Inhibition of photosynthesis by solar radiation in Dunaliella salina: relative efficiencies of UV-B, UV-A and PAR

Inhibition of photosynthesis after exposure to solar radiation was investigated in the marine green alga Dunaliella salina by monitoring photosynthetic optimal quantum yield F_v/F_m and efficiency of oxygen production. Samples were exposed to solar radiation in Ancient Korinth, Greece (37°58′ N, 23°0′ E) in August 1994. Within 30 min, F_v/F_m and efficiency of oxygen production decreased with similar kinetics with increasing exposure time. The inhibition, however, diminished when ultraviolet radiation was progressively excluded by means of colour filter glasses. Samples exposed for 3 h showed complete or partial recovery of photosynthesis, with almost the same rate under all irradition conditions. The fit of the experimental data with an analytical model describing inhibition of photosynthesis as a function of a linear combination of the photon fluence in the UV-B, UV-A and PAR allows one to estimate the relative mean effectiveness for inhibition by the three spectral ranges [about 2 × 10^?4, 4 × 10^?6 and 2 × 10^?7 (μmol photons m^?2)^?1 for UV-B, UV-A and PAR, respectively].     

INHIBITION OF ACETYLCHOLINESTERASE IN VITRO BY PENTYLENETETRAZOL1

Abstract—The effect of pentylenetetrazol (PTZ) on acetylcholinesterase (E.C.3.1.1.7) was studied in vitro. The kinetics of the reaction were studied on AChE in crude homogenates of rat brain and in purified preparations from Electrophorus electricus. The K_m for rat brain AChE was 1·22 × 10^-4m, with a V_max of 1·37 μmol/g/min whereas the K₄ for competitive inhibition of the enzyme by PTZ was 4·7 × 10^-3m. The commercially purified enzyme exhibited a K_m of 1·73 × 10^-4m and a K_i of 1·00 × 10^-3m.     

BENDING STIFFNESS OF CYLINDRICAL PLANT ORGANS WITH A ‘CORE-RIND’ CONSTRUCTION: EVIDENCE FROM JUNCUS EFFUSUS LEAVES

The mechanical behavior of leaves of Juncus effusus L. in bending was investigated in terms of a closed-form analytical solution derived to predict the bending stiffness of a cylindrical sandwich beam consisting of an outer ‘rind’ (sclerenchyma and chlorenchyma) and an inner ‘core’ (aerenchyma). The elastic moduli (E_TOTAL) of intact leaves was measured by means of multiple resonance frequency spectra and compared to that of leaves for which the aerenchymatous core was surgically destroyed. Based on ten leaves, E_TOTAL = 22.33 × 10⁴ ± 5.37 ± 10⁴ kg · cm^–2 while the elastic modulus of the ‘rind’ was 22.29 × 10⁴ ± 5.69 × 10⁴ kg · cm^–2. The elastic modulus of the ‘core’ was estimated at 3.12 × 10⁴ ± 1.42 × 10⁴ kg · cm^–2. Load-deflection curves for three leaf segments indicated leaves were linearly elastic within the range of loading and could be predicted with considerable accuracy based on the closed-form solution. The aerenchymatous core was found to contribute very little to the bending stiffness of leaves, although its contribution appeared to increase as leaf diameter decreased. Leaves mechanically failed by Brazier buckling when excessively loaded and were best considered to mechanically operate as hollow tubes. Nonetheless, the analytical solution for bending stiffness could be applied and, in theory, can be used to predict the behavior of other plant organs with a ‘corerind’ construction.     

Kinetic Properties of Fungal Polyamine Oxidases and Their Application to Differential Determination of Spermine and Spermidine

Polyamine oxidase from Penicillium chrysogenum oxidized spermine rapidly and spermidine slightly at pH 7.5. The apparent Km values for spermine and spermidine were calculated to be 2.25 × 10^?5 m and 9.54 × 10^?6 m, respectively. The relative maximum velocities for spermine and spermidine were 3.37 × 10^?3 m (H₂O₂) per min per mg of protein and 2.08 × 10^?4 m (H₂O₂) per min per mg of protein, respectively. Spermine oxidation of the enzyme was competitively inhibited by spermidine and putrescine. The apparent Ki values by spermidine and putrescine were calculated to be 3.00 × 10^?5 m and 1.80 × 10^?8 m, respectively. On the other hand, polyamine oxidase from Aspergillus terreus rapidly oxidized both spermidine and spermine at pH 6.5. The apparent Km values for spermidine and spermine were 1.20 × 10^?8 m and 5.37 × 10^?7 m, respectively. The relative maximum velocities for spermidine and spermine were 1.55 × 10^?2 m (H₂O₂) per min per mg of protein and 6.20 × 10^?3 m (H₂O₂) per min per mg of protein, respectively.

Differential determination of spermine and spermidine was carried out using the two enzymes. The initial rate was assayed with Penicillium enzyme and the end point was measured afte addition of Aspergillus enzyme. Small amounts of polyamines (25 to 200 nmol of spermine and 25 to 250 nmol of spermidine) were assayed by solving two simultaneous equations obtained from the rate assay method and the end point assay method. The calculated values were in close agreement with those obtained by an amino-acid analyzer.     

THE TIME-COURSE OF PHOTOADAPTATION TO LOW-LIGHT IN PROROCENTRUM MARIAE-LEBOURIAE (DINOPHYCEAE)1

The estuarine dinoflagellate, Prorocentrum mariaelebouriae (Parke & Ballantine 1957) Faust 1974 undergoes increases in pigmentation and photosynthetic efficiency within several days of downward light shifts. These changes can be described by first-order kinetics, as has been reported previously for Chlorophyll (Chl) a in several phytoplankton species. The studies described in this paper were conducted with isolates of populations of Prorocentrum from the Chesapeake Bay. We determined rates of adaptation to low-light for cultures grown at a range of photon flux densities (I₀= 2.65–26.2 E.m^?2, d^?1, shifted to 6.3–7.0% I₀) at three temperatures (10°, 15°, and 20° C), bracketing the conditions this species experiences in situ. In this paper, I report the time-course of changes in α, P_max Chl a, peridinin, and I_k and first-order rate constants, K₁ for changes in α, Chl a and peridinin. cell^?1. K₁ for changes in α cell^?1 averaged 1.58 × 10^?2 h^?1 for conditions encompassing five light treatments and three temperatures; the corresponding mean for Chl a was 1.59 × 10^?2 h^?1. Increases in peridinin measured for five light treatments at 15° C showed a mean K₁ of 1.22 × 10^?2 h^?1, Average percent changes in per cell α, Chl a, and peridinin ranged from 0.4–4.0% h^?1 (10–90% d^?1) following exposure to low-light. Photoadaptive changes are important to Prorocentrum because in nature it occupies turbid waters (K_t≥ 0.5 m^?1) where the mixing depth often exceeds the depth of the photic layer. Cells are entrained beneath a seasonally-stable density discontinuity and are exposed to very low-light (< I E.m^?2.d^?1) for days to weeks during subpycnocline transport. The ability of this species to undergo changes in pigmentation and photosynthetic physiology confers increased efficiency of light harvesting and contributes to this species’survival in the estuary where it is an important component of the dinoflagellate flora.     

Resonance effect of millimeter waves in the power range from 10-19 to 3 × 10-3 W/cm2 on Escherichia coli cells at different concentrations

The effect of millimeter waves (MMWs) on the genome conformational state (GCS) of E. coli AB1157 cells was studied by the method of anomalous viscosity time dependencies (AVTD) in the frequency range of 51.64-51.85 GHz. The 51.755 GHz resonance frequency of the cell reaction to MMWs did not depend on power density (PD) in the range from 10^-19 to 3 × 10^-3 W/cm². The half-width of the resonant reaction of cells showed a sigmoid dependence on PD, changing from 3 MHz to 100 MHz. The PD dependence of the half-width had the same shape for different concentrations of exposed cells (4 × 10⁷ and 4 × 10⁸ cells/ml), whereas the magnitude of the 51.755 GHz resonance effect differed significantly and depended on the PD of MMW exposure. Sharp narrowing of the 51.755 GHz resonance in the PD range from 10^-4 to 10^-7 W/cm² was followed by an emergence of new resonance frequencies. The PD dependence of the MMW effect at one of these resonance frequencies (51.674 GHz) differed markedly from the corresponding dependence at the 51.755 GHz resonance, the power window occurring in the range from 10^-16 to 10^-8 W/cm². The results obtained were explained in the framework of a model of electron-conformational interactions. The frequency-time parameters of this model appeared to be in good agreement with experimental data. © 1996 Wiley-Liss, Inc.     

Tricyclohexyltin hydroxide effects on cationic and substrate activation kinetics of beta-adrenergic–stimulated cardiac Ca2+-ATPase

Previous studies from this laboratory have indicated that tricyclohexyltin hydroxide (Plictran) is a potent inhibitor of both basal- and isoproterenol-stimulated cardiac sarcoplasmic reticulum (SR) Ca²⁺-ATPase, with an estimated IC-50 of 2.5 × 10^?8M. The present studies were initiated to evaluate the mechanism of inhibition of Ca²⁺-ATPase by Plictran. Data on substrate and cationic activation kinetics of Ca²⁺-ATPase indicated alteration of V_max and K_m by Plictran (1 and 5×10^?8M), suggesting a mixed type of inhibition. The beta-adrenergic agonist isoproterenol increased V_max of both ATP- and Ca²⁺-dependent enzyme activities. However, the K_m of enzyme was decreased only for Ca²⁺ Plictran inhibited isoproterenol-stimulated Ca²⁺-ATPase activity by altering both and V_max and K_m of ATP as well as Ca²⁺-dependent enzyme activities, suggesting that after binding to a single independent site, Plictran inhibits enzyme catalysis by decreasing the affinity of enzyme for ATP as well as for Ca²⁺ Preincubation of enzyme with 15 μM cAMP or the addition of 2mM ATP to the reaction mixture resulted in slight activation of Plictran-inhibited enzyme. Pretreatment of SR with 5 × 10^?7M propranolol and 5 × 10^?8M Plictran resulted in inhibition of basal activity in addition to the loss of stimulated activity. Preincubation of heart SR preparation with 5 × 10^?5M coenzyme A in combination with 5 × 10^?8M Plictran partly restored the beta-adrenergic stimulation. These results suggest that some critical sites common to both basal- and beta-adrenergic-stimulated Ca²⁺-ATPase are sensitive to binding by Plictran, and the resultant conformational change may lead to inhibition of beta-adrenergic stimulation.     

Purification and Properties of Formyltetrahydrofolate Synthetase from Spinach

Formyltetrahydrofolate synthetase (E. C. 6. 3. 4. 3) was found in fresh spinach leaves and purified about 60-fold by treatments of ammonium sulfate, protamine sulfate, dialysis, and DEAE-cellulose column chromatography. Some properties of the enzyme were investigated. Optimum pH was found to be 7.5, and optimum temperature was observed to be at 37°C. In the enzyme reaction, FAH₄ and formate were required specifically as the substrates, and Mg⁺⁺ and ATP were essential components. The Michaelis constants for dl-FAH₄, formate, ATP and magnesium chloride were 1.7×10^?3 m, 1.7×10^?2 m, 4.1×10^?4 m and 3.3×10^?3 m, respectively. The primary product formed in the reaction catalyzed by the enzyme was suggested as N¹⁰-formyl-FAH₄ spectrophotometrically. It was observed that the enzyme also catalyzed the reverse reaction. The possible role of the enzyme in plants was discussed.     

In Vitro Culture of Rudimentary Embryos of Ilex paraguariensis: Responses to Exogenous Cytokinins

The effects of benzyladenine (BAP), kinetin (KIN), zeatin (ZEA), isopentenyladenine (2iP), and thidiazuron (TDZ) were studied on in vitro growth of rudimentary embryos of Ilex paraguariensis St. Hil. Heart stage zygotic embryos were removed from seeds of immature, light green fruits and cultured aseptically on quarter-strength Murashige and Skoog medium containing 3% sucrose, 0.65% agar, and supplemented with or without three concentrations of BAP, KIN, ZEA, 2iP, or TDZ. Cultures were incubated in darkness at 27 ± 2°C. Media containing 4.4 × 10⁻⁶ m BAP, 4.6 × 10⁻⁶ m KIN, or 4.9 × 10⁻⁶ m 2iP were totally ineffective in inducing embryo growth after culture for 28 days. However, lower concentrations of these compounds (4.4 × 10⁻⁸ m BAP, 4.6 × 10⁻⁸ m KIN, 4.5 × 10⁻⁸ m ZEA, or 4.9 × 10⁻⁸ m 2iP) promoted embryo growth. TDZ at 9.9 × 10⁻⁹ m, 9.9 × 10⁻⁸ m, or 9.9 × 10⁻⁷ m induced embryo growth at similar rates. The maximum percentage of embryos converted to seedlings was achieved when the medium was supplemented with 4.5 × 10⁻⁷ m ZEA. Received August 1, 1997; accepted February 19, 1998     

Quenching Effect of Tocopherols on the Methyl Linoleate Photooxidation and Their Oxidation Products

The quenching effect of α-, γ- and δ-tocopherols on the methylene blue sensitized photo- oxidation of methyl linoleate was investigated, and the ¹O₂. quenching ability of tocopherols was determined. The ¹O₂ quenching rate constants for α-, γ- and δ-tocopherols in ethanol were estimated to be 2.6 × 10⁸ m^?1 sec^?1, 1.8 × 10⁸ m^?1 sec^?1 and 1.0 × 10⁸ m^?1 sec^?1, respectively. And the rate constants for the chemical reaction between each tocopherol and ¹O₂ were 6.6 × 10⁶ m^?1 sec^?1, 2.6 × 10⁶ m^?1 sec^?1 and 0.7 × 10⁶ m^?1 sec^?1 for α-, γ- and δ-tocopherols, respectively. The results show that α-tocopherol is the most effective compound toward ¹O₂ among the three tocopherols. The photooxidation of each tocopherol produced two peroxides which, after chemical reduction, were identified to be tocopherol hydroquinone by gas chromatography-mass spectrometry analysis. The photooxidation mechanism of these tocopherols was assumed to be different from that of autoxidation.     

Characterization and Regulatory Properties of Glucose-6-Phosphate Dehydrogenase from Black Gram (Phaseolus mungo)

Glucose-6-phosphate dehydrogenase (E.C. 1.1.1.49) was partially purified by fractionation with ammonium sulfate and phosphocellulose chromatography. The K_m value for glucose-6-phosphate is 1.6 × 10^?4 and 6.3 × 10^?4M at low (1.0–6.0 × 10^?4M) and high (6.0–30.0 × 10^?4M) concentrations of the substrate, respectively. The K_m value for NADP⁺ is 1.4 × 10^?5M. The enzyme is inhibited by NADPH, 5-phosphoribosyl-1-pyrophosphate, and ATP, and it is activated by Mg²⁺, and Mn²⁺. In the presence of NADPH, the plot of activity vs. NADP⁺ concentration gave a sigmoidal curve. Inhibition of 5-phosphoribosyl-1-pyrophosphate and ATP is reversed by Mg²⁺ or a high pH. It is suggested that black gram glucose-6-phosphate dehydrogenase is a regulatory enzyme of the pentose phosphate pathway.     

Catalytic properties of three lactate dehydrogenases from potato tubers (Solanum tuberosum)

Two l-lactate dehydrogenase isoenzymes and one dl-lactate dehydrogenase could be separated from potato tubers by polyacrylamide-gel electrophoresis. The enzymes are specific for lactate, while β-hydroxybutyric acid, glycolic acid, and glyoxylic acid are not oxidized. Their pH optima are pH 6.9 for the oxidation and 8.0 for the reduction reaction.The K_m values for l-lactate for the two isoenzymes are 2.00 × 10^?2 and 1.82 × 10^?2, m. In the reverse reaction the affinities for pyruvate are 3.24 × 10^?4 and 3.34 × 10^?4, m. Both enzymes have similar affinities for NAD and NADH (3.00 × 10^?4; 4.00 × 10^?4, and 8.35 × 10^?4; 5.25 × 10^?4, m).The dl-lactate oxidoreductase may transfer electrons either to NAD or N-methyl-phenazinemethosulfate. The K_m values of this enzyme for l-lactate are 4.5 × 10^?2, m and for d-lactate 3.34 × 10^?2, m. Its affinity for pyruvate is 4.75 × 10^?4, m. The enzyme is inhibited by excess NAD (K_m = 1.54 × 10^?4, M) and has an affinity toward NADH (K_m = 5.00 × 10^?3, M) which is about one tenth of that of the two isoenzymes of l-lactate dehydrogenase.     

PROPERTIES OF TYROSINE HYDROXYLASE IN PERIPHERAL NERVES

Approximately 80 per cent of tyrosine hydroxylase activity in bovine mandibular nerve and rabbit sciatic nerve was soluble, and the rest of the activity was particle-bound. The soluble enzyme in bovine mandibular nerve was isolated by ammonium sulphate fractionation (25–35 per cent saturation). The enzyme had a pH optimum at 5·9 in Tris-acetate buffer, and at 6·5 in Tris-HCl or phosphate buffer. The enzyme required a tetrahydropteridine cofactor. K_m values toward various tetrahydropteridines such as l -erythro-tetrahydrobiopterin (a probable natural cofactor), 2-amino-4-hydroxy-6-methyltetrahydropteridine, and 2-amino-4-hydroxy-6,7-dimethyltetrahydropteridine were 2 × 10⁻⁵m , 5 × 10⁻⁵m and 4 × 10⁻⁴m , respectively. The K_m value for tyrosine at 1 × 10⁻³m -2-amino-4-hydroxy-6-methyltetrahydropteridine as a cofactor was 5 × 10⁻⁵m . The enzyme activity was markedly stimulated with Fe²⁺ or catalase, but Fe²⁺ gave higher activity. The activity was inhibited with α, α′-dipyridyl, l -α-methyl-p-tyrosine, and various catecholamines. Among catecholamines, dopamine was the most potent inhibitor. l -5-Hydroxytryptophan was an inhibitor as potent as dopamine. Neither d -5-hydroxytryptophan nor 5-hydroxytryptamine inhibited the enzyme. The inhibition by l -5-hydroxytryptophan was partially competitive with tetrahydrobiopterin at concentrations higher than 9 × 10⁻⁵m , and partially uncompetitive at concentrations lower than 9 × 10⁻⁵m . The addition of heparin or lysolecithin did not affect enzyme activity with tetrahydrobiopterin as cofactor.     

Cell density dependent response of E. Coli cells to weak ELF magnetic fields

The effects of weak magnetic fields of extremely low frequency (ELF) on E. coli K12 AB1157 cells were studied by the method of anomalous viscosity time dependencies (AVTD). E. coli cells at different densities within a range of 5 × 10⁵–10⁹ cell/ml were exposed to ELF (sinusoidal, 30 μT peak, 15 min) at a frequency of 9 Hz. A transient effect with maximum 40–120 min after exposure was observed. Kinetics of the per-cell-normalised ELF effects fitted well to a Gaussian distribution for all densities during exposure. A maximum value of these kinetics and a time for this maximum were strongly dependent on the cell density during exposure. These data suggest a cell-to-cell interaction during response to ELF. Both dependencies had three regions close to a plateau within the ranges of 3 × 10⁵ − 2 × 10⁷ cell/ml, 4 × 10⁷ − 2 × 10⁸ cell/ml and 4 × 10⁸–10⁹ cell/ml and two rather sharp transitions between these plateaus. The effect reached a maximum value at a density of 4 × 10⁸ cell/ml. Practically no effect was observed at the lowest density of 3 × 10⁵ cell/ml. The data suggested that the ELF effect was mainly caused by a secondary rather than a primary reaction. The filtrates from exposed cells neither induced significant AVTD changes in unexposed cells nor increased the ELF effect when were added to cells before exposure. The data did not provide evidence for significant contribution of stable chemical messengers, but some unstable compounds such as radicals could be involved in the mechanism of cell-to-cell interaction during response to ELF. The results obtained were also in accordance with a model based on an re-emission of secondary photons during resonance fluorescence. Bioelectromagnetics 19:300–309, 1998. © 1998 Wiley-Liss, Inc.     

Membrane transport of L-triiodthyronine by human red cell ghosts.

L-T₃ transport has been investigated in human red cell ghosts. Determination of initial T₃ uptake revealed two separate saturable uptake systems, one with a K_m of 1.6 × 10^?8M, the other with a K_m of 3.3 × 10^?6M. Binding experiments resulted in two dissociation constants, 1.4 × 10^?7M.and 2.6 × 10^?6M. Uptake was dependent on the ghost volume, indicating an intravesicular location of T₃. The T₃ was concentrated 6 times by the ghosts. Ouabain reduced the uptake by the low K_m system, but was without effect on the high K_m system. Thus evidence is provided both of binding of T₃ to the ghost membrane and of its uphill transport across the membrane.     

Properties of glutamate dehydrogenase from Beta vulgaris

Glutamate-NAD oxidoreductase, E.C. 1.4.1.3 (GDH), from seedlings of Beta vulgaris cv. Rota, Jahnsch Peragis Comp., was enzymatically characterized. This enzyme with molecular weight of 2.6 × 10⁵ has a pH optimum of around 8 for animation of α-KGA and around 9.5 for the desamination of glutamate. The apparent K_m for α-KGA is 6.7 × 10^?4M, for NH₃ 2.5 × 10^?3M, for NADH 3.2 × 10^?5M and for NAADPH 5.5 × 10^?4M. NAD¹ inhibits the reaction non-competitively when NADPH serves as substrate. The apparent K¹ is 4.5 × 10^?4M. The data are discussed on relation to the properties of GDH from other plant sources.     

PHOTOPHYSIOLOGICAL EVIDENCE OF NUTRIENT LIMITATION OF PLATELET ICE ALGAE IN MCMURDO SOUND, ANTARCTICA

Seasonally changing photophysiological and biochemical characteristics of sea ice microalgae are interpreted with respect to light availability and measurements of nutrient concentration made at high vertical resolution (12.5 cm) during a dense bloom in the platelet ice layer of McMurdo Sound during a 6-week study in austral spring of 1989. Platelet ice algae remained highly shade adapted throughout the spring as shown by their low photoadaptive index (E_k, 3.7–8.4 μmol photons·m⁻²·s⁻¹), low mean specific absorption coefficient (<0.009 m² mg⁻¹ Chl a), high optical cross-sectional area of photosystem II (σ_PSII, 3.0–8.2), and high molar ratio of fucoxanthin:chlorophyll a (mean = 1.62 ± 0.15 SD). Between 24 October and 8 November, the algae exhibited a photoacclimative response that was marked by a 30% decrease in photosynthetic efficiency (α^B), a 75% decrease in maximum photosynthetic rate (P^B/_m), and a 60% increase in σ_PSII. The photochemical conversion efficiency at photosystem II (F_v/F_m= ca. 0.5) and the quantum yield of photosynthesis (Ø_C= 0.062– 0.078 mol C mol⁻¹ photons) were ca. 80% of their maximal values. After 8 November, changes in algal photophysiology and biochemistry, which were inconsistent with a photoacclimation response, suggest that the platelet ice algae near the platelet/congelation ice interface became increasingly nutrient limited. The number of pennate diatoms increased threefold to 150 × 10⁹ cells m⁻³ between 8 and 14 November, then remained unchanged throughout the remainder of the field season. Following the increase in cell number, F_v/F_m, Ø_C, and C:Chla decreased by >40%, σ_PSII increased by 70%; and the biochemical ratios C:N and C:Si increased 25%–30%. Nutrient depletion was apparent from the high-resolution vertical profiles, but nutrient concentrations limiting algal growth were not observed. However, nutrient concentrations at the likely site of nutrient limitation near the platelet/congelation ice interface were not measured, indicating that higher resolution sampling is necessary to fully characterize this highly variable habitat.     

KINETICS OF AMINO ACID INFLUX INTO NITELLA FLEXILIS1

The uptake of amino acids by Nitella flexilis has been investigated. Influx of glycine, alanine, and valine appears to be a diffusive process. Influx ranged from 0.14 to 0.06 and 0.04 pmoles/(cm)(sec), respectively. Aspartic acid uptake is an active transport mechanism. The V_max is 2.8 pmoles/(cm)(sec); the transport constant (Michaelis constant) K_m, 7.8 × 10^?3 M. The uptake of arginine is apparently due to 2 transport systems, one with a V_max and K_m of 3.1 pmoles/(cm)(sec) and 3.2 × 10^?3M, respectively. The second system has a V_max of 1.4 pmoles/(cm)(sec) and a K_m of 2.1 × 10^?4 M. The possibility that the second system is diffusive has been considered.     

UV-induced mutagenesis at the hypoxanthine—guanine phosphoribosyl transferase locus in two L5178Y mouse lymphoma cell strains with different UV sensitivities

Two strains of L5178Y mouse lymphoma cells, L5178Y-R (LY-R) and L5178Y-S (LY-S), differ markedly in their sensitivity to 254 nm UV radiation (D₀ = 0.7 and 5.5 J/m²; n = 6.0 and 2.0 for LY-R and LY-S cells, respctively). In this study, the frequency o hypoxanthine-guanine-phosporibosyl-transferase-deficient mutants was determined, using 6-thioguanine (TG) as a selective agent, in populations of LY-R and LY-S cells exposed to various fluences of UV radiation. The spontaneous mutation frequency for LY-R cells was (3.7 ± 0.6) × 10^?5 TG^r mutants per viable cell, and the UV induction rate was (2.2 ± 0.8) × 10^?4 TG^r mutants per viable cell, per J/m². Both spontaneous and induced mutantion frequencies were much lower for LY-S cells. The sopntaneous mutation frequency for these cells were too low to make its measurement practicable ( < 0.0013 × 10^?5 TG^r mutants per viable cell). Mutation induction rate was (4.2 ± 2.2) × 10^?7 TG^r mutants per viable cell, per J/m². These differences in mutability do not appear to be due to gene duplication in LY-S cells, or to selective growth disadvantage of LY-S-derived TG-resistant mutants. Possible mechanisms underlying the differences in mutability of LY-R and LY-S cells are considered.     

Steroid-protein interactions. Fluorescence quenching of progesterone-binding globulin and alpha1-acid glycoprotein upon binding of steroids.

The influence of progesterone and four other steroids on the intrinsic fluorescence of progesterone-binding globulin was investigated. The corresponding effect of progesterone on α₁-acid glycoprotein was also studied. The intrinsic fluorescence of the progesterone-binding globulin and of α₁-acid glycoprotein was quenched by about 60 and 17%, respectively, upon forming stoichiometric complexes with progesterone. Graphical analysis of fluorescence quenching titrations with progesterone gave affinity constants at 23 °C of 2 × 10⁹m^?1 for progesterone-binding globulin and 1 × 10⁶m^?1 for α₁-acid glycoprotein. With progesterone-binding globulin, affinity constants of 1 × 10⁹m^?1 were determined for desoxycorticosterone, 1 × 10⁸m^?1 for testosterone, and 2 × 10⁶m^?1 for cortisol. The fluorescence quenching of PBG by 5-pregnen-3β-ol-20-one, 5α-pregnanedione, and 5β-pregnanedione, steroids lacking the Δ⁴-3-keto grouping, was too small to be evaluated; however, binding of the pregnanediones to progesterone-binding globulins was demonstrated when the progesterone-progesterone-binding globulin complex was “unquenched” as a result of competitive displacement of progesterone by addition of the pregnanediones. The quenching phenomenon is assumed to be mainly due to radiationless transfer from protein to the near uv (n → π1) absorption band of steroids containing the Δ⁴-3-keto chromophore.