bluuebunny/crossref_metadata_2025 huggingface dataset:
Mechanism of the Enzymatic Synthesis of Cardiolipin in
Escherichia coli
Authors: Hirschberg Carlos B. , Kennedy Eugene P.
Published: May 2006

In previous studies, the enzymatic conversion of phosphatidylglycerol to cardiolipin (diphosphatidylglycerol) in cell-free preparations from
E. coli
was shown to be stimulated by the addition of CDP-dipalmitin, suggesting the participation of the cytidine coenzyme as phosphatidyl donor. The present communication, however, presents three lines of evidence supporting the following mechanism for the synthesis of cardiolipin in
E. coli
.

2 Phosphatidylglycerol → cardiolipin + glycerol

When CDP-dipalmitin labeled with
32
P in the phosphatidyl moiety was incubated with phosphatidyl[2-
3
H]-glycerol, the cardiolipin produced in the enzymatic reaction was labeled with tritium, but not with
32
P. Thus, CDP-diglyceride stimulates the reaction but does not participate as phosphatidyl donor. When [
28
P]phosphatidyl[2-
3
H]glycerol was used as substrate, the ratio of tritium to
32
P in the cardiolipin product was only half of that in the starting phosphatidylglycerol, consistent with the elimination of 1 mol of glycerol during conversion to cardiolipin. Finally, free glycerol produced during the reaction has been unambiguously identified by phosphorylation with ATP in a reaction catalyzed by glycerol kinase (EC 2.7.1.30), followed by chromatographic isolation of labeled
sn
-3-glycero-3-phosphate.
Ribosomes Cannot Interact Simultaneously with Elongation Factors EF Tu and EF G
Authors: Richman Nathan , Bodley James W.
Published: May 2006

Prior binding of EF G and GDP to 70S ribosomes from
Escherichia coli
prevents the subsequent binding of aminoacyl-tRNA, mediated by EF Tu. However, the interaction of EF Tu·GTP·aminoacyl-tRNA with the 30S subunit, which results in aminoacyl-tRNA binding without GTP hydrolysis, appears to be unaffected by EF G, GDP, and fusidic acid. We conclude that elongation factors Tu and G cannot interact simultaneously with the ribosome. The simplest interpretation of these and earlier data is that EF G and EF Tu interact with the same, or overlapping, 50S ribosomal sites in the course of GTP hydrolysis associated with translocation and aminoacyl-tRNA binding, respectively. In any event, these factors must alternate in binding to the ribosome in the course of each elongation cycle.
Degradation of Single- and Double-Stranded RNA by Frog Virus 3
Authors: Palese Peter , Koch Gebhard
Published: May 2006

Purified preparations of frog virus 3 possess ribonuclease activities directed against single-and double-stranded RNA. Double-stranded RNAs isolated from purified reovirus type 3 and from HeLa cells infected with poliovirus and single-stranded poliovirus RNA from purified virus are readily degraded by incubation with frog virus 3. The mode of action of the nucleases is endonucleolytic. Under the assay conditions used for the viral enzyme, crude extracts of uninfected HeLa, L, and baby hamster kidney cells did not show enzyme activity against double-stranded RNA but exhibited activity against single-stranded RNA. The dependence of the viral nucleases on divalent cations for optimal activity and the inhibition of the cleavage of single-stranded RNA by 0.2 M NaCl suggests that the enzymes are either virus-coded or virus-induced.
Initiation of Hemoglobin Synthesis: Comparison of Model Reactions That Use Artificial Templates with Those Using Natural Messenger RNA
Authors: Crystal Ronald G. , Anderson W. French
Published: May 2006

Partial reactions designed to study the individual steps in the initiation of protein synthesis may be divided into two groups: artificial models using artificial mRNA templates and natural models using naturally occurring mRNA. The requirements for each of the reticulocyte initiation factors, IF-M
1
, IF-M
2
, and IF-M
3
, and for GTP are examined using these models in order to determine if either type of model is a valid representation of the events occurring in natural initiation.

Binding of the initiator tRNA, Met-tRNA
F
, to endogenous mRNA requires IF-M
1
, IF-M
2
, and GTP. A requirement for hydrolysis of GTP is not found when the artificial template ApUpG is used since GDPCP may be substituted for GTP. Met-tRNA
F
bound to the template ApUpG by IF-M
1
+ IF-M
2
can form a peptide bond with the aminoacyl-tRNA analog, puromycin. Met-tRNA
F
bound by IF-M
1
+ IF-M
2
to the initiator codon of natural globin mRNA, however, cannot form a peptide bond with either puromycin or valine-tRNA unless IF-M
3
is also present. The requirements for Met
F
-valine synthesis on exogenous globin mRNA are the same as the requirements on endogenous mRNA. Synthesis of the initial tripeptides of the α and β chains of rabbit globin, Met
F
-Val-Leu and Met
F
-Val-His, requires, in addition, leucyl-tRNA and histidyl-tRNA.

It appears, therefore, that model systems that use natural messenger RNA can duplicate the factor and energy requirements of natural initiation, but that the model systems thus far studied that use artificial messengers as templates do not.
Inhibition of DNA Synthesis in Animal Cells by Ethylene Diamine Tetraacetate, and Its Reversal by Zinc
Authors: Rubin Harry
Published: May 2006

During an investigation of the role of ions in growth regulation, it was found that ethylene diamine tetraacetate (EDTA) inhibits DNA synthesis about 10-fold in cultures of chick embryo cells, while ethylene glycol bis(β-amino ethyl ether)-
N,N
′-tetraacetate has no effect. RNA synthesis is only slightly inhibited by EDTA, and protein synthesis is unaffected. EDTA is inhibitory to DNA synthesis at a concentration much lower than that of either Ca
++
or Mg
++
present in the growth medium. The inhibition is prevented by the addition of Zn
++
at a much lower concentration than that of the EDTA. Other metal ions are ineffective. The inhibition of DNA synthesis only becomes apparent after more than 6 hr of incubation with EDTA, and descends to its final level by 15 hr. Complete restoration of the original rate of DNA synthesis is achieved within 8-10 hr by the addition of Zn
++
. The low rate of DNA synthesis that occurs in a density-inhibited culture, is refractory to further inhibition by EDTA. Rous sarcoma cells are less sensitive to inhibition by EDTA than normal cells, but the sensitivity of both is increased by reducing the concentration of Ca
++
in the medium. DNA synthesis in mouse 3T3 cells is also inhibited by EDTA. It is concluded that Zn
++
is a continuing requirement for DNA synthesis in cultured vertebrate cells, and it is suggested that the availability of Zn
++
within the cell may play a role in the regulation of cell multiplication.
Enhancement by Interferon of the Specific Cytotoxicity of Sensitized Lymphocytes
Authors: Lindahl Pernilla , Leary Patricia , Gresser Ion
Published: May 2006

Mouse interferon preparations enhanced the specific cytotoxicity of sensitized lymphocytes for allogeneic target tumor cells. The factor responsible for the enhancement of cytotoxicity could not be dissociated from the antiviral activity of interferon by standard physicochemical means. Thus, in addition to its well-known antiviral activity, and its effect on cell division, interferon also appears to enhance a specialized cellular function. It is suggested that a common mechanism of action underlies these seemingly different biologic phenomena, and that interferon may play a role in the regulation of fundamental cellular processes.
Electromechanochemical Model of Mitochondrial Structure and Function
Authors: Green David E. , Ji Sungchul
Published: May 2006

A new model of mitochondrial structure and function is proposed on the basis of four fundamental assumptions: (
a
) electrons and protons are separated in the electron transfer complexes; (
b
) the ATPase undergoes conformational state-transitions induced by an electric field; (
c
) energy is transferred by an electric field effect; and (
d
) a conformationally strained protein system can be relaxed via a bond-forming chemical reaction. The model can explain all of the mitochondrial coupled processes and, in addition, it provides a reasonable rationalization of the correlation between mitochondrial structure and function.
New Method to Prepare
N-t
-Butoxycarbonyl Derivatives and the Corresponding Sulfur Analogs from di-
t
-Butyl Dicarbonate or di-
t
-Butyl Dithiol Dicarbonates and Amino Acids
Authors: Tarbell D. Stanley , Yamamoto Yutaka , Pope Barry M.
Published: May 2006

Di-
t
-butyl dicarbonate and one of its dithiol analogs, practical methods of preparation for which are given, react with amino-acid esters to form the
N-t
-butoxycarbonyl (
t
-BOC) derivatives and the thiol analogs in good yield under mild conditions. The thiol analogs are stable to acidic conditions, which rapidly remove the
t
-BOC group itself.
t
-Butyl trimethylsilyl carbonate forms a (CH
3
)
3
Si ether from a
N
-thiol-
t
-BOC serine methyl ester. The
N
-thiol-
t
-BOC group can be removed from the —NHCOSR (R =
t
-butyl) by heating with peroxide-acetic acid.

Action of the dicarbonates described above has not been attended by racemization in the cases examined. The two dicarbonates may be useful as agents for selective blocking and deblocking of amino or other groups.
The Effect of Aldosterone on the Accumulation of Adenosine 3′:5′-Cyclic Monophosphate in Toad Bladder Epithelial Cells in Response to Vasopressin and Theophylline
Authors: Stoff Jeffrey S. , Handler Joseph S. , Orloff Jack
Published: May 2006

Vasopressin and theophylline both increase the content of adenosine 3′:5′-cyclic monophosphate (cAMP) in epithelial cells of the urinary bladder of toads (
Bufo marinus
). Incubation of the tissue with 0.2 μM aldosterone markedly increases this response to the two agents; incubation for a similar time without steroid reduces the response. The permeability responses (sodium transport and water flow) of the intact tissue to vasopressin, theophylline, and exogenous cAMP are also considerably greater in bladders incubated with aldosterone than without. The results are interpreted as indicating that the foregoing permissive effects of aldosterone on the permeability responses to vasopressin and theophylline are mediated by a steroid-dependent increase in the accumulation of cAMP in the pertinent epithelial cells, probably as a consequence of a diminution in the rate of degradation of the intracellular nucleotide.
Effect of Insulin on Protein Synthesis in Skeletal Muscle of an Isolated Perfused Preparation of Rat Hemicorpus
Authors: Jefferson L. S. , Koehler J. O. , Morgan H. E.
Published: May 2006

A method for perfusion
in vitro
of a preparation of rat hemicorpus was developed for study of the metabolism of skeletal muscle. The preparation was stable during perfusion, as indicated by maintenance of ATP concentration, perfusion pressure, and oxygen consumption for up to 90 min. The perfused hemicorpus provided the following advantages for study of protein synthesis in skeletal muscle: (
a
) hormones and substrates reached the muscle cells through an intact capillary bed, and (
b
) the preparation included the psoas muscle, which was sufficiently large to allow measurements of intermediates in the pathway of protein synthesis and was readily homogenized for preparation of ribosomes and ribosomal subunits. Perfusion of psoas muscle from fasted rats with buffer containing glucose and insulin reduced the concentration of ribosomal subunits and increased phenylalanine incorporation as compared to perfusion with buffer containing glucose alone. In addition, the hormone increased glucose uptake from the perfusate and inhibited release of free fatty acids from the preparation. When the muscle was perfused with buffer that contained glucose and palmitate, the concentration of ribosomal subunits and phenylalanine incorporation were unchanged. Since fatty acid is known to stimulate protein synthesis in heart muscle, these results indicated that rates of protein synthesis in heart, but not in skeletal, muscle would be maintained during fasting or in diabetic animals by increased plasma concentration of fatty acid.
Genetic Variants of Glucose 6-Phosphate Dehydrogenase from Human Erythrocytes: Unique Properties of the A
-
Variant Isolated from “Deficient” Cells
Authors: Babalola Olaniyi , Cancedda Ranieri , Luzzatto Lucio
Published: May 2006

The A
-
type of glucose 6-phosphate dehydrogenase (EC 1.1.1.49) has been isolated from human erythrocytes deficient in this enzyme. The specific activity of the purified protein is similar to that previously reported for the enzyme isolated from normal, nondeficient erythrocytes. During the purification procedure, a portion of the A
-
enzyme converts spontaneously, from the native “fraction I”, to a “fraction II” having different kinetic and chromatographic properties. The conversion of fraction I to II can be reproduced freely by treatment with iodosobenzoate, and fraction II can be converted back to fraction I by treatment with dithioglycol. We suggest that fraction II is an enzyme species in which one or more sulfhydryl groups have been oxidized to disulfide(s). The tendency to oxidation appears to be a property specific to the A
-
variant and may represent the basis for its rapid rate of inactivation and consequent deficiency
in vivo
.
Separation of Reticulocyte Initiation Factor M
2
Activity into Two Components
Authors: Shafritz David A. , Prichard Philip M. , Gilbert Jeffrey M. , Merrick William C. , Anderson W. French
Published: May 2006

IF-M
2
, one of three initiation factors isolated by DEAE-cellulose chromatography from the 0.5 M KCl-wash fraction of rabbit reticulocyte ribosomes, has been separated by Sephadex G-200 chromatography into two components: IF-M
2A
and IF-M
2B
. IF-M
2A
elutes near the void-volume, while IF-M
2B
, which is much smaller in molecular weight than IF-M
2A
, elutes slightly after a hemoglobin marker. In the presence of the other appropriate factors, both IF-M
2A
and IF-M
2B
are required to stimulate poly(U)-directed polyphenylalanine synthesis at low Mg
++
concentration, ApUpG-directed Met-tRNA
F
binding to washed reticulocyte ribosomes, and initiation of globin synthesis from endogenous mRNA. IF-M
2A
stimulates ribosome-dependent GTP hydrolysis, while IF-M
2B
does not; IF-M
2B
stimulates ApUpG-directed fMet-tRNA
F
binding in the presence of IF-M
1
, while IF-M
2A
does not. Although IF-M
2A
and IF-M
2B
can be distinguished from each other by size and by activity, a distinct function for IF-M
2B
has not yet been found. Therefore, its precise role in the initiation process remains unclear.
Detection of DNA Synthesis in Intact Organisms with Positron-Emitting [
Methyl
-
11
C]Thymidine
Authors: Christman David , Crawford Elizabeth J. , Friedkin Morris , Wolf Alfred P.
Published: May 2006

11
CO
2
produced in the Brookhaven 152-cm cyclotron was converted to formaldehyde, which in turn was used for the enzymatic conversion of deoxyuridine-5′-phosphate to [
11
C]thymidylate. Enzymatic treatment of the nucleotide with alkaline phosphatase gave [
11
C]thymidine.

The preparation of [
11
C]thymidine from cyclotron-generated
11
CO
2
required 110 min (about 5 half-lives): 35 min for the synthesis of H
11
CHO, 25 min for the enzymatic conversion to [
11
C]thymidylate, 20 min for column chromatography, 5 min for phosphatase treatment, 10 min for evaporation, 2 min for filtration through an anion-exchange resin, and 13 min for miscellaneous manipulations.

Positron-emitting [
11
C]thymidine and [
11
C]thymidylate were used for
in vivo
tracer studies of DNA synthesis in mice for periods of up to 3 hr. Findings with carbon-11 were consistent with earlier studies in which carbon-14 and tritium-labeled thymidine were used. For example, 3 hr after injection of [
11
C]thymidine, spleen DNA was labeled to a much greater extent than was liver DNA.
Initiation and Reinitiation of DNA Synthesis during Replication of Bacteriophage T7
Authors: Dressler David , Wolfson John , Magazin Marilyn
Published: May 2006

In its first round of replication, the T7 chromosome follows a simple pattern, as viewed in the electron microscope. The iniation of DNA synthesis occurs about 17% from the genetic left end of the viral DNA rod. Bidirectional DNA synthesis from this origin then generates a replicating intermediate that we call an “eye form.” In the eye form, when synthesis in the leftward direction reaches the left end of the viral chromosome, the molecule is converted into a Y-shaped replicating rod. The remaining growing point continues synthesis rightward, until presumably it runs off the right end of the DNA rod, thus terminating replication.

Numerous T7 chromosomes were found in which a second round of replication had begun before the first round had finished. Analysis of these reinitiated DNA molecules showed that the second round of replication, like the first, began 17% from the end of the chromosome and involved bidirectional DNA synthesis.
Transport of Magnesium by a Repressible and a Nonrepressible System in
Escherichia coli
Authors: Nelson David L. , Kennedy Eugene P.
Published: May 2006

Wild-type cells of
E. coli
take up Mg rapidly when grown in media with high (10 mM) or with low (50 μM) concentrations of Mg. Previous work has provided evidence that a transport system for Mg in
E. coli
also functions for the uptake of Co. Mutants resistant to Co have now been found to possess a system for Mg uptake that is repressed by high concentrations of Mg in the growth medium, and does not function effectively for the uptake of Co. The rate of Mg uptake by such mutants increases at least 50-fold within one generation after a transfer from growth medium with high Mg to one with low Mg concentrations. We conclude that there are at least two systems for the uptake of Mg in
E. coli
. System I is nonrepressible and can catalyze the uptake of Co as well as Mg; it is present in wild type but not in Co-resistant mutants. System II is repressible and has little affinity for Co; it is present in both wild-type and mutant strains.
Role of DNA Polymerase I and the
rec
System in Excision-Repair in
Escherichia coli
Authors: Cooper Priscilla K. , Hanawalt Philip C.
Published: May 2006

The DNA polymerase I-deficient mutant
polA
1 is shown to perform an
increased
amount of UV-stimulated repair synthesis relative to its
pol
+
parent. In contrast, a
recA recB
double mutant is found to perform little detectable repair synthesis. Analysis of the density distribution of sheared DNA of the
recA recB
mutant reveals that none of the repair synthesis in this strain is in the large repair patches previously demonstrated by us in wild-type strains. These results are interpreted in terms of a model involving both DNA polymerase I and the
rec
system in the excision-repair process, with polymerase I performing an efficient short patch repair and
rec
system enzymes producing predominantly large patches of repair synthesis.
Evidence for Reversible Inactivation of Induced Tyrosine Aminotransferase in Rat Liver
In Vivo
Authors: Grossman Albert , Boctor Amal
Published: May 2006

Induction of tyrosine aminotransferase (EC 2.6.1.5) with cortisol is followed by a rapid phase of inactivation of the enzyme, during which administration of puromycin causes a sharp
increase
in enzyme activity. The increase is not due to puromycin-resistant synthesis of protein, since precipitation of the enzyme with antibodies to tyrosine aminotransferase after pulse labeling inhibited incorporation of radioactive amino acids. Reactivation of the enzyme was specific, since the structurally similar dimethylaminopurine and purine aminonucleoside were ineffective. An action of puromycin other than its capacity to inhibit protein synthesis was required for reactivation, since cycloheximide did not demonstrate such an effect.
In vitro
studies with free and bound polysomes, isolated during the phase of inactivation, indicated that reactivation by puromycin was not due to release of nascent tyrosine aminotransferase peptides. We conclude that the enzyme is initially reversibly inactivated; therefore, the overall degradation of this intracellular protein is a multistep phenomenon.
Interferon Messenger RNA: Translation in Heterologous Cells
Authors: Maeyer-Guignard Jaqueline De , Maeyer Edward De , Montagnier Luc
Published: May 2006

A viral inhibitor with the characteristics of mouse interferon is produced by avian and simian cells preincubated with RNA extracted from interferon-producing mouse cells. Similarly, RNA extracted from interferon-producing monkey cells induces a monkey interferon-like substance in avian cells and also in a line of simian cells, VERO, which normally lacks the capacity to produce its own interferon. In both cases, the RNA effect is inhibited by treatment of the receptor cells by cycloheximide, but not by actinomycin D. We conclude that interferon messenger RNA has been translated in the receptor cells. Thus, the production of interferon in heterologous cells can be used as a sensitive assay of interferon messenger RNA.
Interaction of Eukaryote Elongation Factor EF 1 with Guanosine Nucleotides and Aminoacyl-tRNA
Authors: Moon Hong-Mo , Redfield Betty , Weissbach Herbert
Published: May 2006

Evidence for two species of elongation factor 1 (EF 1
A
and EF 1
B
) from calf brain has been obtained by molecular sieve chromatography on Sephadex G-150. A high molecular weight form, EF 1
A
, interacts with GTP to form an EF 1
A
-GTP complex. GDP also reacts with EF 1, but unlike the reaction with GTP, an EF 1
B
-GDP complex is formed that contains a lower molecular weight and labile species of EF 1. The results also indicate that EF 1
A
-GTP reacts with aminoacyl-tRNA to form an aminoacyl-tRNA-EF 1
B
-GTP complex. These results are discussed with regard to the role of EF 1 in aminoacyl-tRNA binding to ribosomes.
Inhibition of Influenza Virus Replication by α-Amanitin: Mode of Action
Authors: Mahy B. W. J. , Hastie N. D. , Armstrong Sylvia J.
Published: May 2006

The replication of influenza virus in chick embryo fibroblast cells is inhibited by α-amanitin added during the first 2 hr of infection at concentrations similar to those required to inhibit cellular DNA-dependent RNA polymerase form II
in vivo
. Of two periods of increased RNA synthesis observed in cells infected with influenza virus, only the first, occurring from 0 to 2 hr after infection, is sensitive to α-amanitin. During this early period, there is a stimulation of the activity of DNA-dependent RNA polymerase II of nuclei isolated from infected cells. The data suggest that DNA transcription mediated by polymerase II is essential for influenza virus replication.
Conformational Studies on Tocinamide and Deaminotocinamide by 220 MHz Nuclear Magnetic Resonance Spectroscopy
Authors: Brewster Anne I. , Glasel J. A. , Hruby V. J.
Published: May 2006

The 220 MHz spectra of tocinamide and deaminotocinamide, the ring moieties of oxytocin and of deamino-oxytocin, respectively, were investigated in [
U
-
2
H]dimethylsulfoxide solution. Extensive decoupling and exchange experiments allow complete spectral assignment and determination of many coupling constants. Circular dichroism spectra assigned a right-hand screw sense to the C-S-S-C group. The conformations of tocinamide and deaminotocinamide are different from each other and from those suggested for the ring portions of oxytocin and deamino-oxytocin. The relationship of this difference to biological activity is commented upon. The importance of the interaction of the side chain with the ring in oxytocin and deamino-oxytocin is emphasized.
Control of Globin Synthesis in Cell-Free Preparations of Reticulocytes by Formation of a Translational Repressor That is Inactivated by Hemin
Authors: Gross M. , Rabinovitz M.
Published: May 2006

The control of globin synthesis by hemin in cell-free preparations of rabbit reticulocytes is mediated by an inhibitor (translational repressor) of globin chain initiation that is inactivated by hemin. When the ribosome-free supernatant fraction is warmed at 34° without hemin, it quickly acquires the ability to inhibit a fresh cell-free preparation. If the warmed supernatant fraction is further incubated with hemin, its inhibitory activity is lost. This reversible inhibitor, which is observable only during the early period of incubation without hemin, is distinct from an irreversible inhibitor that becomes prevalent during longer incubations. The reversible inhibitor may be an intermediate in the formation of the irreversible inhibitor. The sensitivity of the reversible inhibitor to inactivation by hemin, which permits resumption of globin synthesis in both intact cells and lysates, indicates that the inhibitor is a physiological regulator.
Separation and Translation of the mRNAs Coding for α and β Chains of Rabbit Globin
Authors: Temple Gary F. , Housman David E.
Published: May 2006

A method is presented to separate the mRNAs coding for α and β globins of rabbit reticulocytes. 10S RNA was extracted from the light and heavy polysomes created by incubation of reticulocytes with L-
O
-methylthreonine, and assayed for mRNA activity in an ascites cell-free extract. Tryptic digests of the
in vitro
products demonstrated that the heavy polysomes yielded β globin mRNA at least 90% free of α mRNA activity, and that the light polysomes yielded α mRNA at least 70% free of β mRNA activity.
Adenovirus Messenger RNA in Mammalian Cells: Failure of Polyribosome Association in the Absence of Nuclear Cleavage
Authors: McGuire P. M. , Swart C. , Hodge L. D.
Published: May 2006

The nuclear synthesis of adenovirus-specific RNA late in the infectious cycle in the presence of toyocamycin (an adenosine analogue) has been investigated. There is reduced synthesis of viral RNA with an accumulation of virus-specific RNA in the molecular weight range of at least 4 to 8 × 10
6
. No new viral RNA associates with cytoplasmic polyribosomes. In addition, hybridization competition experiments indicate a 70% competition between these large nuclear transcripts and polyribosome-associated viral RNA that was synthesized in the absence of inhibitor. These data are consistent with the following interpretations: complete nuclear processing of viral RNA is necessary for polyribosome association, and precursor viral message(s) contain sequences that are lost normally during post-transcriptional processing.
Introduction of the 3-Deoxy Group in 3,6-Dideoxyhexoses: A Novel Cofactor Function for Pyridoxamine-5′-Phosphate
Authors: Gonzalez-Porque Pedro , Strominger Jack L.
Published: May 2006

A cofactor needed for the reduction of CDP-4-keto-6-deoxy-D-glucose to CDP-4-keto-3,6-dideoxy D-glucose has been detected and purified. Its spectra in neutral, acid, and basic solutions, its behavior on thin-layer chromatography, and other properties led to its identification as pyridoxamine-5′-phosphate. Moreover, authentic pyridoxamine-5′-phosphate substituted for the purified cofactor with about the same specific activity. Other vitamin B
6
derivatives did not stimulate the reaction, and inhibition was observed with some of them.
Proventriculus of a Marine Annelid: Muscle Preparation with the Longest Recorded Sarcomere
Authors: Castillo José Del , Anderson Margaret , Smith David S.
Published: May 2006

Each muscle of the syllid (Annelida: Polychaeta) proventriculus, the region of the gut posterior to the pharynx, contains a single zigzagging Z band, flanked on each side by a sequence of I-A-H-A-I bands defined by thick (60-90 nm) and thin (5 nm) filaments. The thick filaments show a 14-nm periodicity similar to paramyosin. The muscle cell terminates at the level of the outer I bands. The muscle does not include a complete sarcomere in the strict sense, as defined by a pair of Z bands, but the equivalent H-H distance reaches about 40 μm. Electrophysiological evidence suggests that contraction and relaxation of the cell are, respectively, associated with depolarizing and hyperpolarizing potentials.
Sanfilippo Syndrome: Profound Deficiency of Alpha-Acetylglucosaminidase Activity in Organs and Skin Fibroblasts from Type-B Patients
Authors: O'brien John S.
Published: May 2006

Cultured skin fibroblasts from two patients with Sanfilippo syndrome, Type B were strikingly deficient in α-acetylglucosaminidase activity (α-2-acetamido-2-deoxy-D-glucoside acetamidodeoxyglucohydrolase, EC 3.2.1.X). A similar deficiency was found in frozen organs from two other patients. A partial deficiency of α-acetylglucosaminidase was found in cultured skin fibroblasts from both parents of one patient. Soluble endogenous inhibitors did not account for the enzyme deficiency. Other lysosomal hydrolases were normal or increased in cultured fibroblasts from patients with this disease. No deficiency of α-acetylglucosaminidase is present in other genetic mucopolysaccharidoses, including Sanfilippo Type A.
An Engineering Study of the Peristaltic Drive of Axonal Flow
Authors: Biondi Robert J. , Levy Martin J. , Weiss Paul A.
Published: May 2006

By determination of the rheological flow curve of minute quantities of axoplasm drawn into a microcapillary tube connected to a high vacuum, extruded axoplasm was shown to behave as a pseudoplastic fluid with a viscosity of 10
6
-times that of water and without significant signs of time-dependent thixotropic or viscoelastic properties. A theoretical analysis of peristaltic pumping of such pseudoplastic fluids by sinusoidal surface waves was combined with experimental studies of a mechanical model designed to simulate peristaltic drive. The correlation of the respective data permitted quantitative predictions for the peristaltic mechanism of axonal flow, with speed being a function of peristaltic wave geometry and fluid properties, yielding a theoretical mean value of 0.45 mm/day, i.e., of the same order as that observed in the living nerve fiber.
On the Role of RNA in Gene Amplification
Authors: Brown Ronald D. , Tocchini-Valentini Glauco P.
Published: May 2006

A complex of ribosomal DNA with RNA has been isolated from ovaries of metamorphosing tadpoles of
Xenopus laevis
. The complex is disrupted by treatments that destroy hydrogen bonds, and the sedimentation of the DNA within the complex is sensitive to RNase. We suggest that the RNA-ribosomal DNA complex is an intermediate in the synthesis of amplified ribosomal DNA and that the RNA is a template. In addition, in a preliminary attempt to mimic the amplification process
in vitro
, we have demonstrated the use of this RNA from the complex as a template for DNA synthesis by a DNA polymerase isolated from ovaries of
Xenopus
.
DNA Polymerase of Chicken Embryo: Purification and Properties
Authors: Stavrianopoulos Jannis G. , Karkas John D. , Chargaff Erwin
Published: May 2006

The almost complete purification of the previously reported DNA polymerase of chicken embryo is described; in the final isolation step the enrichment of the enzyme, which comprises about 0.01% of the protein extractable from 10-day-old embryos, is more than 9000-fold. The enzyme is a basic protein (pI = 9.15), of molecular weight about 27,000. It exhibits no nuclease activity, requires Mn
2+
ion for activity, and may contain Zn. It is very sensitive to oxidation.
Polypeptide Chain Initiation in Eukaryotes: Functional Identity of Supernatant Factor from Various Sources
Authors: Zasloff Michael , Ochoa Severo
Published: May 2006

Eukaryotic cells contain polypeptide chain initiation factors that, like the prokaryotic initiation factor IF-2, promote the AUG-dependent binding of fMet-tRNA
f
to the small ribosomal subunit. The bound amino-acyl-tRNA is directly convertible to fMet-puromycin upon addition of 60S subunit. The reaction is sensitive to initiation inhibitors such as aurintricarboxylic acid and edeine but, unlike its prokaryotic counterpart, it does not require GTP. Factors that catalyze the binding and fMet-puromycin reactions with ribosomal subunits from
Artemia salina
embryos are present in postribosomal supernatants of
Artemia
, mouse fibroblasts (L cells), and rat liver, as well as in salt washes of rabbit reticulocyte ribosomes. However, whereas all three supernatant factors, like
Escherichia coli
IF-2, are sensitive to SH-binding reagents, the reticulocyte factor is not. The rat liver and
Artemia
factors function indiscriminately with
Artemia
or rat liver ribosomes, but the
Artemia
factor and
E. coli
initiation factor IF-2 are not interchangeable.
Increased Susceptibility of Cells Treated with Interferon to the Toxicity of Polyriboinosinic·Polyribocytidylic Acid
Authors: Stewart William E. , De Clercq Erik , Billiau Alfons , Desmyter Jan , De Somer Pierre
Published: May 2006

Concentrations of the synthetic polymer polyriboinosinic·polyribocytidylic acid that produced no detectable toxicity in normal L cells produced marked cytotoxicity in L cells treated with interferon. This increase in the susceptibility of cells to the toxicity of the polymer was also observed in human cells and secondary mouse embryo cells treated with homologous interferons before exposure to the polynucleotides. The degree of enhancement of toxicity was dependent on the concentration of interferon to which the cells were exposed. The ratio of antiviral activity induced by interferon to enhancement of toxicity by interferon remained constant through about 1000-fold purification. Various interferon preparations induced by viruses or by polyriboinosinic·polyribocytidylic acid
in vivo
or
in vitro
, and international reference standard interferons all exhibited enhancement of toxicity. Both enhancement of toxicity and antiviral activity were destroyed by trypsin and by incubation at 56° for 1 hr, did not act on heterologous cells, were not sedimented by ultracentrifugation, and were not inactivated by ribonuclease, deoxyribonuclease, irradiation with ultraviolet light, or exposure to a pH of 2.
RNA-Linked Nascent DNA Fragments in
Escherichia coli
Authors: Sugino Akio , Hirose Susumu , Okazaki Reiji
Published: May 2006

Nucleic acid that is extracted from
E. coli
labeled by a brief pulse of [
3
H]dT and depatured by treatment with heat, formamide, or formaldehyde bands in a region with a density higher than that of single-stranded
E. coli
DNA in a Cs
2
SO
4
equilibrium density gradient. If treated with alkali or RNase, it then exhibits the density of single-stranded DNA. These results suggest the presence of a short strand of RNA covalently linked to the nascent DNA. Evidence for the presence of covalently linked RNA-DNA molecules is also obtained by pulse labeling with [
3
H]U. Analyses of nascent nucleic acids from cells pulse labeled for various times, and of the molecules with different sizes, support the hypothesis that the short DNA fragments are formed by extension of even shorter RNA chains, which are synthesized on the parental DNA strands and are removed before ligation of the DNA fragments. The synthesis of the RNA segment of the RNA-DNA molecule is much less sensitive to rifampicin than is the synthesis of bulk RNA.
Polyadenylic Acid at the 3′-Terminus of Poliovirus RNA
Authors: Yogo Yoshiaki , Wimmer Eckard
Published: May 2006

Poliovirus RNA that has been derivatized at the 3′-end with NaIO
4
-NaB
3
H
4
yields, after hydrolysis with alkali or RNase T2, predominantly labeled residues of modified adenosine; no labeled nucleoside derivative is produced by digestion with RNase A or RNase T1. The 3′-terminal bases of the RNA are, therefore,...ApA
OH
. Hydrolyzates of poliovirus [
32
P]RNA, after exhaustive digestion with RNase T1 or RNase A, contain, besides internal oligonucleotides, polynucleotides resistant to further action of ribonucleases T1 and A, respectively; these polynucleotides were isolated by membrane-filter binding or ion-exchange chromatography. The sequence of the T1-resistant polynucleotide was determined to be (Ap)

n

A
OH
, that of the RNase A-resistant polynucleotide was GpGp(Ap)

n

A
OH
. The chain length (
n
) of the polyadenylic acid, as analyzed by different methods, averages 89 nucleotides. Gel electrophoresis revealed heterogeneity of the size of poly(A). Poliovirus RNA, when labeled
in vitro
at the 3′-end, contains [3′-
3
H]poly(A); when labeled
in vivo
with [
3
H]A, it contains [
3
H](Ap)

n

A
OH
. The data establish that... YpGpGp(Ap)
[unk]
A
OH
is the 3′-terminal sequence of poliovirus RNA, Type 1 (Mahoney). Since this mammalian virus reproduces in the cell cytoplasm, these observations may modify prior interpretations of the function of polyadenylate ends on messenger RNAs.
In Vitro
Photoreactions of Chlorophyll and Photosynthetic Energy Conversion: Chlorophyll-Quinone-Ethanol at Low Temperature as an Analog of Photosystems I and II
Authors: Harbour John R. , Tollin Gordon
Published: May 2006

Illumination at low temperatures of ethanol solutions of
p
-benzoquinone, alone or in the presence of chlorophyll or pheophytin, leads to the reversible formation of radical complexes. Based on
g
-value determinations and deuterium-substitution effects, the radicals are identified as a semiquinone and an ethoxy radical. In the porphyrin-sensitized system, ternary complexes are produced. Chlorophyll also forms a cation radical upon illumination, whereas pheophytin does not. The generation of this cation radical is independent of the presence of quinone, and proceeds via the chlorophyll singlet state, as opposed to quinone and ethoxy radical production, which involve triplet states. The significance of these reactions to photosynthetic energy conversion is discussed.
Partial Restoration of Normal Functional Properties in Carboxypeptidase A-Digested Hemoglobin
Authors: Bonaventura Joseph , Bonaventura Celia , Giardina Bruno , Antonini Eraldo , Brunori Maurizio , Wyman Jeffries
Published: May 2006

In the absence of organic phosphates human hemoglobin A digested with carboxypeptidase A (des His, Tyr β) has high ligand affinity, a greatly reduced Bohr effect, and no heme-heme interaction. Under these conditions, it shows the simple, homogeneous ligand-binding kinetics characteristic of noncooperative heme proteins in which the high combination velocity for both O
2
and CO accounts, to a larger extent, for the increased affinity for both these ligands.

Addition of inositol hexaphosphate dramatically alters the functional properties of this digested hemoglobin. The Bohr effect is greatly increased, and at neutral pH the protein shows significant, though still reduced, heme-heme interaction, together with a 5-fold decrease in affinity. In the presence of saturating amounts of the organic phosphate, the value of
n
is pH dependent, dropping from 1.9 at pH 5.8 to 1.3 at pH 8.6. After inositol hexaphosphate addition, the combination of the deoxy form of the digested hemoglobin with CO is 10-times slower than that observed in the absence of the inorganic phosphate; also the combination with CO after flash photolysis is biphasic and is similar, in many respects, to that observed for unmodified hemoglobin. Besides these functional changes, addition of inositol hexaphosphate to the modified deoxyhemoglobin results in an increase in the extinction coefficient at 430 nm similar to that observed on mixing the isolated α and β chains of normal hemoglobin. The results are consistent with the idea that inositol hexaphosphate shifts an equilibrium between high- and low-affinity forms of the protein.
Sites of Replication of Chromosomal DNA in a Eukaryotic Cell
Authors: Fakan Stanislav , Turner Gail N. , Pagano Joseph S. , Hancock Ronald
Published: May 2006

In mouse cells (line P815), newly synthesized DNA labeled for 20-30 sec during exponential growth is found by electron microscope autoradiography at sites throughout the cell nucleus. These sites are relatively more concentrated in the peripheral region of the nucleus (averaged over a random population of S-phase cells), probably reflecting a higher local concentration of DNA in this region. Newly synthesized DNA is not preferentially associated with purified nuclear envelopes, but is found in a fraction of the chromosomal deoxynucleoprotein whose buoyant density in CsCl after formaldehyde treatment is about 1% lower than that of the deoxynucleoprotein peak. Kinetics experiments suggest that this material is a precursor of mature deoxynucleoprotein; it may represent regions of deoxynucleoprotein containing replicating DNA and the additional proteins involved in DNA replication. Other complexes of newly replicated DNA that are found in the interphase after phenol extraction of nuclei are formed during the extraction procedure, probably due to the partially single-stranded nature of replicating DNA, and do not appear to exist
in vivo
.
Newly Recognized Vital Nitrogen Cycle
Authors: Johnston Harold
Published: May 2006

Soil and sea bacteria produce a small amount of nitrous oxide (N
2
O); a small part of this N
2
O is photochemically converted to nitric oxide (NO) in the stratosphere. This process has recently been shown to be the principal source of the active oxides of nitrogen (NO and NO
2
) in the stratosphere. The active oxides of nitrogen catalytically destroy ozone, and NO and NO
2
appear to be a principal factor in the natural ozone balance. Stratospheric ozone is the only effective shield for the surface of the Earth against the harsh ultraviolet radiation between 300 and 250 nm. Thus, soil bacteria indirectly control the intensity of ultraviolet radiation reaching the Earth's surface. This subcycle of the major nitrogen cycle involves a relatively small amount of nitric oxide, estimated to be between 0.26 and 1.2 × 10
9
kg of NO per year on a worldwide basis. A recent estimate of the future nitric oxide emission in the stratosphere by the Concord supersonic transport is 0.37 × 10
9
kg/year on a world-wide basis, which is similar to the amount estimated from the natural source.
An Acceptor-Dependent Polyglycerolphosphate Polymerase
Authors: Mauck John , Glaser Luis
Published: May 2006

A method for the extraction of polyglycerolphosphate polymerase from
Bacillus subtilis
membranes is described. Further purification by ion-exchange chromatography yields an enzyme fraction totally dependent on the addition of a heat-stable factor for activity. This factor was isolated from cell membranes and acts as an acceptor of glycerolphosphate units. The acceptor contains glycerolphosphate, glucosamine, and fatty acids, but its precise chemical structure has not yet been elucidated.
Purification by Affinity Chromatography of the Molecular Forms of Acetylcholinesterase Present in Fresh Electric-Organ Tissue of Electric Eel
Authors: Dudai Yadin , Silman Israel , Shinitzky Meir , Blumberg Shmaryahu
Published: May 2006

An acetylcholinesterase inhibitor-Sepharose conjugate was prepared by coupling a derivative of the powerful acetylcholinesterase inhibitor,
N
-methylacridinium, to CNBr-activated Sepharose. Use of this conjugate permitted direct purification, by affinity chromatography, of the two molecular forms of acetylcholinesterase, 14 and 18 S, present in fresh electric organ tissue. The purified 14S and 18S acetylcholinesterases retained the capacity to aggregate at low ionic strength displayed by crude extracts of the enzyme. The major polypeptide components of the 14S and 18S enzymes, as revealed by acrylamide gel electrophoresis, closely resemble those observed in the 11S form of acetylcholinesterase, previously purified after tryptic digestion of electric-organ tissue.
Is the Gene the Unit of Selection? Evidence from Two Experimental Plant Populations
Authors: Clegg M. T. , Allard R. W. , Kahler A. L.
Published: May 2006

The dynamics of gametic frequency change have been analyzed in two experimental plant populations. Individual plants were scored for their genotype at four enzyme loci, and four-locus joint gametic frequencies were estimated from the genotypic data over generations. Striking correlations developed in allelic state over loci, including correlations between nonlinked loci, as these populations evolved. Furthermore, the same pair of four-locus complementary gametic types came into marked excess in the late generations of both populations. The results demonstrate that natural selection acted to structure the genetic resources of these populations into sets of highly interacting, coadapted gene complexes. They also provide evidence, at the level of the gene, that selection operating on correlated multilocus units is an important determinant of population structure.
Identification of an Antigen from Normal Human Tissue That Crossreacts with the Carcinoembryonic Antigen
Authors: Kleist S. Von , Chavanel G. , Burtin P.
Published: May 2006

A glycoprotein present in normal human tissue is characterized that is neither organ- nor tumor-specific (nonspecific crossreacting antigen) and that crossreacts (by the Ouchterlony double-diffusion technique) with the carcinoembryonic antigen. This immunological relationship indicates common determinants on the molecules of both antigens. We demonstrate that the nonspecific crossreacting antigen is not a fragment of the carcinoembryonic antigen molecule.
Decarboxylation of 3,4-Dihydroxyphenylalanine (DOPA) by Erythrocytes: A Reaction Promoted by Methemoglobin and Other Ferriheme Proteins
Authors: Tate Suresh S. , Orlando Joseph , Meister Alton
Published: May 2006

The decarboxylation of DOPA by erythrocyte hemolysates differs from DOPA decarboxylation catalyzed by aromatic aminoacid decarboxylases that contain vitamin B
6
in several significant respects. The ability of erythrocyte hemolysates to decarboxylate DOPA is associated with interaction between DOPA and methemoglobin; the ferriheme protein is reduced and DOPA is decarboxylated, probably after oxidation to a quinone intermediate. An analogous reaction takes place between DOPA and other ferriheme proteins, such as metmyoglobin and cytochrome
c
. This phenomenon may be of significance in relation to the side effects observed in patients with Parkinson's disease who are treated with very large doses of DOPA.
Isolation of Supercoiled Colicinogenic Factor E
1
DNA Sensitive to Ribonuclease and Alkali
Authors: Blair D. G. , Sherratt D. J. , Clewell D. B. , Helinski D. R.
Published: May 2006

The synthesis of the covalently-closed, circular DNA form of colicinogenic factor E
1
(
Col
E
1
) continues in
Escherichia coli
cells after the addition of chloramphenicol. A large portion of the purified supercoiled
Col
E
1
DNA molecules made in the presence of chloramphenicol are converted to the open circular DNA form after treatment with alkali (pH 13), RNase A, or RNase H. These treatments do not significantly affect the covalently-closed form of
Col
E
1
DNA isolated from normally growing
E. coli
cells. The open circular product resulting from treatment of supercoiled
Col
E
1
DNA with RNase A possesses a single break in one strand of the circular duplex. The site sensitive to RNase A occurs with equal probability in either of the complementary strands. Both synthesis of
Col
E
1
DNA and the formation of supercoiled
ColE
1
DNA sensitive to RNase A or alkali are prevented by the inhibitor of RNA synthesis, rifampicin. These results indicate that covalently-closed
Col
E
1
DNA containing one or more ribonucleotides accumulates during
Col
E
1
replication in the presence of chloramphenicol. It is proposed that this incorporated RNA served as a primer during the initiation of synthesis of
Col
E
1
DNA and that its removal from the circular DNA is inhibited in cells incubated in the presence of chloramphenicol.
Translation of Reovirus Messenger RNAs Synthesized
In Vitro
into Reovirus Polypeptides by Several Mammalian Cell-Free Extracts
Authors: McDowell Maxson J. , Joklik Wolfgang K. , Villa-Komaroff Lydia , Lodish Harvey F.
Published: May 2006

Single-stranded reovirus RNA, synthesized
in vitro
by reovirus cores, functioned as messenger RNA in cell-free extracts prepared from several mammalian cells: Krebs II mouse ascites cells, mouse L cells, Chinese hamster ovary cells, HeLa cells, and rabbit reticulocytes. As shown by acrylamide gel electrophoresis, all eight polypeptides known to be specified by reovirus were synthesized in the reticulocyte system. In the other extracts, from 5 to 7 complete virus proteins were made.