Effect of 3-isobutyl-1-methylxanthine on HCO3− transport in turtle bladder: Evidence for electrogenic HCO3− secretion

Ouabain-treated turtle bladders bathed on both surfaces by identical HCO₃^?/CO₂-containing, Cl^?-free Na⁺ media exhibit a short-circuit current (I_sc) and transepithelial potential (p.d.) serosa electronegative to mucosa. Addition of 3-isobutyl-1-methylxanthine (IBMX), an inhibitor of cyclic nucleotide phosphodiesterase, rapidly reverses the direction of the I_sc and p.d.. The IBMX-induced reversal of I_sc and p.d. is (1) dependent on the presence of HCO₃^? (and CO₂) in the serosal bathing fluid, (2) independent of Na⁺ and other ions in the bathing medium, (3) decreased by inhibitors of carbonic anhydrase or oxidative metabolism, (4) increased by the serosal addition of cyclic AMP or the disulfonic stilbene, SITS. The results constitute evidence that the reversed I_sc elicited by IBMX represents electrogenic secretion of HCO₃^?.     

Moniezia expansa: Subcellular distribution,kinetic properties of acetylcholinesterase and effects of inhibitors and anthelmintics

Acetylcholinesterase (EC 3:1:1:7) has been demonstrated biochemically within partially purified whole worm homogenates of Moniezia expansa. Linear activity occurred with temperature, enzyme concentration, and time. The pH optimum was 8.5 and the Michaelis constant 2.8 mM with inhibition by excess substrate. Inhibitor and specific substrate studies indicated that butyrylcholinesterase was probably absent. The molecular weight of AChE was in excess of 300,000. Greatest activity occurred in the 22,000 and 100,000g particulate fractions. Ultrastructural staining showed that activity was restricted to the ribosomes and cisternae of the rough endoplasmic reticulum. Quinacrine hydrochloride caused 48% inhibition of AChE at 10^?3M and haloxon (di(2-chloroethyl)-3-chloro-4-methyl-7-coumarinyl phosphate) caused 97% inhibition at 10^?4M. No appreciable inhibition (< 25%) occurred with 10^?4M bunamidine hydroxynaphthoate, bephenium hydroxynaphthoate, pyrantel tartrate, p-toluoyl phenyl hydrazone, dichlorphen, thiabendazole, mebendazole, fenbendazole, cambendazole, albendazole, parbendazole, oxibendazole, oxfendazole, praziquantel, piperazine adipate, arecoline hydrobromide, and sodium acetarsol.     

UV-Vis and fluorimetric Al, Zn, Cd and Pb complexation studies of two 3-hydroxyflavones and a 3-hydroxythioflavone

The complexation of Al³⁺, Zn²⁺, Cd²⁺ and Pb²⁺ by the 3-hydroxyflavones: 3-hydroxy-2-(2-methoxyphenyl)-4H-1-benzopyran-4-one (H1) and 3-hydroxy-2-(4-methoxyphenyl)-4H-1-benzopyran-4-one (H2), and by the 3-methoxythioflavone: 3-hydroxy-2-(2-methoxyphenyl)-4H-1-benzopyran-4-thione (H3) have been studied spectrophotometrically and fluorimetrically to determine the corresponding complexation constants, K_sp and K_fl, in 5:95 water:ethanol (v/v) solution for which [HClO₄] was either 10⁻² or 10⁻⁵ mol dm⁻³ and I = 0.10 mol dm⁻³ (NaClO₄) at 298.2 K. Complexation occurs dominantly through the deprotonated ligand for [Al(1)]²⁺ and [Al(2)]²⁺ for which log K_sp = 4.51 and 4.73, respectively, in 10⁻² mol dm⁻³ HClO₄ and 4.21 and 4.61 in 10⁻⁵ mol dm⁻³ HClO₄. For Pb²⁺ complexation by H1, H2 and H3 is characterized by log K_sp = 2.20, 2.57 and 3.22, respectively, in 10⁻² mol dm⁻³ HClO₄ and 4.70, 5.38 and 5.74 in 10⁻⁵ mol dm⁻³ HClO₄. Equilibrium mixtures of [Pb(H1)]²⁺ and [Pb1]⁺, [Pb(H2)]²⁺ and [Pb2]⁺, and [Pb(H3)]²⁺ and [Pb3]⁺ appear to be formed. Complexation of Zn²⁺ and Cd²⁺ by all three ligands was only detected in 10⁻⁵ mol dm⁻³ HClO₄. For Zn²⁺ complexation by H1, H2 and H3 log K_sp = 3.22, 3.74 and 4.46 and for Cd²⁺ the corresponding values are 2.39, 2.40 and 3.72 for Cd²⁺. Only [Al1]²⁺ and [Al2]²⁺ show significant fluorescence and are characterized by log K_fl = 6.30 and 7.49 in 10⁻² mol dm⁻³ HClO₄.     

Inhibition of human initiator caspase 8 and effector caspase 3 by cross-class inhibitory bovSERPINA3-1 and A3-3

Serpins are a superfamily of structurally conserved proteins. Inhibitory serpins use a suicide substrate-like mechanism. Some are able to inhibit cysteine proteases in cross-class inhibition. Here, we demonstrate for the first time the strong inhibition of initiator and effector caspases 3 and 8 by two purified bovine SERPINA3s. SERPINA 3-1 (uniprotkb:Q9TTE1) binds tighly to human CASP3 (uniprotkb:P42574) and CASP8 (uniprotkb:Q14790) with k_ass of 4.2 × 10⁵ and 1.4 × 10⁶ M⁻¹ s⁻¹, respectively. A wholly similar inhibition of human CASP3 and CASP8 by SERPINA3-3 (uniprotkb:Q3ZEJ6) was also observed with k_ass of 1.5 × 10⁵ and 2.7 × 10⁶ M⁻¹ s⁻¹, respectively and form SDS-stable complexes with both caspases. By site-directed mutagenesis of bovSERPINA3-3, we identified Asp³⁷¹ as the potential P1 residue for caspases. The ability of other members of this family to inhibit trypsin and caspases was analysed and discussed.

Structured summary

MINT-7234656: CASP8 (uniprotkb:Q14790) and SERPINA3-1 (uniprotkb:Q9TTE1) bind (MI:0407) by biochemical (MI:0401)MINT-7234634: SERPINA3-3 (uniprotkb:Q3ZEJ6) and CASP3 (uniprotkb:P42574) bind (MI:0407) by biochemical (MI:0401)MINT-7234663: CASP8 (uniprotkb:Q14790) and SERPINA3-3 (uniprotkb:Q3ZEJ6) bind (MI:0407) by biochemical (MI:0401)MINT-7234625: SERPINA3-1 (uniprotkb:Q9TTE1) and CASP3 (uniprotkb:P42574) bind (MI:0407) by biochemical (MI:0401)     

Zinc as a co-factor for an enzyme involved in exsheathment of Haemonchus contortus

The activity of concentrated exsheathing fluid of Haemonchus contortus against isolated sheaths was not inhibited by ethylenediamine tetra-acetic acid (EDTA), 10^?2 M, even when the concentrations of Mg and Mn were < 4 × 10^?4 M and < 0·9 × 10^?6 M respectively. Purified or diluted solutions of exsheathing fluid, even in the presence of Mg²⁺, 10^?3 M, were inhibited. Leucine aminopeptidase (LAP) in exsheathing fluid was active even at concentrations of Mg < 1·3 × 10^?5M. Concentrated solutions were partially inhibited by EDTA, 10^?2 M, at low concentrations of Mg; inhibition was increased in diluted and purified preparations.1,10-phenanthroline (Ophen) strongly inhibited exsheathing activity (Zn < 1 × 10^?6 M). When Zn²⁺, 10^?3 M was added, the inhibition was abolished. The hydrolysis of l-leucinamide was greatly increased in the presence of Ophen, 10^?4 M; this effect was abolished by adding Zn²⁺, 10^?3 M.It is suggested that exsheathing fluid from at least some ‘strains’ of H. contortus contains a Zn metallo-enzyme, probably LAP, which is involved in the process of exsheathment.     

The yield of chlorophyll a fluorescence as a means to test the various deexcitation mechanisms in the antenna system of green plants

With the aid of measurements of the fluorescence yield, the efficiency of the various deexcitation mechanisms of an exciton in the light-harvesting system has been determined. For this purpose, the fluorescence of dark-adapted as well as of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU)-treated and preilluminated leaves of Zea mays L. was excited by single ultrashort laser pulses of different energies. The experimental results have served for the fitting of solutions of rate equations, which describe the deexcitation by linear relaxation processes like fluorescence and radiationless transitions, by annihiation of excitons, and by traps both in the ground state and in an excited state. We have obtained the following results: a ratio of antenna chlorophyll molecules to Photosystem II traps of 600:1, an annihilation constant γ = 2·10^?8 cm³·s^?1, a mean trapping time of $\text{t}\text{?}\text{=0.5}\text{ns}$, a trapping probability for traps in the ground state of 2·10^?8 cm³·s^?1, and 6·10^?9 cm³·s^?1 for traps in an excited state.     

Calcium accumulation by chick intestinal mitochondria: Regulation by vitamin D-3 and 1,25-dihydroxyvitamin D-3

We have evaluated the effect of vitamin D-3 and its metabolite 1,25-dihydroxyvitamin D-3 on Ca²⁺ accumulation by chick intestinal mitochondria. Ca²⁺ accumulation appears to occur in two phases: an early, transient accumulation into an Na⁺-labile pool followed by an ATP-dependent accumulation into an Na⁺-resistant pool. Ca²⁺ accumulation is extensive at free Ca²⁺ concentrations greater than 3 · 10^?6 M in the presence of ATP. Ruthenium red and dinitrophenol block Ca²⁺ accumulation, but atractyloside does not. Oligomycin blocks ATP-supported accumulation completely with a partial inhibition of ATP and malate-supported accumulation. Little difference could be found in mitochondrial preparations from vitamin D-deficient chicks compared to those from vitamin D-3 (or 1,25(OH)₂D-3)-supplemented chicks with respect to respiratory control, oxygen consumption, efficiency of oxidative phosphorylation, affinity for Ca²⁺, or the rate and extent of ATP-supported Ca²⁺ accumulation. Intestinal cytosol stimulated Ca²⁺ accumulation, but this was not specific with respect to vitamin D status or tissue of origin, nor was it duplicated by chick intestinal Ca²⁺-binding protein. 30 ng/ml 1,25(OH)₂D-3 stimulated Ca²⁺ accumulation directly, regardless of the presence of intestinal cytosol. Other vitamin D metabolites were less potent: 25-hydroxyvitamin D-3 > 24,25-dihydroxyvitamin D-3 = vitamin D-3. Since increasing the free Ca²⁺ concentration from 3 · 10^?6 to 1 · 10^?5 M increased Ca²⁺ accumulation approx. 50-fold, whereas direct stimulation by 1,25(OH)₂D-3 in vitro increased Ca²⁺ accumulation less than 2-fold, we conclude that 1,25(OH)₂D-3 influences mitochondrial accumulation of Ca²⁺ in vivo primarily by altering cytosol concentrations of free Ca²⁺.     

Theoretical study and rate constant calculations for the reactions of SiHX3 with CF3 and CH3 radicals (X = F, Cl)

Theoretical investigations were carried out on the multi-channel reactions CF₃ + SiHF₃, CF₃ + SiHCl₃, CH₃ + SiHF₃, and CH₃ + SiHCl₃. Electronic structures were calculated at the MP2/6-311+G(d,p) level, and energetic information further refined by the MC-QCISD (single-point) method. The rate constants for major reaction channels were calculated by the canonical variational transition state theory with small-curvature tunneling correction over the temperature range of 200–1,500 K. The theoretical rate constants were in good agreement with the available experimental data and were fitted to the three parameter expression: k _1a(T) = 2.93 × 10^?26 T ^4.25 exp (?318.68/T), and k _2a(T) = 3.67 × 10^?22 T ^2.72 exp (?1,414.22/T), k _3a (T) = 7.00 × 10^?24 T ^3.27 exp (?384.04/T), k _4a(T) = 6.35 × 10^?22 T ^2.59 exp (?603.18/T) (in unit of cm³molecule^?1s^?1) are given. Our calculations indicate that hydrogen abstraction channel is the major channel due to the smaller barrier height among four channels considered.

Active site-specifically reconstituted nickel(II) horse liver alcohol dehydrogenase: Optical spectra of binary and ternary complexes with coenzymes,coenzyme analogues,substrates, and inhibitors

Insertion of nickel ions into the empty catalytic site of horse liver alcohol dehydrogenase yields an active enzyme with 65% metal substitution and about 12% intrinsic activity. The electronic absorption spectrum is characterized by bands at 357 nm (2900 M^?1 cm^?1, 407 nm (3500 M^?1 cm^?1), 505 nm (300 M^?1 cm^?1), 570 nm (?130 M^?1 cm^?1), and 680 nm (?80 M^?1 cm^?1). The absorption and CD spectra are similar to those of nickel(II) azurin and nickel(II) aspartate transcarbamoylase and prove coordination of the nickel(II) ions to sulfur in a distorted tetrahedral coordination geometry. Changes of the spectra upon ligand binding at the metal or conformation changes of the protein induced by coenzyme, or both, indicate alterations of the metal geometry.The chromophoric substrate trans-4-(N, N-dimethylamino)-cinnamaldehyde forms a ternary complex with Ni(II) liver alcohol dehydrogenase and the coenzyme analogue 1,4,5,6-tetrahydronicotinamide-adenine-dinucleotide, stable between pH 6 and 10. The corresponding ternary complex with NADH is only stable at pH > 9.0. The spectral redshifts induced in the substrate are 11 nm larger than those found in the zinc enzyme. We suggest direct coordination of the substrate to the catalytic metal ion which acts as a Lewis acid in both substrate coordination and catalysis.     

Development of selective inhibitors for the treatment of rheumatoid arthritis: (R)-3-(3-(Methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)pyrrolidin-1-yl)-3-oxopropanenitrile as a JAK1-selective inhibitor

A series of 3(R)-aminopyrrolidine derivatives were designed and synthesized for JAK1-selective inhibitors through the modification of tofacitinib’s core structure, (3R,4R)-3-amino-4-methylpiperidine. From the new core structures, we selected (R)-N-methyl-N-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine as a scaffold for further SAR studies. From biochemical enzyme assays and liver microsomal stability tests, (R)-3-(3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)pyrrolidin-1-yl)-3-oxopropanenitrile (6) was chosen for further in vivo test through oral administration. Compound 6 showed improved selectivity for JAK1 compared to that of tofacitinib (IC₅₀ 11, 2.4?×?10², 2.8?×?10³, and 1.1?×?10²?nM for JAK1, JAK2, JAK3, and TYK2, respectively). In CIA and AIA model tests, compound 6 exhibited similar efficacy to tofacitinib citrate.     

Kinetics of nucleotide binding to chloroplast coupling factor (CF1)

Studies of the kinetics of association and dissociation of the formycin nucleotides FTP and FDP with CF₁ were carried out using the enhancement of formycin fluorescence. The protein used, derived from lettuce chloroplasts by chloroform induced release, contains only 4 types of subunit and has a molecular weight of 280 000.In the presence of 1.25 mM MgCl₂, 1 mol of ATP or FTP is bound to the latent enzyme, with K_d = 10^?7 or 2 · 10^?7, respectively. The fluorescence emission (λ_max 340 nm) of FTP is enhanced 3-fold upon binding, and polarization of fluorescence is markedly increased. The fluorescence changes have been used to follow FTP binding, which behaves as a bimolecular process with K₁ = 2.4 · 10⁴ M^?1 · s^?1. FTP is displaced by ATP in a process apparently involving unimolecular dissociation of FTP with k_?1 = 3 · 10^?3 s^?1. The ratio of rates is comparable to the equilibrium constant and no additional steps have been observed.The protein has 3 sites for ADP binding. Rates of ADP binding are similar in magnitude to those for FTP. ADP and ATP sites are at least partly competitive with one another.The kinetics of nucleotide binding are strikingly altered upon activation of the protein as an ATPase. The rate of FTP binding increases to at least 10⁶ M^?1 · s^?1. This suggests that activation involves lowering of the kinetic barriers to substrate and product binding-dissociation and has implications for the mechanism of energy transduction in photophosphorylation.     

The kinetics and specificity of electron transfer from cytochromes and copper proteins to P700

The rates of electron transfer to P700 from plastocyanin and cytochrome f have been compared with those from three other c-type cytochromes and azurin, a copper protein resembling plastocyanin. Three different disruptive techniques were used to expose P700; digitonin, Triton X-100 and sonication. The following rate constants were measured at 25 °C, pH 7.0, with digitonin-treated chloroplasts: plastocyanin, 8 · 10⁷ M^?1 · s^?1; red-algal cytochrome c-553, 1.9 · 10⁷ M^?1 · s^?1; Pseudomonas cytochrome c-551, 8 · 10⁶ M^?1 · s^?1; azurin, ? 3 · 10⁵ M^?1 · s^?1; cytochrome f, ? 2 · 10⁴ M^?1 · s^?1; mammalian cytochrome c, ? 2 · 10⁴ M^?1 · s^?1. For electron transfer from plastocyanin, the effects of ionic strength, pH and temperature were also studied, and saturation effects found in earlier work were avoided by a full consideration of the various secondary reactions and inclusion of superoxide dismutase. The relative rates are discussed in relation to photosynthetic electron transport.     

Novel Potato Micro-Tuber-Inducing Compound, (3R,6S)-6-Hydroxylasiodiplodin,from a Strain of Lasiodiplodia theobromae

A novel potato micro-tuber-inducing compound was isolated from the culture broth of Lasiodiplodia theobromae Shimokita 2. The structure of the isolated compound was determined as (3R,6S)-6-hydroxylasiodiplodin by means of spectroscopic analyses, the modified Mosher method, and chemical conversion. The compound showed potato micro-tuber-inducing activity at a concentration of 10^?4 M, using the culture of single-node segments of potato stems in vitro.     

A specific prostaglandin I2 synthetase inhibitor, 3-hydroperoxy-3-methyl-2-phenyl-3H-indole.

Inhibitory effects of 3-hydroperoxy-3-methyl-2-phenyl-3H-indole(HPI) on prostaglandin endoperoxide synthase(EC 1.14.99.1) and prostaglandin I₂(PGI₂) synthetase were compared with those of 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid, namely, 15-hydroperoxyarachidonic acid(15-HPAA) and tranylcypromine (TCP). Sheep seminal vesicle microsomes were used as a source of prostaglandin endoperoxide synthase and bovine aortic microsomes as that of PGI₂ synthetase. 15-HPAA and HPI inhibited PGI₂ synthetase with IC₅₀s of 5 × 10^?7 and 3.5 × 10^?6 M, respectively, whereas neither compound had effect on prostaglandin endoperoxide synthase at the concentration inhibiting PGI₂ synthetase by 90%. TCP was a weak(IC₅₀ = 5 × 10^?4M) PGI₂ synthetase inhibitor with low specificity.     

The isolation and characterization of 3-(2-carboxyethyl)cytosine following in vitro reaction of β-propiolactone with calf thymus DNA

The new adduct 3-(2-carboxyethyl)cytosine (3-CEC) was isolated following in vitro reaction of the carcinogen β-propiolactone (BPL) with calf thymus DNA. The structure of 3-CEC was confirmed by synthesis from BPL and dCyd. Reaction of BPL with cCyd (pH 7.0–7.5, 37°C) gave 3-(2-carboxyethyl)deoxycytidine (3-CEdCyd) (9% yield) and 3,N⁴-bis(2-carboxyethyl)deoxycytidine (3,N⁴-BCEdCyd) (0.6% yield). 3-CEdCyd and 3,N⁴-BCEdCyd were hydrolyzed (1.5 N HC1, 100°C, 2 h) to 3-CEC and 3,N⁴-bis(2-carboxyethyl)cytosine (3,N⁴-BCEC), respectively. The structure of 3-CEC was assigned on the basis of UV and NMR spectra and the electron impact (EI) mass spectra of 3-CEC and a tri-trimethylsilyl (TMS) derivative of 3 CEC as well as deuterated (d₂₇) tri-TMS derivative of 3-CEC. The structure of 3,N⁴-BCEC was assigned on the basis of UV spectra and the EI mass spectra of a tri-TMS derivative. EI and isobutane chemical ionization mass spectra of 3-methylcytosine (3-MeCyt) and a di-TMS derivative of 3-MeCyt were obtained and were helpful in deducing the structures of 3-CEC and 3,N⁴-BCEC. This is the first report of the alkylation by BPL of an exocyclic atom on a base in DNA. Compound 3,N⁴-BCEC was not detected in BPL-reacted calf thymus DNA. The relative amounts of 1-(2-carboxyethyl)adenine (1-CEA), 7-(2-carboxyethyl)guanine (7-CEG), 3-(2-carboxyethyl)thymine (3-CET) and 3-CEC isolated from BPL-reacted DNA following perchloric acid hydrolysis were 0.23, 1.00, 0.39 and 0.41 respectively, when the alkylation reaction was conducted in phosphate buffer at 0–5°C and pH 7.5 and 0.10, 1.00, 0.29 and 0.28 respectively when the reaction was conducted in H₂O at 37°C and pH 7.0–7.5.     

Cell Cycle activation and ouabain-sensitive ion movements of 3T3 and C3H-10T½ fibroblasts

The introduction of either PGF_2α (10^?7 M) or TPA (10^?7 M) stimulated, ouabain-sensitive ⁸⁶Rb⁺ influx at 30 min in postconfluent 3T3-4 mouse fibroblast cultures by 117% and 124%, respectively. Both TPA and PGF_2α at these concentrations stimulated the incorporation of ³H-TdR into DNA. TPA had the greatest stimulatory effect, which was similar to that obtained with 10% fetal calf serum. In accord with the idea that modulation of membrane processes such as Na⁺/K⁺ pump activity in fibroblasts may reflect important events related to the initiation of DNA synthesis, it was observed that in both 3T3-4 and C3H-1 0T½ cells there were parallel increases in ³H-TdR incorporation and ouabain-sensitive ⁸⁶Rb⁺ influxes with 10^?7 M TPA, whereas PGF_2α stimulated a significant increase in ³H-TdR incorporation in 3T3-4 but not C3H-10T½ cells and only marginal increases in ouabain-sensitive ⁸⁶Rb⁺ influx in both. Therefore, although there appears to be a close correlation between Na⁺/K⁺ pump activation and subsequent S-phase entry following TPA stimulation, a similar correlation for PGF_2α cannot be confirmed.     

Binding of 14-3-3 proteins to a single stranded oligodeoxynucleotide aptamer

A synthetic library of ca. 10¹³ single stranded oligodeoxynucleotides, each comprising a randomized 40mer sequence and homogeneous 10mer flanking regions, was screened for binding to recombinant human 14-3-3γ. A single aptamer, which showed similar affinities (K_D ∼ 10⁻⁸ M) for six isoforms of the protein, has been shown to bind to undenatured 14-3-3 protein in the cerebral spinal fluid of scrapie infected sheep.     

Radiosynthesis and evaluation of [11C]CMP,a high affinity GSK3 ligand

Dysfunction of GSK3 is implicated in the etiology of many brain, inflammatory, cardiac diseases, and cancer. PET imaging would enable in vivo detection and quantification of GSK3 and can impact the choice of therapy, allow non-invasive monitoring of disease progression and treatment effects. In this report, the synthesis and evaluation of a high affinity GSK3 ligand, [¹¹C]2-(cyclopropanecarboxamido)-N-(4-methoxypyridin-3-yl)isonicotinamide, ([¹¹C]CMP, (3), (IC₅₀?=?3.4?nM, LogP?=?1.1) is described. [¹¹C]CMP was synthesized in 25?±?5% yield by radiomethylating the corresponding phenolate using [¹¹C]CH₃I. The radioligand exhibited modest uptake in U251 human glioblastoma cell lines with ～50% specific binding. MicroPET studies in rats indicated negligible blood–brain barrier (BBB) penetration of [¹¹C]CMP, despite its high affinity and suitable logP value for BBB penetration. However, administration of cyclosporine prior to [¹¹C]CMP injection showed significant improvement in brain radioactivity uptake and the tracer binding. This finding indicates that [¹¹C]CMP might be a P-gp efflux substrate and therefore has some limitations for routine in vivo PET evaluations in brain.     

A diffusional analysis of the temperature sensitivity of the Mg2+-induced rise of chlorophyll fluorescence from pea thylakoid membranes

The kinetics of the chlorophyll fluorescence rise induced by adding 20 mM MgCl₂ to a suspension of isolated pea chloroplasts treated with 3-(3,4-dichlorophenyl)-1, 1-dimethylurea (DCMU) have been examined experimentally and theoretically as a function of temperature. The application of similarity arguments and particle aggregation theory to the experimental results suggests that at the first approximation, the salt-induced time-dependent fluorescence changes may be described by the diffusion-controlled lateral movement of Photosystem II pigment-protein complexes. From an analysis of the temperature dependence of the fluorescence changes, estimates obtained for the lateral diffusion coefficients were 1.85 · 10^?12–3.08 · 10^?11 cm²/s over the temperature range 10°C ? T?30°C.     

Inhibition by dexamethasone of the in vitro transport of 3-O-methylglucose into rat thymocytes

Reduced glucose transport across the plasma membrane and reduced phosphorylation may both be responsible for the early inhibitory effect of physiological concentrations of glucocorticoids on glucose uptake by rat thymocytes.The early inhibitory effects of glucocorticoids (5 · 10^?7 M dexamethasone) on glucose consumption and ¹⁴CO₂ formation from d-[U-¹⁴C]glucose were reproduced.The total uptake curve of 4.8 μM $\text{3-O-[}{}^{14}\text{C}\text{]}\text{methyl-}\text{d}\text{-glucose}$ was biexponential with $\text{t}\text{1}\text{2}$ of 1.1 min and 36 min, respectively, the rapid part comprising about 50% of the equilibrated intracellular water space. The latency of the effect of 5 · 10^?7 M dexamethasone on $\text{3-O-[}{}^{14}\text{C}\text{]}\text{methyl-}\text{d}\text{-glucose}$ uptake ranged from 15 to 100 min and the inhibition varied from 15 to 55% independently of the lag period. The effect of 3-O-methylglucose concentration on the initial uptake by steroid-responsive cell preparations was tested after 45 min of preincubation with or without 5 · 10^?7 M dexamethasone. In 12 experiments dexamethasone reduced V from 1.36 ± 0.16 mmol · min^?1 · l^?1 cell water to 0.81 ± 0.10 mmol · min^?1 · l^?1 cell water with insignificant change of K_m (6.0 mM versus 5.9 mM). Dexamethasone had similar effect after 90 or 120 min.The variabilities of control cell transport capacity, the lag period and the magnitude of the dexamethasone effect could not be accounted for by changes in pH, effects of cell density, concentrations of albumin, ethanol, nucleosides, pyruvate or correlated to age and sex of the rats. In conclusion the inhibition of glucocorticoids on glucose consumption by thymocytes appears to be an inhibited plasma membrane transport capacity.