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THE INDUCIBILITY AND ONTOGENY Of RAT-LIVER UDP-GLUCURONYLTRANSfERASE TOWARD fUROSEMIDE

BIOCHEMICAL PHARMACOLOGY 1986年第21期35卷 3777-3782页

作者： RACHMEL, A HAZELTON, GA UNIV KANSAS MED CTR DEPT PEDIAT KANSAS CITY KS 66103 USA UNIV KANSAS MED CTR DEPT TOXICOL & THERAPEUT KANSAS CITY KS 66103 USA

furosemide (f) conjugation with glucuronic acid is the main pathway of f metabolism in humans and experimental animals. In order to study rat liver microsomal UDP-glucuronyltransferase (UDP-GT) activity towards f we developed an in vitro assay in which the conjugation product, furosemide 1-0-acyl glucuronide (fG) was separated and quantitatively determined by reverse phase high pressure liquid chromatography. The optimal conditions of the reaction were established and the apparent Km for f and UDP-glucuronic acid (UDPGA) were 0.22 and 1.76 mM, respectively. Substrate inhibition of UDP-GT toward f occurred at f concentrations higher than 1.5 mM. Developmental changes in f glucuronidation were compared to the ontogeny of UDP GT activity toward two other acceptors, 1-naphthol and esterone that are known to have different patterns of maturation. f glucuronidation was 26% of adult activity at 18 days of gestation, reached 48% at birth and gradually increased to 250% of adult activity at 22 days of age. Glucuronidation of 1-naphthol and estrone attained 87% and 44% of adult activity at 22 days of gestation, 37% and 66% in six-day-old rats and 100% and 427% of adult activity in 22-day-old rats, respectively. The effect of 3-methylcholanthrene (3-MC), phenobarbital (PB) and ***.-carbonitrile (PCN) on f UDP-GT was studied and compared to their effect on 1-naphthol and estrone glucuronidation. PB, 3-MC and PCN increased f-UDP-GT activity to 208% 282% and 342% of vehicle-treated animals, respectively, while f pretreatment did not affect the conjugation of f. In comparison, 1-naphthol glucuronidation was preferentially induced by 3-MC (4.4-fold of control) while estrone glucuronidation was induced by PB and PNC (4.9- and 2.5-fold of control, respectively). These studies suggest that several forms of UDP-GT activities, which differ in their ontogeny and inducibility patterns, are involved in the glucuronidation of f in vitro.

关键词： f furosemide fG furosemide 1-0-acyl glucuronide UDP-GT UDP-glucuronyltransferase UDPGA UDP-glucuronic acid 3-MC 3-methylcholanthrene PB phenobarbital PCN pregnenolone-16α-carbonitrile CHAPS 3 [(cholamidopropyl) dimethylamonio]-1-propranesulfonate

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AN ELECTROCHEMICAL STUDY Of ENERGY-DEPENDENT POTASSIUM ACCUMULATION IN ESCHERICHIA-COLI .11. THE TRK SYSTEM IN ANAEROBICALLY AND AEROBICALLY GROWN CELLS

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BIOELECTROCHEMISTRY AND BIOENERGETICS 1986年第3期15卷 417-426页

作者： MARTIROSOV, SM TRCHOUNIAN, AA YEREVAN STATE UNIV FAC BIOL DEPT BIOPHYS EREVAN 375049 ARMENIA USSR

The interaction of H+-ATPase complex ***.f0 and the Trk system of K+ accumulation was studied in Escherichia coli grown quasi-anaerobically in peptone media with glucose (anaerobia) and aerobically in salt medium with succinate (aerobia). In anaerobia the Trk system takes part in H+-K+ exchange, displaying km = 3.7 mM, vmax = 1.6 mM g-1 min-1 and in aerobia in the Trk system has Km = 3.4 mM, vmax = 0.45 mM g-1 min-1. The K+ accumulation is blocked by DCC in anaerobia and by cyanide together with DCC in aerobic, whereas protonophores and arsenate block the K+ uptake in bacteria grown under either condition. Valinomycin decreases the K+ accumulation in anaerobic and increases (or has no effect) that in aerobia. ***.f0 is sensitive and the Trk system is insensitive to variation of the external osmotic pressure in both cases. The ratio H+:K+ is stable and equal to 2:1 in anaerobia and is changed from 0.5 to 5.0 in aerobia in response to variation of pH, K+ activity and temperature. Q10 is about 2.8 both for ***.f0 and for the Trk system in anaerobia, but 2.4 and near 1.0 respectively in aerobia. The distribution of K+ in anaerobia is 2500 (potassium equilibrium potential of -210 mV) which is much more than the measured Um of -145 mV. The distribution of K+ in aerobia is 720, which is in good conformity with the measured membrane protential of -175 mV. The structural association of ***.f0 and the Trk system, forming a H+-K+ pump, has been assumed previously to take place in anaerobically grown E. coli;these systems operate separately in aerobic cells. According to Bakker and Helmer et al. the Trk system transport K+ along the electrical field, its function being regulated by ATP.

关键词： EC refers to proteins from E.coli cells upshock refers to an increase and downshock to a decrease in medium osmolarity f 1 ·f 0 and Trk are the systems for ATP-dependent transfer of H + and K + , respectively f 1 is ATPase, f 0 the proteolipid incorporated in a membrane U m the membrane potential DCC is N,N′-dicyclohexylcarbodiimide CCCP is carbonylcyanide- m -chlorphenylhydrazine TPP + is tetraphenylphosphonium cation EDTA is ethylenediaminetetraacetic acid k m and ν max are the Michaelis constant and maximal rate, respectively Q 10 is Van't Hoffs coefficient.

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ESTIMATION Of MICROBIAL MASS CONCENTRATION BASED ON fLUOROMETRIC MEASUREMENT Of CELL PROTEIN

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JOURNAL Of fERMENTATION TECHNOLOGY 1986年第5期64卷 411-417页

作者： TAYA, M AOKI, N KOBAYASHI, T Department of Chemical Engineering Faculty of Engineering Nagoya University Chikusa-ku Nagoya 464 Japan

To develop a convenient method for the estimation of microbial cell mass, the fluorescence intensity of protein (f290) was measured in cell extracts from Escherichia coli, Ruminococcus albus, and Aspergillus niger. Cell extracts obtained by sodium dodecyl sulfate (SDS)-boiling treatment after sonication had high f290 intensities under an excitation wavelength (EXW) of 290 nm and an emission wavelength (EMW) of 340 nm. On the basis of the fluorescence spectra, we confirmed that the fluorescence of the extract was caused by protein in the cell, especially by tryptophan residues in the proteins. Carbohydrate, lipids, nucleic acids, and vitamins from the cells did not interfere significantly with the f290 intensity. The intensity and cell mass concentration were related linearly below 0.15 g-cell/l, and the ratio of the intensity to the cell concentration was evaluated as a constant for each microorganism. It was thus possible to monitor the microbial cultures using the f290 intensity as an indicator, even in culture systems where optical and gravimetric measurements of cell mass were not effective.

关键词： BSA bovine serum albumin DCW dry cell weight, g/ l EMW emission wavelength nm EXW excitation wavelength nm f 290 fluorescence intensity of protein -MLSS mixed liquor suspended solids OD optical density - SDS sodium dodecyl sulfate X cell concentration, g/ l

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QUANTITATIVE STRUCTURE-ACTIVITY RELATIONSHIP Of CARBONYLCYANIDE PHENYLHYDRAZONES AS UNCOUPLERS Of MITOCHONDRIAL OXIDATIVE-PHOSPHORYLATION

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BIOCHIMICA ET BIOPHYSICA ACTA 1986年第1期851卷 93-98页

作者： BALAZ, S STURDIK, E DURCOVA, E ANTALIK, M SULO, P Department of Biochemistry and Microbiology Slovak Polytechnical University 812 37 Bratislava Czechoslovakia

The dependence of the uncoupling activity in the series of 16 carbonylcyanide phenylhydrazones on their physico-chemical properties (partition coefficient, dissociation constant and rate constant for reaction with thiols) is investigated using two physiologically based models, one for protopophoric mechanism of uncoupling and the other assuming the covalent modification of a membrane constituent to be the key step in this process. As indicated by uptake experiments, at the given conditions a lipophilic-hydrophilic equilibrium is attained without any loss of the compounds via chemical reactions. Using this fact to reduce the number of adjustable parameters, a better fit to the data on stimulation of respiration is obtained with the former (protonophoric) model.

关键词： Uncoupler Oxidative phosphorylation Carbonylcyanide phenylhydrazone (Rat liver mitochondria) CCP carbonylcyanide phenylhydrazones K a dissociation constant P apparent partition coefficient k∗ rate constant for reaction of CCP with protein k rate constant for reaction with mercaptoacetic acid n number of points r correlation coefficients s standard deviation f f -test

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fORMATION Of 4,8-DIMEIQX fROM THE MODEL SYSTEM fRUCTOSE, ALANINE AND CREATININE - COMPARISON WITH THE ISOMERIC 5,8-DIMEIQX

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MUTATION RESEARCH 1986年第1期174卷 5-9页

作者： NYHAMMAR, T GRIVAS, S OLSSON, K JAGERSTAD, M SWEDISH UNIV AGR SCI DEPT CHEM & MOLEC BIOL POB 7015 S-75007 UPPSALA SWEDEN UNIV LUND CTR CHEM DEPT NUTR S-22100 LUND SWEDEN

In a previous paper, the main mutagenic compound isolated from the model reaction system D-fructose, DL-alanine and creatinine was tentatively identified as 4,8-DiMeIQx. Its mutagenic activity and spectra characteristics have now been compared with those of the isomer 5,8-DiMeIQx. The comparison clearly demonstrates that the isolated compound was indeed 4,8-DiMeIQx. This finding is in agreement with the hypothesis that sugars, amino acids and creatinine present in meat may be the precursors of the mutagenic imidazoquinolin- and imidazoquinoxalin-2-amines (IQ compounds).

关键词： IQ, 3-methyl-3 H -imidazo[4,5- f ]quinolin-2-amine MeIQ, 3,4-dimethyl-3 H -imidazo[4,5- f ]quinolin-2-amine MeIQx, 3,8-dimethyl-3 H -imidazo[4,5- f ]quinoxalin-2-amine 4,8-DiMeIQx, 3,4,8-trimethyl-3 H -imidazo[4,5- f ]quinoxalin-2-amine 5,8-DiMeIQx, 3,5,8-trimethyl-3 H -imidazo[4,5- f ]quinoxalin-2-amine HPLC, high performance liquid chromatography

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APOLIPOPROTEIN-B, APOLIPOPROTEIN-CIII AND APOLIPOPROTEIN-E IN 2 MAJOR SUBPOPULATIONS Of LOW-DENSITY LIPOPROTEINS

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BIOCHIMICA ET BIOPHYSICA ACTA 1986年第2期879卷 126-133页

作者： LEE, DM ALAUPOVIC, P UNIV OKLAHOMA HLTH SCI CTR DEPT BIOCHEM & MOLEC BIOL OKLAHOMA CITY OK 73104 USA

To determine the concentration and distribution of apolipoproteins C-III and E in low density lipoproteins (LDL) of d 1.025-1.043 g/ml, fresh human plasma was fractionated by single-spin density gradient ultracentrifugation into five layers. Two major subpopulations including layer 2 (d 1.025-1.029 g/ml) and layer 3 (d 1.032-1.043 g/ml) were isolated and characterized by determination of flotation coefficient, neutral lipids and apolipoproteins B, C-III and E. The apolipoprotein B/C-III/E ratio of layer 2 was 100/(3.3 .+-. 2.0)/(5.1 .+-. 2.9) (wt/wt) and that of layer 3 was 100/(0.61 .+-. 0.32)/(0.58 .+-. 0.29) (wt/wt). These weight ratios corresponded to molar ratios of 1.0/(1.90 .+-. 1.16)/(0.74 .+-. 0.42) and 1.0/(0.34 .+-. 0.18)/(0.08 .+-. 0.04), respectively. Layer 2 contained 6-23% of the total plasma apolipoprotein B or 7-27% of total LDL2 (d 1.019-1.063 g/ml) apolipoprotein B. Layer 3 contained 41-65% of plasma apolipoprotein B or 62-86% of of LDL2 apolipoprotein B. About 5-17% of apolipoprotein C-III and 8-30% of apolipoprotein E in plasma are distributed in layers 2 and 3 with the majority present in layer 2. These results show an evident apolipoprotein heterogeneity of LDL2 isolated from normolipidemic subjects. Moreover, they show that the relatively small amounts of apolipoprotein C-II and apolipoprotein E in lower-density segments of LDL2 take on a greater significance when presented in molar rather than weight concentrations. The existence of different ratios of apolipoprotein C-III/apolipoprotein E in layer 2 and layer 3 suggest the presence in LDL2 of varying amounts of several discrete apolipoprotein B- and/or apolipoprotein C-II- and apolipoprotein E-containing lipoprotein particles.

关键词： Apolipoprotein B Apolipoprotein C-III Apolipoprotein E LDL subpopulation (Human plasma) LDL low-density lipoproteins ( d 1.006–1.063 g/ml S f 0–20) LDL 1 ( d 1.006–1.063 g/ml S f 12–20) LDL 2 ( d 1.019–1.063 g/ml S f 0–12) VLDL very low-density lipoproteins ( d < 1.006 g/ml) GSH glutathione, reduced form M r , molecular weight estimated on the basis of relative mobility of proteins on SDS-polyacrylamide gel electrophoresis

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RELATIONSHIP Of STEADY-STATE PHOTOSYNTHESIS TO fLUORESCENCE IN EUKARYOTIC ALGAE

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BIOCHIMICA ET BIOPHYSICA ACTA 1986年第2期849卷 183-192页

作者： fALKOWSKI, PG WYMAN, K LEY, AC MAUZERALL, DC ROCKEFELLER UNIV NEW YORK NY 10021 USA

The change in fluorescence yield (.DELTA..vphi.) was measured in five species [Thalassiosira weisflogii, Skeletonema costatum, Isochrysis galbana, Dunaliella tertiolecta, and Chlorella vulgaris] of eucaryotic algae using a ''pump and probe'' flash technique. The half-time for the oxidation of Q, which was measured by varying the delay time between actinic (pump) and measuring (probe) flashes, averaged 400 .mu.s and was unaffected by background irradiance between 1012 and 1016 quanta .cntdot. cm-1 .cntdot. s-1. The absorption cross-section of PS II traps was measured by varying the intensity of the actinic flash. These cross-sections did not change (.+-. 10%) with background irradiance. The cross-section data can be fitted to a cumulative one-hit Poisson distribution. In the steady state, the relationship between .DELTA..vphi. and photosynthetic oxygen evolution was highly nonlinear and cannot be explained by energy transfer between PS II units. Using the criteria of .DELTA..vphi., about 15% of the PS II traps remain open at light saturation of O2 evolution. Conversely, at low irradiance levels, capable of stimulating much less than 1% of the maximum steady-state photosynthetic rate, the fluorescence yield decreases by as much as 25% from the dark-adapted value. furthermore, the data suggest that a long-lived quencher of fluorescence is formed at moderate to continuous irradiance levels, at least 1016 quanta .cntdot. cm-2 .cntdot. s-1. Our results suggest that cyclic electron flow around PS II occurs under normal physiological conditions and is especially pronounced in chlorophytes.

关键词： Chlorophyll fluorescence Oxygen evolution Photosynthesis Photosystem II fluorescence yield (Eukaryotic algae) Δφ t change in fluorescence yield ( = (f s − f p ) f p ), measured at delay time, t , between ‘pump’ and probe flash f p fluorescence yield of a weak probe flash f s fluorescence yield of a weak probe flash following a ‘pump’ flash f b fluorescence yield from the continuous background light J continuous irradiance level P steady-state photosynthetic oxygen evolution at any irradiance P max maximum steady-state photosynthetic oxygen evolution at saturating continuous irradiance RC reaction center DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea CCCP carbonyl cyanide m -chlorophenylhydrazone

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AMINO-ACID-SEQUENCE Of THE VARIABLE REGION Of HEAVY-CHAIN IN IMMUNOGLOBULIN (MOT) HAVING UNUSUAL PAPAIN CLEAVAGE SITES

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MOLECULAR IMMUNOLOGY 1986年第2期23卷 169-174页

作者： KOJIMA, M KOIDE, T ODANI, S ONO, T NIIGATA PREFECTURE PUBL HLTH LAB CLIN PATHOL SECT NIIGATA 95021 JAPAN NIIGATA UNIV SCH MED DEPT BIOCHEM NIIGATA 951 JAPAN

An IgGl(.lambda.) Mot myeloma protein showed a unique susceptibility toward papain digestion. The fab fragment of Mot was more digestible with papain than the fc fragment. This phenomenon was found to occur by unusual cleavage of the fd fragment with papain. Determination of the complete primary structure of the V region of the H chain of Mot identified two papain cleavage sites in the second complementarily-determining region (CDR). Amino acid sequence of the cleavage sites was .**GRAPHIC**. (.dwnarw. = cleavage site). In the vicinity of these cleavage sites, many hydrophilic and polar residues are present and the predicted secondary structure near these cleavage sites suggested that this region was exposed on the surface of the molecule, and that the unusual papain cleavage of the IgG Mot might be caused by a unique conformation of the molecule, making it highly susceptible to enzyme digestion.

关键词： H heavy L light V variable C constant CDR complementarity-determining region fR framework region PeCys S -pyridylethylcysteine PTH phenylthiohydantoin HPLC high-performance liquid chromatography M r molecular mass PCA pyrrolidone carboxylic acid A alanine B aspartic acid or asparagine C cysteine D aspartic acid E glutamic acid f phenylalanine G glycine H histidine I isoleucine K lysine L leucine M methionine N asparagine P proline Q glutamine R arginine S serine T threonine V valine W tryptophan Y tyrosine Z PCA

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BINDING Of NITROPYRENES AND BENZO[A]PYRENE TO MOUSE LUNG DEOXYRIBONUCLEIC-ACID AfTER PRETREATMENT WITH INDUCING AGENTS

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BIOCHEMICAL PHARMACOLOGY 1986年第13期35卷 2129-2134页

作者： HOWARD, AJ MITCHELL, CE DUTCHER, JS HENDERSON, TR MCCLELLAN, RO LOVELACE BIOMED & ENVIRONM RES INST INHALAT TOXICOL RES INST ALBUQUERQUE NM 87185 USA

In assessing the biological effects of exposure to a complex chemical mixture, it is important to determine how the behavior of one compound may be influenced by the presence of other compounds in the mixture. In this study the effect of pre-exposure to an organic extract of diesel exhaust or to selected compounds in diesel exhaust on the binding of diesel exhaust compounds to DNA was determined. The amount of radiolabel covalently bound to mouse lung DNA following intratracheal administration of radiolabeled benzo[a]pyrene (BaP), 1-nitropyrene, 1,3,6-trinitropyrene, or a mixture of dinitropyrene was determined following pretreatment with benzo[a]pyrene, 1-nitrophrene, and diesel exhaust extract. Male CD-1 mice, 15-18 weeks of age, received 10 mg/kg of putative inducing agents by intratracheal instillation and, after 24 hr, 0.03 to 1.2 mg/kg radiolabeled putative DNA binding agents. Lung DNA was extracted, and covalent binding was quantitated by liquid scintillation spectroscopy. 1-Nitropyrene was a potent lung DNA binding agent in the absence of inducing agents [Covalent Binding Index (CBI) = 970] and was extremely potent after benzo[a]pyrene pretreatment (CBI = 21,540, comparable to the CBI for aflatoxin B1). Similar results were obtained for DNA binding of dinitropyrene and trinitropyrene with and without BaP pretreatment. DNA binding of BaP was lower (CBI = 40) and less inducible (BaP-pretreatment CBI = 230). Pretreatment with diesel extract caused an elevation in the binding of benzo[a]pyrene but little or no elevation in the binding of the nitropyrenes. Pretreatment with 1-nitropyrene did not increase significantly DNA binding of any of the agents tested. These results indicate that nitropyrenes bind readily to lung DNA and this binding may be increased in the presence of respirable mixtures, especially those containing inducing agents such as BaP.

关键词： AHH aryl hydrocarbon hydroxylase BaP benzo[ a ]pyrene CBI Covalent Binding Index HPLC high-performance liquid chromatography ITRI Inhalation Toxicology Research Institute R f relative migration in a TLC experiment

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PROTEIN-PHOSPHORYLATION AS A CONTROL fOR EXCITATION-ENERGY TRANSfER IN RHODOSPIRILLUM-RUBRUM

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fEBS LETTERS 1986年第1期200卷 144-148页

作者： HOLMES, NG ALLEN, Jf Department of Plant Sciences University of Leeds Leeds LS2 9JT England

Changes in cooperativity of the light-harvesting system of Rhodospirillum rubrum were monitored in whole cells by measurements of in vivo bacteriochlorophyll fluorescence yield transients. Under non-cooperative conditions a polypeptide of 10.5 kDa (provisionally identified as B880-β) was specifically phosphorylated; under cooperative conditions a polypeptide of 13 kDa (provisionally identified as B880-α) was specifically phosphorylated. These results are discussed in relation to the regulation of excitation energy distribution in purple photosynthetic bacteria and other photosynthetic organisms.

关键词： Photosynthesis Purple bacteria Light-harvesting complex Photosynthetic unit Protein phosphorylation Excitation transfer Bacteriochlorophyll fluorescence f o fluorescence yield when all photochemical traps are open f m fluorescence yield when all photochemical traps are closed f t fluorescence yield at time, t LHC-II light-harvesting chlorophyll a/b pigment-protein complex 1

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