bluuebunny/crossref_metadata_2025 huggingface dataset:
The amino acid sequence of the enterotoxin from Clostridium perfringens type A
Authors: Richardson Michael , Granum Per Einar
Published: July 2002

The amino acid sequence of the enterotoxin from Clostridium perfringens type A was determined by analysis of peptides derived from the protein by digestion with trypsin chymotrypsin, thermolysin, pepsin, a lysinespecific protease, S. aureus V8 protease and a proline‐specific protease, and fragments generated by cleavage with cyanogen bromide or by dilute acetic acid in 7 M guanidine HCl. The sequence which is complete except for the definite order of 3 small peptides between residues 88 and 103 consists of 309 amino acids and contains a correction to our preliminary announcement [(1984) FEMS Symp. 24, 329‐330].
Rapid in vitro desensitization of the testosterone response in rat Ley dig cells by sub‐active concentrations of porcine luteinizing hormone
Authors: Guillou Florian , Martinat Nadine , Combarnous Yves
Published: July 2002

We have studied in rat Leydig cells, the effect of sub‐active concentrations of porcine LH on the subsequent stimulation of the cAMP and testosterone production by a sub‐maximal concentration of pLH or hCG. We found that extremely low concentrations of pLH (0.01–2.0 ) were able to induce rapidly a partial but highly significative desensitization of the testosterone response without affecting the cyclic AMP response. These data indicate that desensitization of the steroidogenic response might be due to some lesion beyond cAMP formation or at the level of one discrete compartment of cyclic AMP, directly involved in the control of steroidogenesis. Moreover, our data strongly suggest that the basal circulating concentrations of LH can exert an inhibitory control on the testosterone response to LH pulses in vivo.
Cloning of the β‐phycocyanin gene from Anacystis nidulans
Authors: Lind Lisbet K. , Kalla S.Roger , Lönneborg Anders , Öquist Gunnar , Gustafsson Petter
Published: July 2002

The β‐phycocyanin gene, pcyB, of Anacystis nidulans was isolated from an Escherichia coli λ‐phage bank by the use of synthetic oligonucleotides derived from the 170 amino acid sequence of the β‐phycocyanin protein. Two positive, overlapping λ‐clones were found. Partial DNA sequencing of one of the clones gave a deduced amino acid sequence which was in full agreement with a portion of the published sequence of A. nidulans β‐phycocyanin. A comparison with the published DNA sequence for β‐phycocyanin from Agmenellum quadruplicatum shows a DNA sequence homology of 70.4% over the sequenced region.
An oxygen‐evolving complex with a simple subunit structure ‐ ‘a water‐plastoquinone oxidoreductase” ‐ from the thermophilic cyanobacteriumSynechococcussp.
Authors: Satoh Kazuhiko , Ohno Takashi , Katoh Sakae
Published: July 2002

An oxygen‐evolving complex has been highly purified from the thermophilic cyanobacteriumSynechococcussp. The complex, which reproducibly showed 5 major polypeptide bands of 47, 40, 35, 30 and 9 kDa on SDS‐polyacrylamide gel electrophoresis and contained 3.2 Mn per QA, had an oxygen‐evolving activity of 300–400chl per h in the presence of 5 mM MnCl2; or CaCl2. The complex most likely represents a minimum functional unit of the photosynthetic oxygen evolution.
Modification of photosystem II activity by protein phosphorylation
Authors: Hodges M. , Packham N.K. , Barber J.
Published: July 2002

Phosphorylation of proteins within pea thylakoid membranes decreases photosystem II (PSII) mediated electron transfer at saturating light intensities which, according to changes in room temperature chlorophyll fluorescence transients, is due to a modification in the electron transfer from QAto QB. However, a previously reported increase in the ability of DCMU to inhibit PS2 electron flow to DCPIP, as a consequence of protein phosphorylation, was not observed, although changes in DCMU efficacy were found to depend upon the redox state of the plastoquinone (PQ) pool.
Resonance energy transfer evidence for two attached states of the actomyosin complex
Authors: Bhandari Dhan G. , Trayer Hylary R. , Trayer Ian P.
Published: July 2002

Resonance energy transfer measurements were made between a donor fluorophore, N‐(bromacetyl)‐N'‐(1‐sulpho‐5‐naphthyl) ethylenediamine, located on the single cysteine of the Al light chain of myosin S1(A1), and an acceptor fluorophore, 5‐(iodoacetamido)fluorescein, sited on Cys‐374 of actin. In the binary rigor complex a transfer efficiency of 24% was noted, representing a spatial separation of about 6 nm. When the same measurements were made using a stable analogue of S1·ATP, in which the fast reacting SH1 thiol group is crosslinked to another thiol group in the 20 kDa domain of S 1, the 2 fluorophores were found to have moved closer together by ⩾ 3 nm. This provides, for the first time, direct experimental evidence for a change in structure of the myosin crossbridge that could account for tension generation.
Association of the 33‐kDa polypeptide with the 43‐kDa component in photosystem II particles
Authors: Isogai Yasuhiro , Yamamoto Yasusi , Nishimura Mitsuo
Published: July 2002

The organization of polypeptides and Mn atoms in the oxygen‐evolution system was studied by analyzing the effects of protease treatment on photosystem (PS) II particles from spinach chloroplasts. Hydrolysis of the 47‐ and 43‐kDa polypeptides of the PS II core complex had similar profiles of dependence on trypsin concentration in the treatment of control PS II particles and NaCl‐washed particles, which were devoid of the peripheral 24‐ and 18‐kDa polypeptides. In CaCl2‐washed particles, which were depleted of the 33‐kDa polypeptide besides the 24‐ and 18‐kDa polypeptides, the 43‐kDa polypeptide was much more sensitive to trypsin than the 47‐kDa polypeptide. Chymotrypsin treatment gave similar results. These findings suggest that the 33‐kDa polypeptide is associated to the 43‐kDa polypeptide and shields it from tryptic attack. Changes of the amount of Mn in the PS II and salt‐washed particles on protease treatment indicated a heterogeneity of binding sites of Mn in the PS II particles
Primary structure of a cationic Cu,Zn Superoxide dismutase
Authors: Schininà M.E. , Barra D. , Gentilomo S. , Bossa F. , Capo C. , Rotilo G. , Calabrese L.
Published: July 2002

The complete amino acid sequence of Cu,Zn Superoxide dismutase from sheep erythrocytes has been determined. The sequence is very similar to that of the bovine enzyme, having the same number of residues (151) and only two substitutions in the ‘hypervariable’ region (residues 17–30). The 5 overall substitutions confer a positive charge on the sheep enzyme at neutral pH (pI≈8). This charge is localized outside the active site region. The catalytic efficiency of the sheep enzyme is 15% less than that of the cow enzyme, confirming the hypothesis that the enzyme activity is related to the concentration of positive surface charge near the active site channel.
Guanine nucleotide‐binding protein in sea urchin eggs serving as the specific substrate of islet‐activating protein, pertussis toxin
Authors: Oinuma Masayuki , Katada Toshiaki , Yokosawa Hideyoshi , Ui Michio
Published: July 2002

A GTP‐binding protein serving as the specific substrate of islet‐activating protein (IAP), pertussis toxin, was partially purified from Lubrol extract of sea urchin egg membranes. The partially purified protein possessed two polypeptides of 39 and 37 kDa; the 39 kDa polypeptide was specifically ADP‐ribosylated by IAP and the 37 kDa protein cross‐reacted with the antibody prepared against purified βγ‐subunits of αβγ‐heterotrimeric IAP substrates from rat brain. Incubation of this sea urchin IAP substrate with a non‐hydrolyzable GTP analogue resulted in a reduction of the apparent molecular mass on a column of gel filtration as had been the case with purified rat brain IAP substrates, suggesting that the sea urchin IAP substrate was also a heterooligomer dissociable into two polypeptides in the presence of GTP analogues. Thus, the 39 and 37 kDa polypeptides of the sea urchin IAP substrate correspond to the α‐ and β‐subunits, respectively, of mammalian IAP substrates which are involved in the coupling between membrane receptor and effector systems.
TSH‐induced cyclic AMP production in an ovine thyroid cell line: OVNIS 5H
Authors: Fayet Guy , Aouani Azédine , Hovsépian Sonia
Published: July 2002

The TSH‐induced cyclic AMP response was studied using a 3‐year‐old ovine thyroid cell line TSH‐independent for growth: OVNIS 5H. The kinetics of cyclic AMP production was followed both in cell layers and in cell culture media, with or without phosphodiesterase inhibitor. It is noteworthy that following the first wave in cyclic AMP obtained within minutes, we observed later a sustained exponential increase in cyclic AMP during the 5 days following TSH stimulation. A bioassay of TSH was derived allowing measurement of 1 TSH from a crude bTSH preparation.
Comparison of the structure of turtle pancreatic ribonuclease with those of mammalian ribonucleases
Authors: Beintema Jaap J. , van der Laan Jan M.
Published: July 2002

There are 33 invariant amino acid positions out of 132 positions in 42 investigated sequences of ribonucleases from a number of mammalian species and a reptile (snapping turtle, Chelydra serpentina). These invariant residues are unequally distributed over 3 different parts of the molecule. The lobe of the S‐protein part of the molecule, which lacks one disulfide bridge and has two shortened loops in turtle ribonuclease, has the lowest percentage of invariant residues, although the active‐site residue His 119 is located in this part.
Destabilization by high pH of the S1‐state of the oxygen‐evolving complex in photosystem II particles
Authors: Plijter Johan J. , de Groot Alex , van Dijk Minse A. , van Gorkom Hans J.
Published: July 2002

The S‐state distribution in a dark‐adapted photosystem II preparation, isolated from spinach chloroplasts, can be changed by pH. The period 4 oscillation of the oxygen‐releasing reaction, as measured by UV absorbance changes, is shifted by one flash at pH 8.3. This is not due to the generation of the EPR signal II on the first flash, or to the loss of a positive charge to some other electron donor after the first flashes. It is concluded that at pH 8.3 the S1‐state is reduced to S0. The half‐time of the reduction of S1 to S0 is 45 s, and the reduction is reversed in the dark if the pH is readjusted to pH 6.0. The oxygen release at pH 8.3 is irreversibly slowed down to about 3.2 ms.
Thyrotropin and cyclic AMP regulation of ras proto‐oncogene expression in cultured thyroid cells
Authors: Dere Willard H. , Hirayu Hideshi , Rapoport Basil
Published: July 2002

We examined the effect of thyrotropin (TSH) on intracellular levels of c‐ras mRNA in a line of differentiated rat thyroid cells obtained from normal Fischer rat thyroids. These cells are totally dependent on TSH for growth. TSH stimulation of quiescent cells increased c‐ras mRNA content, with a maximal response (730% of basal) after 6 h, and a decline towards basal levels after 24 h. Dibutyryl cAMP and forskolin mimicked this stimulatory effect of TSH on c‐ras, but did not enhance β‐actin mRNA content. This study demonstrates hormonal and cyclic nucleotide control of c‐ras expression in a well‐differentiated, non‐tumorogenic mammalian cell.
Complete amino acid sequence of 33 kDa protein isolated from spinach photosystem II particles
Authors: Oh-oka H. , Tanaka S. , Wada K. , Kuwabara T. , Murata N.
Published: July 2002

The amino acid sequence of the 33 kDa protein isolated as an extrinsic protein from spinach photosystem II particles was determined by using solid‐phase sequencing and conventional procedures. The 33 kDa protein was found to be composed of 248 amino acid residues, to lack histidine and to have a molecular mass of 26 663 Da, which was considerably smaller than the value deduced from SDS‐polycrylamide gel electrophoresis. The sequence of the 33 kDa protein was compared with those of the bacterial Superoxide dismutases (SOD) with Mn atoms at an active site. A part of the sequence of the 33 kDa protein was similar to a region in Mn‐SODs from Bacillus stearothermophilus and Escherichia coli, which was expected to be the Mn‐binding site.
The chlorophylla/b‐proteins of PS I and PS II are immunologically related
Authors: Evans Paula K. , Anderson Jan M.
Published: July 2002

Polyclonal antisera were raised against three apoproteins of the light‐harvesting chlorophylla/b‐protein complex of photosystem I (LHC I) from spinach. These antisera were found to cross‐react with all four LHC I and both LHC II apoproteins on immunoblots. Affinity purification of the cross‐reactive antibodies on either the ‘native’ LHC II complex or on the 28 kDa apoprotein of LHC II immobilized on nitrocellulose yielded antibodies which recognize the same apoproteins as the crude sera. Partial peptide mapping of LHC I and LHC II apoproteins withStaphylococcus aureusV8 protease andN‐chlorosuccinimide showed different patterns for each polypeptide. These data demonstrate that the four polypeptides of LHC I are distinct from each other and from the two major polypeptides of LHC II but all six are immunologically related.
Acyclic model peptides with ionophoretic activity
Authors: Shastri B.P. , Rehse P.H. , Attah-Poku S.K. , Ananthanarayanan V.S.
Published: July 2002

Two linear synthetic peptides, N‐tBoc‐Pro‐Gly‐Ala‐NHCH3 and N‐tBoc‐Pro‐D‐Ala‐Ala‐NHCH3, have previously been shown by us to complex with Ca2+ and form 2:1 (peptide:calcium) complexes. Here we report their binding to Pr3+ and demonstrate, by 1H NMR, the peptide‐mediated transport of Pr3+ across dimyristoylphosphatidylcholine unilamellar vesicles via a 2:1 ion‐sandwich complex.
Blood glucose and liver glycogen in the rat
Authors: Kaminsky Yury G. , Kosenko Elen A.
Published: July 2002

In rats, chronic ethanol consumption results in moderate hypoglycemia with no great changes in the diurnal patterns of blood glucose and hepatic glucose. Marked alterations of the diurnal rhythm of liver glycogen were observed in ethanol‐fed animals, the phase of the rhythm being shifted and the daily mean and minimum levels being lowered. After withdrawal of ethanol consumption, blood glucose remained at a nearly constant level for 18 h, but then decreased sharply causing the liver/blood gradient of glucose to increase up to 4. Liver glycogen varied with a small amplitude and a nearly inverted phase in withdrawn rats as compared to controls.
Reconstitution of photosynthetic water‐splitting activity by the addition of 33 kDa polypeptide to urea‐treated PS II reaction center complex
Authors: Tang Xiao-Song , Satoh Kimiyuki
Published: July 2002

Photosystem IIReaction centerReconstitutionUrea treatmentManganese binding33 kDa polypeptideOxygen evolution
Regulation of cholesterol ester hydrolase by cyclic AMP‐dependent protein kinase
Authors: Colbran Roger J. , Garton Andrew J. , Cordle Susan R. , Yeaman Stephen J.
Published: July 2002

Cholesterol ester hydrolase
Phosphorylation
Steroidogenesis
Onset of ATP synthesis in spinach chloroplasts after single‐turnover flashes in relation to adenylate kinase activity
Authors: Peters R.L.A. , van Kooten O. , Vredenberg W.J.
Published: July 2002

Flash‐induced ATP synthesis was investigated in chloroplasts isolated from spinach varieties with a pronounced difference in their adenylate kinase activity. In chloroplasts with high adenylate kinase activity the first 4 flashes induced ATP hydrolysis which persisted for an extended dark period at which the membrane remained energized. In chloroplasts showing low adenylate kinase activity, ATP hydrolysis could only be induced by a successive flash train of at least 60 flashes or by a 30–60 s period of continuous pre‐illumination. The activity of the adenylate kinase must be considered when the kinetics of initial ATP synthesis and of P515 in light flashes are studied.
Putative ammo‐terminal presequence for β‐subunit of plant mitochondrial F1ATPase deduced from the amino‐terminal sequence of the mature subunit
Authors: Kobayashi Kazuto , Iwasaki Yukimoto , Sasaki Takuji , Nakamura Kenzo , Asahi Tadashi
Published: July 2002

The α‐ and β‐subunits of sweet potato mitochondrial F1ATPase were purified from the F1 complex by gel filtration and ion‐exchange high‐performance liquid chromatography. Isoelectric focussing and N‐terminal amino acid sequencing indicated that the purified β‐subunit contains at least two polypeptides similar to each other. The N‐terminal 18 amino acid sequence of the β‐subunit showed homology to the amino acid sequence of the tobacco mitochondrial F1ATPase β‐subunit precursor deduced from the nucleotide sequence [(1985) EMBO J. 4, 2159‐2165] between residues 56 and 73, suggesting that the N‐terminal 55 amino acids of the tobacco precursor constitute the presequence required for mitochondrial targetting.
Novel aryl ether rearrangement catalyzed by lignin peroxidase of Phanerochaete chrysosporium
Authors: Miki Keiji , Renganathan V. , Gold Michael H.
Published: July 2002

Lignin peroxidase catalyzed the rearrangement of 2‐(2́‐methoxy‐4́‐propylphenoxy)‐1,3‐dihydroxypropane I to 3‐(2́‐methoxy‐4́‐propylphenoxy)‐1,2‐dihydroxypropane VI with a maximum product yield of 64%. The reaction was strongly inhibited by oxygen. The enzyme also catalyzed the rearrangement of 2‐(2́‐methoxy‐4́‐propylphenoxy) ‐1‐hydroxypropane IV to form 1‐(2́‐methoxy‐4́‐propylphenoxy)‐2‐hydroxypropane VIII. When IV was labeled with 18O in the 1‐hydroxyl group, all of the 18O was retained in the ether oxygen of the product VIII. A mechanism explaining these results involves: (i) one electron oxidation of the substrate to form an aryl cation radical; (ii) nucleophilic attack by the hydroxyl oxygen on the cation radical with release of a proton, resulting in a dioxolane radical intermediate; (iii) cleavage of the latter and subsequent hydrogen abstraction to form the rearranged product.
Site of action of a halogenated 4‐hydroxypyridine on ferredoxin‐catalysed cyclic photophosphorylation
Authors: Patel Pravin K. , Bendall Derek S. , Ridley Stuart M.
Published: July 2002

Tetrabromo‐4‐hydroxypyridine (J820) inhibited ferredoxin‐catalysed cyclic photophosphorylation at micromolar concentrations but did not inhibit or uncouple the AQS‐catalysed system. At 2 μM it did not abolish the slow phase of the electrochromic shift or affect the turnover of cytochromesb‐563 andf. At higher concentrations (10 μM) it decreased the rate of re‐reduction of cytochromef, whilst inhibiting the reduction of cytochromeb‐563. It is concluded that tetrabromo‐4‐hydroxpyridine does not bind to the quinone reduction site of the cytochromebfcomplex, but inhibits the putative ferredoxin‐plastoquinone reductase.
Control of tubulin gene expression during metacyclogenesis of Trypanosoma cruzi
Authors: Rondinelli Edson , de Moura-Neto Rodrigo Soares , Suva Rosane , de Almeida Soares Célia Maria , Carvalho José Francisco , de Castro Firmino Torres
Published: July 2002

During differentiation of the dividing epimastigote to the non‐dividing metacyclic trypomastigote form of the parasitic protozoan Trypanosoma cruzi there is a marked reduction in the rate of synthesis of the major proteins α‐ and β‐tubulin. Our results indicate that the control of synthesis of these proteins during the differentiation event is exerted at the level of α‐ and β‐tubulin mRNA accumulation.
Protonation of a novel intermediate P is involved in the M → bR step of the bacteriorhodopsin photocycle
Authors: Drachev L.A. , Kaulen A.D. , Skulachev V.P. , Zorina V.V.
Published: July 2002

A novel intermediate (P) of the bacteriorhodopsin (bR) photocycle, appearing between M412 and bR is described. Like bR, intermediate P shows an absorption maximum at 560–570 nm. However, the extinction coefficient of P is somewhat lower than that of bR. Moreover, there are some differences in spectra of bR and P at wavelengths shorter than 450 nm. The P → bR transition correlates with the absorption of H+ from the water medium. The following conditions proved to be favourable for the detection of the new intermediate: a high salt concentration, low light intensity and low temperature (0.5°C). The P → bR transition is strongly decelerated by a small amount of Triton X‐100. Illumination of P does not produce M412 before bR is formed. It is assumed that M412 converts to P when the Schiff base is protonated by a proton transferred from a protein protolytic group which participates in the inward H+‐conductivity pathway. Reprotonation of this group results in the conversion of P to bR. No more than 1 H+ is transported per bR photocycle.
An energy‐dependent, transient peak in the minute range decay of luminescence, present in CO2‐accumulating cells of Scenedesmus obliquus
Authors: Sundblad L.-G. , Palmqvist K. , Samuelsson G.
Published: July 2002

Photosynthetic O2 evolution in the green alga, Scenedesmus obliquus, was shown to be more sensitive to the uncoupler FCCP when assayed in a low Ci medium than in a high Ci medium, indicating the action of an energy‐dependent mechanism for Ci uptake. Low Ci adapted algae exhibited characteristic luminescence decay kinetics with a transient peak 20–60 s after excitation. This peak was abolished by addition of FCCP and HCO−
3. The effect caused by HCO−
3 was partially reversed by methyl viologen. In view of the results obtained, a model is presented in order to discuss the origin of the transient luminescence peak.
Ca2+ and phorbol ester synergistically induce HL‐60 differentiation
Authors: Tyers Michael , Harley Calvin B.
Published: July 2002

Exposure of HL‐60 cells to subthreshold concentrations of TPA caused monocytic differentiation only when cells were cotreated with the Ca2+ ionophore A23187. Phorbol ester dose‐response curves for growth arrest and enzymatic markers of differentiation were shifted to lower concentrations when the ionophore was present. Expression of a monocyte/granulocyte cell surface antigen also occurred only when cells were treated with both agents. Similar effects were seen with other active but not inactive phorbol esters and with another Ca2+ ionophore. The Ca2+ component of phosphoinositide‐based signalling may thus play a role in HL‐60 differentiation.
Two species of phospholipase C isolated from lymphocytes produce specific ratios of inositol phosphate products
Authors: Carter Helen R. , Bird Ian M. , Smith Anthony D.
Published: July 2002

Chromatography of the soluble porcine lymphocyte phospholipase C on cellulose phosphate resolves the activity into two species. An HPLC method is described for separating the enzyme products. Ins 1,2 > P and Ins 1P. Use of these methods reveals that the two iso‐enzymes liberate, with a high degree of reproducibility, characteristic ratios of the two products. This suggests that the amount of each product produced is an inherent property of the enzyme mechanism.
Demonstration of mRNAs for oxytocin and prolactin in porcine granulosa and luteal cells
Authors: Einspainer Ralf , Pitzel Lutz , Wuttke Wolfgang , Hagendorff Gerd , Preuß Klaus-Dieter , Kardalinou Efterpi , Scheit Karl Heinz
Published: July 2002

The relative levels of mRNAs for relaxin, prolactin, inhibin and oxytocin have been measured in porcine granulosa as well as luteal cells by hybridisation to single‐stranded synthetic DNA. The likelihood of a paracrine function of oxytocin and prolactin in the porcine ovary was inferred from the invitro effects of both both hormones on progesterone secretion of ovarian cells. Both hormones were found to inhibit progesterone secretion of luteal cells. In contrast, only prolactin but not oxytocin stimulated progesterone secretion in granulosa cells.
Effect of γ‐butyrolactone derivative on an artificial membrane
Authors: Gräfe U. , Schlegel R. , Grigoriev P.A. , Römer W.
Published: July 2002

trans‐2‐(6'Methylheptanol‐1'‐yl)‐3‐hydroxymethyl‐4‐butanolide(II) was shown to increase the passive flux of cations such as Co2+ through a black lipid membrane made from ox brain phospholipids. This membranotropic effect appears to be involved in the activity of II towards blocked mutants of streptomycetes as an autoregulator of cytodifferentiation.
Interaction of chlorophyll a′ with the 65 kDa subunit protein of photosystem I reaction center
Authors: Hiyama Tetsuo , Watanabe Tadashi , Kobayashi Masami , Nakazato Masataka
Published: July 2002

The 65 kDa polypeptide subunit depleted of P700 was prepared from a photosystem I reaction center preparation and mixed with chlorophyll a′ (C‐10 epimer of chlorophyll a) to yield a complex exhibiting a tripleheaded spectrum with absorbance maxima at 673, 692 and 707 nm. The difference spectra (oxidized‐minus‐untreated and light‐minus‐dark) had a major trough at 707 nm and minor ones at 690 and 430 nm. The overall shape of the spectra resembled well that of P700 with a small red shift. A rapidly decaying flash‐induced absorbance change was observed at 430 nm with a half decay time of less than 500 μs in a preparation supplemented with an electron donor system.
Effect of echinomycin on DNA methylation
Authors: Adams R.L.P. , Rinaldi A.
Published: July 2002

Although echinomycin is reported to intercalate and to bind to DNA at CG dinucleotides, the effects of the drug on DNA methylation in vitro and in vivo are much less apparent than are the effects on DNA synthesis and cell growth.
Time‐dependent cGMP‐activated conductance of detached patches of ROS plasma membrane
Authors: Kolesnikov S.S. , Jainazarov A.B. , Fesenko E.E.
Published: July 2002

cGMP markedly increases the cationic conductance of the ‘inside‐out’ patches of rod outer segment plasma membrane when applied to the inner side. The cGMP‐activated conductance of some patches was shown to be time‐dependent. The data obtained suggest that the change of cGMP concentration in the near membrane layer underlies this phenomenon rather than the change in the channel's activity. The hydrolysis and, probably, the desorption of the nucleotide are responsible for this.
Surface enhanced resonance Raman scattering as a probe of the spin state of structurally related cytochromes P‐450 from rat liver
Authors: Kelly K. , Rospendowski B.N. , Smith W.E. , Wolf C.R.
Published: July 2002

Surface enhanced resonance Raman scattering (SERRS) was observed from structurally related drug‐induced rat liver cytochromes P‐450 adsorbed on a silver colloid. Careful control of pH and the sequence of addition of components to the sol is required to prevent protein denaturation at the surface due to conversion to P‐450's biologically inactive form P‐420 or haem loss. A low‐spin P‐450 (PB3a), a mixed low‐and high‐spin P‐450 (PB3b) and a predominantly high‐spin P‐450 (MC1a) were investigated. Spectra recorded in the 1300–1700 cm−1 frequency region, containing the oxidation state marker ν4 at 1375 cm−1 (Fe3+) and spin state markers ν10 (1625 cm−1, high‐spin; 1633 cm−1, low‐spin) and ν19 (1575 cm−1, high‐spin; 1585 cm−1, low‐spin) were used to differentiate between the spin states of the various forms of cytochrome P‐450. As well as the established spin state marker bands, the intensity of a band at 1400 cm−1 appeared to depend on the high‐spin content. Thus, with this method SERRS from silver colloids can be used to determine spin states of related cytochromes P‐450 in dilute solution (10−8 M) and may be of value in studies of protein‐substrate interactions.
The bcr‐c‐abl tyrosine kinase activity is extinguished by TPA in K562 leukemia cells
Authors: Alitalo Riitta
Published: July 2002

Tyrosine kinase activity is associated with the transforming potential of several oncogenes. Human chronic myeloid leukemia (CML) cells and cell lines have been shown to contain an active bcr‐c‐abl p210 tyrosine kinase as a consequence of the Philadelphia chromosomal translocation. In the present work the activity of the c‐abl and c‐src oncogene‐encoded tyrosine kinases was investigated during phorbol diester (TPA) induced differentiation of the K562 CML cells. The high tyrosine kinase activity of p210
bcr
‐c‐
abl
is strongly reduced during the initial 24 h of TPA treatment. In contrast, the activity of the c‐src tyrosine kinase is not changed. No change occurs in the expression of the c‐abl‐specific RNAs during this period. Following the reduction of bcr‐c‐abl kinase activity, cell proliferation is arrested and megakaryoblastic antigens appear on the cells. Sodium butyrate caused a slight decrease in growth rate and of bcr‐c‐abl kinase activity during erythroid differentiation whereas no changes in c‐src or c‐abl tyrosine kinase activities were seen in DMSO‐treated control cells.
Photooxidation products of primaquine Structure, antimalarial activity and hemolytic effects
Authors: Brossi Arnold , Gessner Wieslaw , Hufford Charles D. , Baker John K. , Homo Frederic , Millet Pascal , Landau Irène
Published: July 2002

Photooxidation of primaquine (1) and 5‐hydroxyprimaquine (5) afforded a blue dye for which o‐quinone structure 4 was elaborated. Similar oxidation of N‐ethoxyacetylprimaquine (10) afforded o‐quinone 11. Tissue schizontocidal activity of 4 and 11, and bisquinolylmethine 3 prepared earlier, showed that none of them had noteworthy antimalarial activity, but all three produced methemoglobin.
Experimental and theoretical study of gramicidin P, an analog of gramicidin A with a methylamine C‐terminal
Authors: Trudelle Y. , Daumas P. , Heitz F. , Etchebest C. , Pullman A.
Published: July 2002

Gramicidin P (a gramicidin A in which the ethanolamine C‐terminus is replaced by methylamine) was synthesized and shown to have the same single‐channel conductance behavior as gramicidin A. The results are discussed in connection with the energy profile computed in the presence of water in comparison with the corresponding profile for gramicidin A.
Characterisation of a PS II reaction centre isolated from the chloroplasts ofPisum sativum
Authors: Barber J. , Chapman D.J. , Telfer A.
Published: July 2002

A photosystem II reaction centre has been isolated from peas and found to consist of D1, D2 polypeptides and the apoproteins of cytochromeb‐559, being similar to that reported for spinach by Nanba and Satoh [(1987) Proc. Natl. Acad. Sci. USA 84, 109–112]. The complex binds chlorophylla, pheophytin and the haem of cytochromeb‐559 in an approximate ratio of 4:2:1 and also contains about one molecule of β‐carotene. It binds no plastoquinone‐9 or manganese but does contain at least one non‐haem iron. In addition to a light‐induced signal due to Pheo−seen under reducing conditions, a light‐induced P680+signal is seen when the reaction centre is incubated with silicomolybdate. In the presence of diphenylcarbazide, the P680+signal is partially inhibited and net electron flow to silicomolybdate occurs. This net electron flow is insensitive too‐phenanthroline, 3‐(3,4‐dichlorophenyl)‐1,1‐dimethyl urea and 2‐(3‐chloro‐4‐trifluoromethyl)anilino‐3,5‐dinitrothiophene but is inhibited by proteolysis with trypsin and by other treatments. Fluorescence, from the complex, peaks at 682 nm at room temperature and at 685 nm at 77 K. This emission is significantly quenched when either the P680+Pheo or P680Pheo−states are established indicating that the fluorescence emanates from the back reaction between P680+and Pheo−.
Stimulation of thylakoid energization and ribulose‐bisphosphate carboxylase/oxygenase activation in Arabidopsis leaves by methyl viologen
Authors: Salvucci Michael E. , Portis Archie R. , Heber Ulrich , Ogren William L.
Published: July 2002

The light responses of ribulose‐1,5‐bisphosphate carboxylase/oxygenase (rubisco) activation and light scattering, a measure of the transthylakoid ΔpH, were found to be similar in Arabidopsis leaves. Methyl viologen lowered the light requirements for both activation and scattering, indicating that rubisco activation is related to thylakoid energization status. Nigericin inhibited rubisco activation in spinach protoplasts, providing further evidence for an association between rubisco activation and thylakoid energization. Light scattering and fluorescence quenching signals indicated an overenergization of the thylakoids in an Arabidopsis mutant defective in light activation of rubisco.
A comparison between the multiline EPR signals of spinach andAnacystis nidulansand their temperature dependence
Authors: Aasa Roland , Andréasson Lars-Erik , Lagenfelt Göran , Vänngård Tore
Published: July 2002

The multiline EPR signals arising from manganese in the S2state of the oxygen‐evolving system of spinach and the cyanobacteriumAnacystis nidulanshave very similar properties and are affected identically by NH3, suggesting that the system is highly conserved. The temperature dependence of the signal amplitude follows Curie behavior down to sub‐helium temperatures. This is in contrast to previous reports, which were taken as evidence for a tetrameric manganese cluster. Thus, it seems that it is not yet possible from EPR data alone to distinguish between this model and a dimeric structure.
S‐configuration of (+)‐primaquine by X‐ray analysis of the urea obtained with R‐(+)‐1‐phenylethylisocyanate
Authors: Yu Qian-Sheng , Brossi Arnold , Flippen-Anderson Judith L.
Published: July 2002

The S‐configuration for (+)‐primaquine prepared from the racemate by chemical resolution was established by solid‐state X‐ray analysis of the (+)‐1‐phenylethylurea obtained with R‐(+)‐1‐phenylethylisocyanate.
Concurrent measurements of the inhibition of ATP synthesis and alanine transport in intact cells ofRhodobacter capsulatus
Authors: Crielaard W. , Elferink M.G.L. , Hellingwerf K.J. , Konings W.N. , Myatt J.F. , Jackson J.B.
Published: July 2002

Myxothiazol, antimycin A and carboxylcyanide‐m‐chlorophenylhydrazone (CCCP) all had a more pronounced inhibitory effect on the rate of ATP synthesis than on the rate of alanine transport during illumination of intact cells ofRhodobacter capsulatus. CCCP had the most potent effect and antimycin A the least. The data suggest that the energy‐input requirement for alanine transport is lower than that for ATP synthesis. A new test for direct interactions between the electron transport system and the ATP synthase and alanine transporter is applied to the data. Subject to reservations about the accuracy of rate measurement in intact bacterial cells and about the specificity of inhibition, the test reveals that the proton motive force is not the sole determinant of the rate of ATP synthesis and the rate of alanine transport.
Isolation and characterization of the intact gastrin precursor from a gastrinoma
Authors: Desmond H. , Pauwels S. , Varro A. , Gregory H. , Young Janice , Dockray G.J.
Published: July 2002

Antibodies to the extreme C‐terminal region of human progastrin have been used to monitor the isolation of high‐M
r immunoreactive material in a gastrinoma extract. Microsequence analysis of the product revealed amino acid residues in the first 18 positions corresponding to those predicted from the cDNA sequence for preprogastrin starting at position 22; the sequence and immunochemical data together allow the identification of this material as intact progastrin. Implications for gastrin biosynthesis are discussed.
Pertussis toxin‐dependent and ‐independent hormonal effects on cultured renal epithelioid cells
Authors: Paulmichl M. , Wöll E. , Lang F.
Published: July 2002

The present study has been performed to test for the involvement of pertussis toxin‐sensitive GTP‐binding proteins (G‐proteins) in the cellular transduction of hormone‐induced activation of potassium channels. In Madin Darby canine kidney (MDCK) cells, a permanent cell line from dog kidney, epinephrine, acetylcholine, bradykinin, serotonin and ATP hyperpolarize the cell membrane by activation of potassium channels. In cells pretreated with pertussis toxin the hyperpolarizations elicited by either acetylcholine or serotonin are completely abolished; that following epinephrine is blunted and only transient. The hyperpolarizing effects of ATP or bradykinin are not affected by pertussis toxin. Thus, in MDCK cells both pertussis toxin‐dependent and ‐independent mechanisms operate in parallel to enhance the potassium conductance of the cell membrane.
Evidence that a second stereochemical centre in diacylglycerols defines interaction at the recognition site on protein kinase C
Authors: Bonser Robert W. , Thompson Neil T. , Hodson Harold F. , Beams Richard M. , Garland Lawrence G.
Published: July 2002

The interaction of novel diacylglycerol analogues at the recognition site on protein kinase C has been evaluated using a modified [3H]phorbol dibutyrate binding assay and an established kinase activation assay. Studies with the 3‐methyl analogues of 1,2‐dihexanoyl‐sn‐glycerol have revealed a preferred stereochemical configuration at the C‐3 position. Other chemical modifications have extended existing structure/activity relationships by showing that carbamates and sulphonyl esters cannot substitute for carboxylate esters and that cyclic acyl groups are active. Thus, most, if not all of the functionalities in the diacylglycerol molecule are required for interaction at the receptor on protein kinase C. Stereochemical specificity is required at C2 and C3.
Insulin‐like growth factor 1 and insulin reduce epidermal growth factor binding to Swiss 3T3 cells by an indirect mechanism that is apparently independent of protein kinase C
Authors: Corps Anthony N. , Brown Kenneth D.
Published: July 2002

Insulin‐like growth factor 1 and insulin reduced the binding of 125I‐labelled epidermal growth factor (125I‐EGF) to Swiss 3T3 cells by 15–20% at 37°C, but not at 4°C. Scatchard analysis indicated that IGF‐1 and insulin affected the higher‐affinity component of EGF binding, an effect previously associated with the activation of protein kinase C. However, the inhibition of 125I‐EGF binding by IGF‐1 and insulin was increased, not reduced, when the cells were treated with high concentrations of phorbol esters to down‐modulate protein kinase C. We suggest that IGF‐1 and insulin activate a protein kinase with similar or overlapping specificity to that of protein kinase C.
Expression and electrophysiological identification of the receptor for bombesin and gastrin‐releasing peptide in Xenopus laevis oocytes injected with polyA+ RNA from rat brain
Authors: Meyerhof Wolfgang , Morley Steven D. , Richter Dietmar
Published: July 2002

The receptor for bombesin and the related peptide, gastrin‐releasing peptide (GRP) has been induced in frog oocytes by injection of polyA+ RNA from rat brain. The primed oocytes responded to peptides of the bombesin family (GRP, neuromedin C of bombesin) by showing dose‐dependent oscillations in membrane currents as recorded by the voltage‐clamp method. The induced membrane changes were suppressed when oocytes were pretreated with a bombesin‐receptor antagonist.
The transmembrane distribution of galactolipids in spinach thylakoid inside‐out vesicles is opposite to that found in intact thylakoids
Authors: Siegenthaler Paul-André , Sutter James , Rawyler André
Published: July 2002

The transmembrane distribution of galactolipids has been determined in spinach thylakoid inside‐out vesicles obtained by fragmentation in a Yeda Press followed by partition in an aqueous dextran‐polyethylene glycol two‐phase system. Using the lipase from Rhizopus arrhizus to digest selectively the galactolipids in the outer monolayer of the membrane, we have found the molar outside/inside distribution to be 42±1/58±1 for monogalactosyldiacylglycerol (MGDG) and 82±0.2/18±0.2 for digalactosyldiacylglycerol (DGDG). As expected, the transmembrane distribution of galactolipids in inside‐out vesicles was the opposite to that found in intact thylakoids which, under similar conditions, was found to be 62/38 for MGDG and 20/80 for DGDG. The reliability of the enzymatic approach to determine the transmembrane distribution and the relative arrangement of acyl lipids in the thylakoid membrane are discussed.
Calcium‐dependence of chromogranin A‐catecholamine interaction
Authors: Westermann R. , Stögbauer F. , Unsicker K. , Lietzke R.
Published: July 2002

Major components of the secretory organelle of bovine adrenal medullary cells, the chromaffin vesicles, are the acidic protein chromogranin A, catecholamines and Ca2+. The binding of Ca2+ to chromogranin A has been established. To study the interaction between chromogranin A and catecholamines and its dependence on Ca2+ we immobilized chromogranin A to a newly raised monoclonal antibody. It is shown that chromogranin A can bind (i) about 0.5 mol catecholamines per mol in a non‐calcium‐dependent manner and (ii) about 5 mol per mol in the presence of calcium. These results further support the notion that chromogranin A may act as a secretory granule‐condensing protein.
Oxidation of rat insulin II, but not I, leads to anomalous elution profiles upon HPLC analysis of insulin‐related peptides
Authors: Gross David , Skvorak Anne , Hendrick Grant , Weir Gordon , Villa-Komaroff Lydia , Halban Philippe
Published: July 2002

Rat insulin II, unlike rat insulin I and other non‐rodent insulins, contains a unique methionine residue at position B29. Reversed‐phase HPLC allows for separation of the two rat insulins, with insulin I typically eluting faster than insulin II. An anomalous peak of insulin immunoreactive material was found eluting even faster than insulin I following acid extraction of rat insulin‐producing cells. This early peak co‐eluted with [Met‐O
B29]insulin II suggesting that during cell extraction and subsequent purification steps, rat insulin II is subject to selective oxidation at MetB29. Such oxidation of rat proinsulin II affords improved separation from rat proinsulin I compared to the native form.
Evidence for the presence of two distinct sites of sucrose hydrolysis and glucosyl transfer activities on 1,3‐α‐D‐glucan synthase of Streptococcus mutans
Authors: Yamashita Y. , Hanada N. , Itoh-Andoh M. , Takehara T.
Published: July 2002

1,3‐α‐D‐Glucan synthase of Streptococcus mutans catalyzes both the hydrolysis of sucrose to glucose and fructose, and the glucosyl transfer to glucosyl polymers to yield water‐insoluble glucan. The enzyme catalyzes only sucrose hydrolysis, however, in the absence of 1,6‐α‐D‐glucan as an acceptor. In the present study, we found that glucosyl transfer activity was completely inhibited by the antiserum against isolated 1,3‐α‐D‐glucan synthase but that the sucrose hydrolysis activity was not. The antiserum did not impair the binding of the enzyme to the acceptor. These findings indicate that sucrose hydrolysis and glucosyl transfer occur at two distinct sites on the enzyme.
Chicken red‐sensitive cone visual pigment retains a binding domain for transducin
Authors: Fukada Yoshitaka , Okano Toshiyuki , Artamonov Igor D. , Yoshizawa Tôru
Published: July 2002

Iodopsin (a red‐sensitive cone visual pigment) and rhodopsin (a rod pigment) were isolated from chicken retina. They were separately reconstituted into phosphatidylcholine liposomes and then mixed with rod transducin (Tα and Tβγ) purified from bovine retina. Iodopsin enhanced, only when irradiated, the binding of GppNHp to Tα to a similar extent to irradiated rhodopsin. Furthermore, the binding of GppNHp to Tα in the presence of a photobleaching intermediate of iodopsin preferably required Tβγ‐2 rather than Tβγ‐1, which is very similar in profile to that in the presence of the intermediate of rhodopsin (J. Biol. Chem., in press). These results indicate that the binding domain for transducin in iodopsin should closely resemble that in rhodopsin.
Leupeptin does not affect the normal signal transduction mechanism in platelets
Authors: Alonso M.T. , Sanchez A. , Herreros B.
Published: July 2002

Calpains are Ca2+‐dependent serine proteases that can regulate protein kinase C‐mediated cellular events by cleaving the membrane‐bound native enzyme to yield an activated cytosolic fragment. Inhibition of calpain by leupeptin may cause enhancement or inhibition of cellular functions depending on the nature of the protein kinase C reaction involved. We have studied the effects of leupeptin on platelet responses (aggregation, secretion, thromboxane B2 formation and intracellular Ca2+ and pH changes) induced by either thrombin, collagen or phorbol 12‐myristate 13‐acetate (TPA), which are known to activate protein kinase C by different mechanisms. Only thrombin‐induced responses were inhibited by leupeptin. This suggests that the inhibitory effect of leupeptin is not due to antagonism of calpain in this system, but to direct interference with the proteolytic effect of thrombin.
High resolution emission spectra of one second delayed fluorescence from chloroplasts
Authors: Hideg Éva , Scott R.Q. , Inaba Humio
Published: July 2002

The first well resolved emission spectra of white light‐illuminated spinach chloroplasts at room temperature show that one second delayed fluorescence occurs at 685 nm. We demonstrate that reabsorption of this delayed fluorescence induces the second (probably prompt) emission observed at 730 nm and which we identify with the photosystem I peripheral antenna system.
Synthesis of spin‐labeled 2‐azido‐ATP: Evidence for distinct nucleotide‐binding sites in calcium pump protein from sarcoplasmic reticulum
Authors: Jakobs Pia , Sauer Hubertus E. , McIntyre J.Oliver , Fleischer Sidney , Trommer Wolfgang E.
Published: July 2002

A spin‐labeled and photoreactive derivative of ATP was synthesized with the spin label attached to the 2′‐ or 3′‐position of the ribose moiety and an azido group to C2 of the adenine ring (SL‐2N3‐ATP). Irradiation of this compound at 350 nm generates a nitrene, which then reacts with nucleophiles in its vicinity. SL‐2N3‐ATP, in the presence of Ca2+, was hydrolyzed by the calcium pump protein (Ca2+‐ATPase) of fast twitch skeletal muscle sarcoplasmic reticulum. The SL‐2N3‐ATP—enzyme complex in the absence of Ca2+ exhibited strongly immobilized ESR spectra. ESR spectra obtained after covalent incorporation of SL‐2N3‐ATP into Ca2+‐ATPase and removal of freely tumbling SL‐2N3‐ATP exhibited motionally constrained species indicative of distinct and possibly adjacent ATP‐binding sites. By contrast, with SL‐ATP devoid of the azido group or with the corresponding ‘non‐cleavable’ β,γ‐methylene triphosphate analogue (SL‐AMP‐PCP), two distinct sites were not as well resolved in the ESR spectra due to spectral overlap with the signal from the freely tumbling fraction even with the enhanced spectral resolution provided by perdeuteration of the spin label. Thus, SL‐2N3‐ATP may have general application for ESR studies of ATP‐dependent proteins under conditions in which noncovalent interactions are too weak for motionally restricted species to be resolved.
γ‐Thrombin‐induced phospholipase A2 activation in rabbit platelets: Comparison with α‐thrombin
Authors: Touqui Lhousseine , Jandrot-Perrus Martine , Vargaftig B.Boris
Published: July 2002

γ‐Thrombin stimulated release of [3H]arachidonic acid ([3H]AA) accompanied by a significant production of PAF and lyso‐PAF by rabbit platelets. These responses, which reflect PLA2 activation, were observed after a prolonged lag and to a lower extent when compared to those induced by α‐thrombin which evoked a much higher elevation in intracellular calcium. This elevation together with [3H]AA release were markedly reduced by EDTA. However, addition of ionophore A23187 enhanced the release of [3H]AA by γ‐thrombin to the levels similar to those of α‐thrombin. We conclude that γ‐thrombin is able to activate PLA2 and suggest that calcium influx may be a limiting factor for this activation.
Structural basis for the high activation energy of spectrin self‐association
Authors: Morris S.A. , Eber S.W. , Gratzer W.B.
Published: July 2002

The association of spectrin hetero‐dimer (αβ) to the tetramer (α2β2, which predominates in the cell) is marked by an exceptionally high activation energy, so that the reaction does not proceed measurably in the cold. We have tested the hypothesis that this is due to intra‐dimer association between the α‐ and β‐chain ends, which must be broken before tetramers can form. Two mutant univalent spectrins with association defects at the α and β ends, respectively, and incapable therefore of intra‐dimer bonding, were found to associate rapidly with one another at 4°C. The bimolecular rate constant is greater than for the association of normal dimers by 6 orders of magnitude.