation in
Dictyostelium discoideum
Authors: Mato José M. , Krens Frans A. , Van Haastert Peter J. M. , Konijn Theo M.
Published: May 2006

Suspensions of 3′:5′-cyclic AMP (cAMP)-sensitive cells of
Dictyostelium discoideum
responded to a cAMP pulse with increased 3′:5′-cyclic GMP (cGMP) levels. Under the assay conditions used (2 × 10
8
cells per ml in 10 mM phosphate buffer, pH 6.0) cAMP (5 × 10
-8
M final concentration) increased cGMP levels from 1 pmol per 10
7
cells to 7 pmol per 10
7
cells in 10 sec and basal levels were recovered in 20-25 sec. cGMP accumulation did not occur when cells were in the cAMP-insensitive stage. cAMP-sensitive cells responded with increased cGMP levels when triggered by 5 × 10
-8
M 5′-CH
2
-cAMP or 10
-5
M adenosine-5′-methylmonophosphate (5′-AMPMe) but not after addition of 5 × 10
-8
M 3′:5′-cyclic IMP (cIMP) or 5 × 10
-8
M 5′-AMP. As agonists of cAMP, 5′-CH
2
-cAMP and 5′-AMPMe have, respectively, more than 10% and 1% the chemotactic activity of cAMP, while cIMP has 0.01% the activity of cAMP and 5′-AMP is inactive up to a concentration of 10
-3
M. cAMP-mediated cGMP formation was dependent upon cAMP concentration, with a half-maximal cAMP concentration of about 10
-8
M. This cAMP concentration agrees closely with that necessary for half-maximal receptor occupation. cAMP-mediated cGMP formation was independent of the presence of extracellular Ca
2+
; cell aggregation and chemotaxis were also independent of the presence of external Ca
2+
. Therefore, cAMP action does not depend on stimulation of the Ca
2+
influx. cAMP was found to mediate desensitization of cAMP-dependent cGMP formation. Addition of 5 × 10
-8
M cAMP to sensitive cells induced a desensitization period that lasted 1-5 min. Desensitization was dependent on the cAMP concentration. Finally, we propose that the translation of a chemotactic signal from the cell surface to pseudopod formation in
Dictyostelium
involves changes in the levels of cGMP.
Bacteriophage phiX174: gene A overlaps gene B.
Authors: Weisbeek P J , Borrias W E , Langeveld S A , Baas P D , Van Arkel G A
Published: May 2006

The map position of several phiX174 mutations in the genes A and B was determined by marker rescue with DNA fragments produced by the restriction enzymes Hha I, HindII, Hae III, and Alu I. All the gene B mutants were found to be located within gene A. Genetic complementation and analysis of phage-specific protein synthesis show that, under restrictive conditions, nonsense mutants in gene A do not block the synthesis and activity of the B protein and nonsense mutants in gene B do not affect the gene A function. The map position of the COOH-terminal end of gene A was determined using an amber mutant that synthesizes slightly shortened A and A proteins. It is concluded from these experiments that gene A overlaps gene B completely (or almost completely) and that the overlap region can be translated in two ways with different reading frames: one frame for the synthesis of the A and A proteins and another for the synthesis of the B protein.
Maintenance of genetic variability under mutation and selection pressures in a finite population.
Authors: Li W H
Published: May 2006

Formulas are developed for the distribution of allele frequencies and the mean and variance of heterozygosity under mutation and selection pressures. In large populations, even slight selection drastically changes the shape of the distribution of allele frequencies and reduces heterozygosity. On the other hand, the number of rare alleles in a sample is much less affected by selection. Under genic selection, heterozygosity may decrease with increasing population size. As a test statistic, the variance of heterozygosity can be used to detect the presence of selection, though it is not efficient when selection is very slight.
On the location of palindromes in immunoglobulin genes.
Authors: Wuilmart C , Urbain J , Givol D
Published: May 2006

We present a statistical method for detection of palindromes in mRNA or DNA, starting from the protein sequence. Analysis of immunoglobulin genes by this method demonstrates that palindromic sequences are not randomly distributed. They are located at each side of the hypervariable regions in the variable (V) genes, whereas no such regular design is observed in the constant (C) genes. In addition, palindromic sequences overlap the V-C junction in all immunoglobulin classes and significant palindromes are present near residue 216 of the heavy chain, which is the end of deletions in many heavy chain diseases. The relevance of these palindromes to gene translocation and generation of diversity in antibodies is discussed.
Isolation and identification of aliphatic short-chain acylcarnitines from beef heart: possible role for carnitine in branched-chain amino acid metabolism.
Authors: Bieber L L , Choi Y R
Published: May 2006

Aliphatic acylearnitines isolated from a water-soluble fraction of beef heart have been characterized by gas chromatography and mass spectrometry. The following acyl residues derived from the acylcarnitine fraction were unequivocally identified: acetyl, propionyl, isobutyryl, butyryl, alpha-methylbutyryl, valeryl, isovaleryl, tiglyl, and caproyl. beta-methylcrotonyl and methacrylyl were tentatively identified. This occurrence of considerable quantities of branched-chain acylcarnitines indicates a role for carnitine in branched-chain amino acid metabolism.
Unique DNA associated with mitochondria in the “S”-type cytoplasm of male-sterile maize
Authors: Pring D. R. , Levings C. S. , Hu W. W. L. , Timothy D. H.
Published: May 2006

Mitochondrial DNAs were prepared from maize lines with normal cytoplasm and with the T, C, S, and EP sources of male-sterile cytoplasms. Agarose gel electrophoresis of these preparations revealed a main high-molecular-weight DNA band. In addition, the S cytoplasm was characterized by the presence of two faster migrating DNAs of molecular weight 3.42 to 3.48 × 10
6
and 4.01 to 4.10 × 10
6
. Electron microscopy showed these unique DNAs to be of different length, but their molecular configuration was not clearly established. It is possible that these unique DNAs represent physical evidence of an episomal system previously postulated to function in the S male-sterile cytoplasm.
Hemoglobin F synthesis in vitro: evidence for control at the level of primitive erythroid stem cells.
Authors: Papayannopoulou T , Brice M , Stamatoyannopoulos G
Published: May 2006

The in vitro regulation of fetal hemoglobin (HbF was investigated in clones of cultured adult human erythroid cells by in situ immunofluorescent identification of the hemoglobins synthesized. Formation of Hb F-containing clones was enhanced by erythropoietin and by culture conditions favoring the proliferation of less-differentiated stem cells of the burst-forming-unit type. Burst-forming units differed in their capacity to direct Hb F synthesis in their terminally differentiated progeny. A class of early precursors that can produce descendent stem cells with or without commitment to Hb F production was identified. The findings suggest that the capability for expression of Hb F in terminally differentiated cells of the adult is determined at the level of less-differentiated erythroid stem cells with characteristics of burst-forming units. It is proposed that the regulation of Hb F synthesis in vivo is also linked to the process of differentiation of the erythroid stem cells and that the patterns of Hb F synthesis during ontogeny reflect the attainment of progressively higher levels of differentiation of erythroid stem cells as development proceeds.
Measurement of daily cholesterol synthesis rates in man by assay of the fractional conversion of mevalonic acid to cholesterol.
Authors: McNamara D J , Ahrens E H , Samuel P , Crouse J R
Published: May 2006

A significant correlation has been found in man between total daily cholesterol synthesis rates as determined by sterol balance measurements and the fraction of intravenously administered R-[5-14C]mevalonic acid converted to cholesterol. In 30 studies it was found that the mean daily cholesterol synthesis rates estimated by sterol balance measurements ranged from 395 to 3047 mg/day, whereas the fractional conversion of mevalonate to cholesterol varied from 0.28 to 0.99. The two parameters correlated with a coefficient of 0.87, P less than 0.001. This method for estimating cholesterol synthesis rates requires low doses of radioisotopic materials (25 muCi of [14C]mevalonate and 5 muCi of [3H]cholesterol) and less than 1 hr of the patient's time; it can be repeated at intervals of 3 weeks and reflects cholesterol synthesis over a short period of time.
Blue copper proteins: Synthesis, spectra, and structures of Cu
I
N
3
(SR) and Cu
II
N
3
(SR) active site analogues
Authors: Thompson Jeffery S. , Marks Tobin J. , Ibers James A.
Published: May 2006

The reaction of Cu(SR) or [Cu(SR)][ClO
4
] derivatives (SR =
p
-nitrobenzenethiolate or
O
-ethylcysteinate) with potassium hydrotris(3,5-dimethyl-1-pyrazolyl)borate produces redox pairs of the stoichiometry Cu
I
N
3
(SR) and Cu
II
N
3
(SR). These complexes are well-defined synthetic approximations to the proposed N
3
S binding sites of blue (type 1) copper electron transfer proteins. The compounds were investigated by a variety of chemical and spectral (optical, resonance Raman, and electron paramagnetic resonance) techniques; the complex K[Cu(hydrotris(3,5-dimethyl-1-pyrazolyl)borate)(
p
- NO
2
C
6
H
4
S]-2 acetone was also studied by single-crystal x-ray diffraction methods. The spectrochemical characteristics of the Cu
II
N
3
(SR) species are in large part similar to the native system and thus provide some perspective regarding the origin of the unique type 1 spectral parameters and electron transfer properties.
Tumorigenicity of five dihydrodiols of benz(a)anthracene on mouse skin: exceptional activity of benz(a)anthracene 3,4-dihydrodiol.
Authors: Wood A W , Levin W , Chang R L , Lehr R E , Schaefer-Ridder M , Karle J M , Jerina D M , Conney A H
Published: May 2006

Benz[a]anthracene and the five metabolically possible vicinal trans dihydrodiols of benz[a]anthracene were tested for ability to initiate skin tumors in CD-1 female mice. A single topical application of 0.4-2.0 mumol of hydrocarbon was followed 18 days later by twice weekly applications of the skin promoter 12-O-tetradecanoylphorbol-13-acetate. Comparisons of latency period, percent of mice with tumors, and number of papillomas observed per mouse indicated that benz[a]anthracene 1,2-, 5,6-, 8,9-, and 10, 11-dihydrodiols were all less active tumor initiators than was benz[a]anthracene. The high tumorigenicity of benz[a]anthracene 3,4-dihydrodiol, presumably the result of metabolism to either or both of the diastereomeric benz[a]anthracene 3,4-diol-1,2-epoxides, supports the bay region theory of polycyclic hydrocarbon carcinogenicity and provides the first example of a proximate carcinogenic metabolite that is much more active than the parent hydrocarbon on mouse skin.