Resistance of Tetrahymena to ricin, a toxic enzyme from Ricinus communis.

The protozoan $\text{Tetrahymena}\text{pyriformis}$ was found to be resistant to the toxic action of ricin $\text{in}\text{vivo}$. Isolated $\text{Tetrahymena}$ ribosomes were strongly resistant to the A subunit of ricin when tested in a cell free protein synthesis system under different conditions and also lacked the ability to bind A chain stoichometrically. This suggests that $\text{Tetrahymena}$ is resistant $\text{in}\text{vivo}$ because it contains a ribosome which is not susceptible to the toxic action of ricin.     

Evidence for plastoquinol-cytochrome f/b-563 reductase as a common electron donor to P700 and cytochrome oxidase in cyanobacteria

Membranes isolated from $\text{Nostoc}$ sp. strain MAC and $\text{Anacystis}\text{nidulans}$ displayed spectral changes in the cytochrome $\text{f}\text{b}$ region when examined by reduced $\text{minus}$ oxidized or dual wavelength spectrophotometry under physiological conditions. The same changes accompanied both light-induced (photosynthetic) and oxygen-induced (respiratory) electron transport. Physiological reduction of the cytochrome $\text{f}\text{b}$ moiety was abolished after extraction of plastoquinone but reappeared on reconstitution of the depleted membranes with authentic plastoquinone. Moreover, a mutual inhibition of photosynthetic and respiratory activities could be directly demonstrated with the isolated membranes. From the results it is concluded that the membrane-bound plastoquinol-cytochrome $\text{f}\text{b}$ reductase functions as a common electron donor to both P700 and the cytochrome oxidase in cyanobacteria.     

In vitro synthesis of adrenodoxin and adrenodoxin reductase: Existence of a putative large precursor form of adrenodoxin

Cytoplasmic free and bound polysomes were isolated from bovine adrenal cortex, and used to program $\text{in}\text{vitro}$ protein synthesis in rat liver cell sap and wheat germ lysate systems. Synthesis of adrenodoxin(Ad) and adrenodoxin reductase(AdR) in the cell-free systems was determined by immunoprecipitation using monospecific antibodies, and the sizes of the $\text{in}\text{vitro}$ products were analyzed by SDS-polyacrylamide gel electrophoresis. Ad was synthesized by both free and bound polysomes as a putative large precursor having molecular weight of approximately 20,000 daltons, which was processed to mature size Ad (MW 12,000 daltons) by $\text{in}\text{vitro}$ incubation with adrenal cortex mitochondria. On the other hand, AdR was synthesized only by free polysomes apparently as the mature size product.     

Isolation of a complementing activity for a dna-B mutant

A cell free extract which displays temperature sensitive DNA synthesis in the presence of single strand DNA and ATP was prepared from a $\text{dna-B}$ mutant. Following an activity which would reverse this temperature sensitivity, a protein fraction was isolated. The absence of this fraction in a $\text{dna-B}$ mutant indicates that this protein corresponds to the $\text{Dna-B}$ product.     

Peptidyl transferase substrate activity and inhibition of protein biosynthesis by a hydrophilic-aminoacyl analogue of puromycin.

A puromycin analogue possessing a hydrophilic amino acid, 3′-N-[S-(6-hydroxyhexyl)-$\text{L}$-cysteinyl]puromycin aminonucleoside, has been prepared and examined as a substrate for ribosomal peptidyl transferase. Kinetic studies indicate that this non-aromatic aminoacyl analogue is 95.6% as efficient as the parent antibiotic in the transpeptidation reaction. In addition, the analogue is an effective inhibitor of poly (U) and poly (U,C) directed protein synthesis in an $\text{Escherichia coli}$ cell free system.     

Preparation and biological activity of taxol acetates

Synthesis and biological activity of 2′-acetyltaxol and 7-acetyltaxol are reported. Activity is measured $\text{in}\text{vivo}$ by cytotoxicity toward the macrophage-like cell line J774.2, and $\text{in}\text{vitro}$ by promotion of microtubule assembly in the absence of exogenous GTP. Addition of an acetyl moiety at C-2′ results in loss of $\text{in}\text{vitro}$ activity but not cytotoxicity. The properties of 7-acetyltaxol are similar to those of taxol in its effects on cell replication and on $\text{in}\text{vitro}$ microtubule polymerization. Therefore a free hydroxyl group at C-7 is not required for $\text{in}\text{vitro}$ activity and this position is available for structural modifications.     

Photosynthetic control and photophosphorylation in photosystem II of isolated spinach chloroplasts

Two cycles of photosynthetic control have been observed in isolated spinach chloroplasts in the presence of lipophilic class III electron acceptors, which may accept electrons at PS II. $\text{ADP}\text{O}$ ratios of 0.8 to 0.9 were recorded;rates of oxygen evolution were stimulated by phosphorylating reagents and uncouplers. Addition of the plastoquinone antagonist DBMIB decreased photosynthetic control, oxygen evolution and photophosphorylation. We believe that there is a coupling site associated with PSII which can be rate limiting. Comparison of the $\text{P}\text{2e}$ ratios observed with class I and class III electron acceptors leads us to propose that more than 0.6 and possibly approaching one molecule of ATP can be formed for every pair of electrons transported from water to PSII acceptors.     

Glycosylation of free trans-trans abscisic acid as a contributing factor in bud dormancy break

By means of gas-liquid chromatography determination it was found that progress of dormancy break of almond buds is a function of relative proportions of free and glycoside-bound abscisic acid. Successive stages of bud break manifest a marked increase of bound abscisic acid accompanied by a parallel decrease in endogenous levels of the free form. Of the two stereoisomers involved it was found that while the $\text{cis-trans}$ form maintains a more or less stable level throughout, changes were detected primarily in the $\text{trans-trans}$ form. It is thus postulated that the binding of free hormone as well as its total content are of major physiological importance in the process of bud dormancy break.     

The occurrence of glutamate synthase in algae.

The presence of glutamate synthase in the green algae $\text{Chlorella fusca}$ var. $\text{vacuolata}$ has been demonstrated using a whole cell assay as well as cell free extracts. The assay is complicated by the presence of glutamine (amino): α-oxoglutarate transaminase, but this enzyme can be inhibited by amino oxyacetate. The rates of glutamate synthase activity are sufficient to account for the known rates of nitrate assimilation to occur via the glutamine synthetase/glutamate synthase pathway.     

Difference in microviscosity induced by different cholesterol levels in the surface membrane lipid layer of normal lymphocytes and malignant lymphoma cells

Microviscosity ($\text{}\text{?}$gh) in the surface membrane lipid layer of normal lymphocytes and malignant lymphoma cells, and in liposomes prepared from their lipid extracts, was determined with the aid of the fluorescence polarization properties of 1,6-diphenyl 1,3,5-hextriene embedded in it. The $\text{}\text{?}$gh values, both in intact cells and in the liposomes, are distinctively greater for normal lymphocytes than for the lymphoma cells, whereas the fusion activation energy in both types of cells and liposomes is 8 ± 0.5 kcal/mol. Determination of cholesterol revealed that its relative amount in a lymphoma cell is about half of that of a normal lymphocyte, a difference that may account for the above difference in fluidity. This thesis is supported by the observed changes in $\text{}\text{?}$gh, which follow artificial changes in cholesterol contents in the surface membrane of both cell types. Introduction of exogeneous cholesterol into the cell surface membranes was performed with lecithin-cholesterol (1:1) liposomes, and in lymphoma cells resulted in an increase of $\text{}\text{?}$gh to a level of normal lymphocytes. Extraction of native cholesterol from the cell surface membranes was carried out with lecithin liposomes, and in normal lymphocytes results in a decrease of $\text{}\text{?}$gh to a value similar to that of lymphoma cells. The induced changes in cholesterol contents are practically reversible for both cell types. By virtue of controlling the microviscosity of lipid layers, the level of cholesterol in cell surface membranes may play an important role in determining biological activities of normal and malignant cells.     

Synthesis of carnitine from ε-N-trimethyllysine in post mitochondrial fractions of Neurospora crassa

An enzyme system in the post mitochondrial fraction of $\text{Neurospora}$$\text{crassa}$ when supplemented with appropriate cofactors formed carnitine from ε-N-trimethyllysine. These findings together with previous studies of ε-N-lysine methylation in this fungi, illustrate that carnitine synthesis in $\text{Neurospora}$ differs markedly in certain features from mammalian systems in that the entire synthesis is carried out employing free intermediates and cytosolic enzymes.     

Triplet states in photosynthesis

A comparison of zero field splitting (ZFS) and spin polarization of triplet spectra of bacteriochlorophyll $\text{a in vitro}$ and $\text{in vivo}$ provides support for the special pair model of photoreactive chlorophyll in photosynthetic bacteria. Spin polarization of the triplet spectra is a new and unique probe of primary events in the light conversion act in photosynthesis.     

Effect of N-ethylmaleimide on leucine transport in the chang liver cell. II. Effect on the kinetics of Na+-independent transport

The Na⁺-independent leucine transport system is resolved into two components by their different affinity ($\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ about 44 μM and 8.0 mM) for leucine in the Chang liver cell. Treatment of the cells with $\text{N-}\text{ethylmaleimide}$ (1 mM) specifically stimulates the high-affinity component of the Na⁺-independent system by greatly increasing its $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ value, whereas the $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ value of the low-affinity component is markedly lowered. The stimulatory effect of $\text{N-}\text{ethylmaleimide}$ on leucine transport is reduced by prior treatment of the cells with 2,4-dinitrophenol, but this phenomenon seems to be irrelevant to the ATP-depleting action of the uncoupler. The treatment with 2,4-dinitrophenol has been found not to be inhibitory on the subsequent Na⁺-independent leucine uptake itself. Treatment with dibucaine, a phospholipid-interacting drug, also reduces to varying degrees (depending on its concentration) the stimulatory effect of $\text{N-}\text{ethylmaleimide}$ on the subsequent leucine uptake, although pretreatment with dibucaine can stimulate the Na⁺-independent leucine uptake itself. We conclude that the stimulatory effect of $\text{N-}\text{ethylmaleimide}$ on leucine transport is not correlated with the energy level of cell, but involves the perturbation of the membrane bilayer structures.     

Atomic coordinates for the chlorophyll core of a bacteriochlorophyll a-protein from green photosynthetic bacteria

Preliminary atomic coordinates are presented for the seven bacteriochlorophylls constituting the core of one subunit of the trimeric water soluble bacteriochlorophyll $\text{a}$-protein from the green photosynthetic bacterium $\text{Prosthecochloris aestuarii}$, strain 2K, formerly identified as $\text{Chlorobium limicola}$, strain 2K. The coordinates were derived from a 2.8 Å resolution electron density map based on four isomorphous heavy-atom derivatives, and adjusted to have stereochemically acceptable bond lengths and angles.     

Cysteine as a system-specific substrate for transport system ASC in rat hepatocytes.

The rapid transport of $\text{L}$-cysteine into isolated rat hepatocytes escapes detectable inhibition by 2-(methylamino)-isobutyric acid at levels up to 50 mM. The system transporting cysteine instead is convincingly similar to the $\text{ASC}$ system described for the Ehrlich cell in structural and steric specificity and in pH sensitivity. The Na⁺-dependent uptake of 2-aminoisobutyric acid is almost evenly divided between Systems $\text{A}$ and $\text{ASC}$, showing better accommodation of its two α-methyl groups by $\text{ASC}$ than in the Ehrlich cell. The hepatocyte $\text{ASC}$ system tolerates Li⁺-for-Na⁺ substitution better than does System $\text{A}$, although the tolerance depends on amino acid structure. Adaptive regulation and insulin and glucagon stimulation were not seen under conditions producing these effects for System $\text{A}$.     

The accumulation of glycinamide ribotide by ade3 and ade8 mutants of Saccharomyces cerevisiae

The purine intermediate GAR is present in cell free extracts of $\text{ade3}$ and $\text{ade8}$ mutants of yeast. It is also detectable following acid hydrolysis of extracts of $\text{ade6}$ and $\text{ade7}$ which accumulate FGAR and FGAM respectively. GAR accumulation is repressed by growing cells in high levels of adenine. Neither $\text{ade4}$ nor $\text{ade5}$ accumulate GAR and both prevent accumulation of GAR in $\text{ade3}$ and $\text{ade8}$ and FGAR in $\text{ade6}$. Since $\text{ade3}$ is known to be defective in folate metabolism these results indicate that $\text{ade8}$ is blocked in the conversion of GAR→FGAR.     

Isolation and properties of the cytochrome B-C1 complex from Rhodopseudomonas sphaeroides

A highly purified and active cytochrome $\text{b}$-$\text{c}$₁ complex has been isolated from the chromatophores of the photosynthetic bacteria $\text{Rhodopseudomonas}\text{sphaeroides}$ R-26, through steps of Triton X-100 solubilization, salt fractionation and calcium phosphate column chromatography. The isolated enzyme complex catalyzes fully antimycin A sensitive oxidation of ubiquinol by cytochrome $\text{c}$ with a turnover number of 1500 per minute at 23° based on cytochrome $\text{c}$₁. It contains 8.3 nmoles of cytochromes $\text{b}$ and $\text{c}$₁ per mg protein and shows four polypeptides in the sodium dodecylsulfate polyacrylamide gel electrophoresis.     

The biosynthesis of lubimin from [1-14C]isopentenyl pyrophosphate by cell-free extracts of potato tuber tissue inoculated with an elicitor preparation from Phytophthora infestans

A cell-free enzyme system, which catalyses the incorporation of radiolabel from [$\text{1}\text{2}$-¹⁴C]isopentenyl pyrophosphate into the sesquiterpenoid phytoalexin lubimin, has been prepared from tuber tissue of Solanum tuberosum inoculated with an elicitor preparation from Phytophthora infestans. Biosynthesis of lubimin is optimum at pH 7.$\text{3}\text{2}$-7.5 and is dependent upon Mg²⁺ and NADPH. Lubimin labelling by cell-free enzyme system prepared from tissue 48 hr after treatment with elicitor rises rapidly to a maximum over the first 30 min of incubation and does not decline for a further 150 min. The biosynthetic capacity for lubimin in cell free extracts can be observed as early as 3 hr after inoculation of tuber tissue, and rises to a maximum at about 48 hr after treatment, declining thereafter. Lubimin labelling is inhibited by iodoacetamide, the effect of which is reversed by 3,3-dimethylallylpyrophosphate. Preliminary observations on the cell-free system show that it will also catalyse the incorporation of [2-¹⁴C]mevalonic acid into lubimin in the presence ofan ATP-generating system.     

Cyclic 3',5'-adenosine monophosphate stimulates trehalose degradation in baker's yeast

The levels of cyclic 3′,5′-AMP and trehalose, as well as the specific activity of the trehalase have been investigated in cells of baker's yeast ($\text{Saccharomyces cerevisiae}$) during the lag phase preceding growth. During the first few minutes a substantial increase in the intracellular concentration of cyclic 3′,5′-AMP was observed, followed by a 6–8 fold increase in trehalase activity concomitant with the rapid degradation of trehalose. Cell free extracts prepared from resting yeast were shown to contain a cryptic trehalase, which under physiological conditions could be activated by cyclic 3′,5′-AMP to the same degree as $\text{in vivo}$. These observations suggest that in the lag phase of growth, the level of trehalose in baker's yeast is under control of a system, regulated by the level of cyclic 3′,5′-AMP.     

Kinetics of in vivo binding of antagonist to muscarinic cholinergic receptor in the human heart studied by positron emission tomography

Positron Emission Tomography (PET) was used to analyse $\text{in vivo}$ antagonist binding to human myocardial muscarinic cholinergic receptor. The methiodide salt of the muscarinic antagonist, quinuclidinyl benzilate (MQNB), was labeled with the positron emitter, Carbon-11, and injected intravenously to 8 normal subjects. ¹¹C-MONB concentration was determined $\text{in vivo}$ in the ventricular septum from 40 cross-sectional images acquired at the same transverse level over a period of 70 minutes. In 4 subjects, various amounts of unlabeled atropine were rapidly injected at 20 minutes to study whether atropine competitively inhibited MQNB.The kinetics of binding of ¹¹C-MQNB were not the same $\text{in vivo}$ and $\text{in vitro}$. The apparent dissociation rate of ¹¹C-MQNB $\text{in vivo}$ was much slower (by 1 to 2 orders of magnitude) than that observed $\text{in vitro}$ with ³H-QNB. After atropine injection, ¹¹C-MNQB dissociated from its binding sites at a rate that apparently depended on the amount of atropine present. ¹¹C-MQNB kinetics were analysed with a mathematical model which assumes the existence of a boundary layer containing free ligand in the vicinity of the binding sites. The dissociation rate of the radioligand depends on the probability of its rebinding to a free receptor site.