bluuebunny/crossref_metadata_2025_split huggingface dataset:
The quinone‐reactive Ni/Fe‐hydrogenase of Wolinella succinogenes
Authors: DROSS Felix , GEISLER Viktor , LENGER Ruth , THEIS Friedbert , KRAFFT Torsten , FAHRENHOLZ Falk , KOJRO Elzbieta , DUCHÊNE Andrea , TRIPIER Dominique , JUVENAL Karin , KRÖGER Achim 
Published: March 2005

The hydrogenase (Hyd) isolated from the cytoplasmic membrane of Wolinella succinogenes consists of three polypeptides (HydA, HydB and HydC) and contains cytochrome b (6.4 μmol/g protein), which was reduced upon the addition of H2. The enzyme catalyzed the reduction of 2,3‐dimethyl‐1,4‐naphthoquinone with H2, in contrast to an earlier preparation which was made up of HydA and HydB only and did not contain cytochrome b (Unden, G., Böcher, R., Knecht, J. & Kröger, A. (1982) FEBS Lett. 145, 230–234). This suggests that HydC is a cytochrome b which serves as a mediator in the electron transfer from H2 to the quinone.The hydrogenase genes were cloned, sequenced and identified by sequence comparison with the N‐termini of the three subunits. The three genes were arranged in the order hydA, hydB, hydC, with the transcription start site in front of hydA, and were present only once on the genome. Separated by an intergene region of 69 nucleotides, hydC was followed by at least two more open reading frames of unknown function. The amino acid sequences derived from hydA, hydB and hydC were similar to those of the membrane Ni‐hydrogenases of seven other bacteria. HydA and HydB also showed similarity to the small and the large subunits of periplasmic Ni‐hydrogenases. HydC was predicted to contain four hydrophobic segments which might span the bacterial membrane. Two histidine residues located in hydrophobic segments are conserved in the corresponding sequences of the other membrane hydrogenases and might ligate the haem B.
Phenylalanine positively modulates the cAMP‐dependent phosphorylation and negatively modulates the vasopressin‐induced and okadaic‐acid‐induced phosphorylation of phenylalanine 4‐monooxygenase in intact rat hepatocytes
Authors: DØSKELAND Anne P. , VINTERMYR Olav K. , FLATMARK Torgeir , COTTON Richard G. H. , DØSKELAND Stein Ove 
Published: March 2005

The state of phosphorylation of phenylalanine hydroxylase was determined in isolated intact rat hepatocytes. 32P‐labeled phenylalanine hydroxylase was immunoisolated from cells loaded with 32Pi or from cell extracts ‘back‐phosphorylated’ with [γ‐32P]ATP by cAMP‐dependent protein kinase.The rate of phenylalanine hydroxylase phosphorylation in cells with elevated cAMP was similar to that observed for the isolated enzyme phosphorylated by homogeneous cAMP‐dependent protein kinase. The phosphorylation rate in cAMP‐stimulated cells was increased up to four times (reaching 0.018 s–1) by the presence of phenylalanine, the phosphate content (mol/mol hydroxylase) increasing to 0.5 from the basal level (0.17) in 50 s. The half maximal effect of phenylalanine was obtained at a physiologically relevant concentration (110 μM). The synthetic phenylalanine hydroxylase cofactor dimethyltetrahydropterin also enhanced the cAMP‐stimulated phosporylation of phenylalanine hydroxylase, presumably by displacing the endogenous cofactor, tetrahydrobiopterin.Phenylalanine was a negative modulator of the phosphorylation of phenylalanine hydroxylase induced by incubating cells with vasopressin or with the phosphatase inhibitor okadaic acid. The same site on the phenylalanine hydroxylase was phosphorylated in response to these two agents as in response to elevated cAMP. The available evidence suggested that not only vasopressin, but also okadaic acid, acted by stimulating the multifunctional Ca2+/calmodulin‐dependent protein kinase II or a kinase with closely resembling properties.
The binding of triazine herbicides to the photosynthetic reaction center of Rhodopseudomonas viridis
Authors: EGNER Ursula , HOYER Georg‐Alexander , SAENGER Wolfram 
Published: March 2005

The binding of six herbicides of the triazine family to the photosynthetic reaction center of Rhodopseudomonas viridis was investigated with energy‐minimization techniques, in order to correlate experimental with calculated data. The inhibitors were modeled in the active site according to the X‐ray structure analysis of the complex formed between the triazine terbutryn (2‐ethylamino‐4‐t‐butylamino‐6‐methylthio‐s‐triazine) and the reaction center of R. viridis [Michel, H., Epp, O. & Deisenhofer, J. (1986) EMBO J. 5, 2445–2451].40 different energy minimizations were carried out with varying cutoff radii, partial charges on inhibitor atoms and dielectric constants, i.e. 10 different combinations of these were tested. The impact of these parameters on the calculated binding and interaction energy was either examined for all protein/triazine complexes or, in the case of the dielectric constant, a smaller sample was used. The calculated energies are dominated by van der Waals interactions, which change by up to 20% when extending the cutoff radius from 0.8 nm to 1.5 nm. The use of uniform or distance‐dependent dielectric constant or partial charges on the inhibitor atoms does not severely influence the resulting structures, but shows a great impact on the calculated energies.In the two groups of triazines, each containing three inhibitors with methoxy or methylthio substituents, correlations of biological and calculated data were found quite often, but only once with all six triazines. The energy‐minimized structures were compared and analysed. A third hydrogen bond, not seen in the X‐ray analysis of the reaction center/terbutryn complex, was found between the t‐butylamino moiety of terbutryn (and equivalent moieties in the other triazines) and the carbonyl oxygen of TyrL222.
Differential tissue‐specific expression and induction of cytochrome P450IVA1 and acyl‐CoA oxidase
Authors: BELL David R. , BARS Remi G. , ELCOMBE Clifford R. 
Published: March 2005

We have examined the tissue‐specific expression and inducibility of acyl‐CoA oxidase and cytochrome P450IVA1 (P450IVA1) RNA in rats. Groups of three rats were dosed daily by gavage with methylclofenapate at 25 mg/kg in 5 ml/kg corn oil for nine weeks, or were administered a vehicle control. P450IVA1 and acyl‐CoA oxidase RNA were detected using an RNase protection assay. Similar levels of acyl‐CoA oxidase RNA were present in control liver and kidney, but the level of this RNA in lung, muscle and testis was 6–11%, and in pancreas was 0.13%, of that in liver. Treatment of rats with methylclofenapate led to an 11‐fold induction of acyl‐CoA oxidase RNA in liver and also produced a significant induction of this RNA in kidney, lung, muscle and testis of 1.7‐fold, 1.3‐fold, 2‐fold and 1.7‐fold, respectively. Acyl‐CoA oxidase RNA was not induced in pancreas.P450IVA1 RNA was present in control liver and also in kidney of control rats at 28% of the level in liver. In contrast to acyl‐CoA oxidase RNA, P450IVA1 RNA was not detected in lung, pancreas or testis. Methylclofenapate treatment of rats led to an 18‐fold induction of P450IVA1 RNA in liver, and a sevenfold induction in kidney. Induction of P450IVA1 was not detected in any of the other tissues examined. Quantification of the relative amounts of acyl‐CoA oxidase and P450IVA1 RNA in control liver revealed that acyl‐CoA oxidase RNA was present in a 17.5‐fold molar excess over P450IVA1 RNA.Western blotting with an anti‐P450IVA IgG revealed two bands of similar apparent molecular mass in liver and kidney microsomes, but not in microsomes from the testis of control rats. Methylclofenapate treatment of rats caused an increase in the intensity of these bands in microsomes from liver, but no induction was obvious in kidney. Immunocytochemical staining for both the microsomal P450IVA and peroxisomal acyl‐CoA oxidase proteins was restricted to the proximal convoluted tubule in the kidney cortex, with staining being most intense in the S3 region.
Multiple binding sites in fibronectin and the staphylococcal fibronectin receptor
Authors: BOZZINI Silvia , VISAI Livia , PIGNATTI Patrizia , PETERSEN Torben E. , SPEZIALE Pietro 
Published: March 2005

The binding of fibronectin to Staphylococci exhibits the properties of a ligand‐receptor interaction and has been proposed to mediate bacterial adherence to host tissues.To localize staphylococcal‐binding sites in fibronectin, the protein was subjected to limited proteolysis and, of the generated fragments, Staphylococci appeared to preferentially bind to the N‐terminal fragment. Different fibronectin fragments were isolated and tested for their ability to inhibit 125I‐fibronectin binding to Staphylococci. The results indicate that only the N‐terminal region effectively competed for fibronectin binding. However, when isolated fragments were adsorbed to microtiter wells, we found that two distinct domains, corresponding to the N‐terminal fragment and to the heparin‐binding peptide mapping close to the C‐terminal end of fibronectin, promoted the attachment of both Staphylococcus aureus Newman and coagulase‐negative strain of Staphylococcus capitis 651.These same domains were recognized by purified 125I‐labeled staphylococcal receptor, either when immobilized on microtiter wells or probed after adsorbtion onto nitrocellulose membrane.The heparin‐binding domain is comprised of type‐III‐homology repeats 14, 15 and 16. To determine which repeats participate in this interaction, we isolated and tested repeats type III14 and type III16. We found that the major staphylococcal‐binding site is located in repeat type III14.The staphylococcal receptor bound the N‐terminal domain of fibronectin with a KD of 1.8 nM, whereas the dissociation constant of the receptor molecule for the internal heparin‐binding domain was 10 nM.Since the fusion protein ZZ‐FR, which contains the active sequences of fibronectin receptor (D1‐D3) bound only to the N‐terminus, it is reasonable to assume that the bacterial receptor may have additional binding sites outside the D domains, capable of interacting with the internal heparin‐binding domain of fibronectin.
Lipoamide dehydrogenase from Azotobacter vinelandii
Authors: BENEN Jacques , van BERKEL Willem , VEEGER Cees , de KOK Arie 
Published: March 2005

The 10 C‐terminual residues are not visible in the crystal structure of lipoamide dehydrogenase from Azotobacter vinelandii, but can be observed in the crystal structures of the lipoamide dehydrogenases from Pseudomonas putida and Pseudomonas fluorescens. In these structures, the C‐terminus folds back towards the active site and is involved in interactions with the other subunit.The function of the C‐terminus of lipoamide dehydrogenase from A. vinelandii was studied by deletion of 5, 9 and 14 residues, respectively. Deletion of the last 5 residues does not influence the catalytic properties and conformational stability (thermoinactivation and unfolding by guanidinium hydrochloride). Removal of 9 residues results in an enzyme (enzyme 9) showing decreased conformational stability and high sensitivity toward inhibition by NADH. These features are even more pronounced after deletion of 14 residues (enzyme 14). In addition Tyr16, conserved in all lipoamide dehydrogenases sequenced thus far, and shown from the other structures to be likely to be involved in subunit interaction, was replaced by Phe and Ser. Mutation of Tyr16 also results in a strongly increased sensitivity toward inhibition by NADH. The conformational stability of both Tyr16‐mutated enzymes is comparable to enzyme 9. The results strongly indicate that a hydrogen bridge between tyrosine of one subunit (Tyr16 in the A. vinelandii sequence) and histidine of the other subunit (His470 in the A. vinelandii sequence), exists in the A. Vinelandii enzyme. In the 9 and 14 enzymes this interaction is abolished. It is concluded that this interaction mediates the redox properties of the FAD via the conformation of the C‐terminus containing residues 450–470.
Reaction of myeloperoxidase with its product HOCI
Authors: FLORIS René , WEVER Ron 
Published: March 2005

The reaction of human myeloperoxidase with its product, hypochlorous acid was investigated using both rapid‐scan spectrophotometry and the stopped‐flow technique. In the reaction of myeloperoxidase with hypochlorous acid a primary compound is found with propersties similar to that of compound I and which is converted into compound II. The primary reaction is strongly pH‐dependent. At pH 7.2 the reaction is too fast to be measured but at higher pH values it is possible to determine the apparent second‐order rate constant. Its value decreases to about 2 × 107 M−1· s−1 at pH 8.3 and to 2.3 (±0.4) × 106 M−1· s−1 at pH 9.2, respectively. The dissociation constant for the formation of the primary compound is 25.7 (± 15.3) μM at pH9.2 and about 2.5μM at pH 8.3. The apparent second‐order rate constant for the formation of compound II is hardly affected by pH and varies between 2 to 5 × 104 M−1· s−1 at pH 10.2 and pH 8.3 respectively.Reaction of myeloperoxidase with hypochlorous acid also resulted in irreversible partial bleaching of the chromophore. Chloride, which is a substrate of the enzyme not only protects myeloperoxidase against bleaching by hypochlorous acid but also competitively inhibits the binding of hypochlorous acid to myeloperoxidase, a process which also has been observed in the reaction with hydrogen peroxide. It is concluded that hypochlorous acid binds at the heme iron to form compound I.
The structure of pneumococcal lipoteichoic acid
Authors: BEHR Thomas , FISCHER Werner , PETER‐KATALINIĆ Jasna , EGGE Heinz 
Published: March 2005

Pneumococcal lipoteichoic acid was extracted and purified by a novel, quick and effective procedure. Structural analysis included methylation, periodate oxidation, Smith degradation, oxidation with CrO3, and fast‐atom‐bombardment mass spectrometry. Hydrolysis with 48% (by mass) HF and subsequent phase partition yielded the lipid anchor (I), the dephosphorylated repeating unit of the chain (II) and a cleavage product of the latter (III). The proposed structures are:

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where AATGal is 2‐acetamido‐4‐amino‐2,4,6‐trideoxygalactose, and all sugars are in the pyranose form and belong to the d‐series.Alkaline phosphodiester cleavage of lipoteichoic acid, followed by treatment with phosphomonoesterase, resulted in the formation of II and IV, with IV as the prevailing species:
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The linkage between the repeating units was established as phosphodiester bond between ribitol 5‐phosphate and position 6 of the glucosyl residue of adjacent units. The chain was shown to be linked to the lipid anchor by a phosphodiester between its ribitol 5‐phosphate terminus and position 6 of the non‐reducing glucosyl terminus of I. The lipoteichoic acid is polydisperse: the chain length may vary between 2 and 8 repeating units and variations were also observed for the fatty acid composition of the diacylglycerol moiety. Preliminary results suggest that repeating units II and IV are enriched in separate molecular species. All species were associated with Forssman antigenicity, albeit to a various extent when related to the non‐phosphocholine phosphorus. Owing to its unique structure, the described macroamphiphile may be classified as atypical lipoteichoic acid.
Role of F0 and F1 subunits in the gating and coupling function of mitochondrial H+‐ATP synthase
Authors: ZANOTTI Franco , GUERRIERI Ferruccio , CAPOZZA Giuseppe , FIERMONTE Maria , BERDEN Jan , PAPA Sergio 
Published: March 2005

A study is presented on the role of F0 and F1 subunits in oligomycin‐sensitive H+ conduction and energy transfer reactions of bovine heart mitochondrial F0F1 H+‐ATP synthase.Mild treatment with azodicarboxylic acid bis(dimethylamide) (diamide) enhanced oligomycin‐sensitive H+ conduction in submitochondrial particles containing F1 attached to F0. This effect was associated with stimulation of the ATPase activity, with no effect on its inhibition by oligomycin, and depression of the 32Pi—ATP exchange. The stimulatory effect of diamide on H+ conduction decreased in particles from which F1 subunits were partially removed by urea. The stimulatory effect exerted by diamide in the submitochondrial particles with F1 attached to F0 was directly correlated with a decrease of the original electrophoretic bands of a subunit of F0 (F0I‐PVP protein) and the γ subunit of F1, with corresponding formation of their cross‐linking product. In F0 liposomes, devoid of γ subunit, diamide failed to stimulate H+ conduction and to cause disappearance of F0I‐PVP protein, unless purified γ subunit was added back. The addition to F0 liposomes of γ subunit, but not that of α and β subunits, caused per se inhibition of H+ conduction.It is concluded that F0I‐PVP and γ subunits are directly involved in the gate of the F0F1 H+‐ATP synthase. Data are also presented indicating contribution to the gate of oligomycin‐sensitivity conferral protein and of another protein subunit of F0, F6.
Derived amino acid sequences of the nosZ gene (respiratory N2O reductase) from Alcaligenes eutrophus, Pseudomonas aeruginosa and Pseudomonas stutzeri reveal potential copper‐binding residues
Authors: ZUMFT Walter G. , DREUSCH Andreas , LÖCHELT Sabine , CUYPERS Heinrich , FRIEDRICH Bärbel , SCHNEIDER Beate 
Published: March 2005

The nosZ genes encoding the multicopper enzyme nitrous oxide reductase of Alcaligenes eutrophus H16 and the type strain of Pseudomonas aeruginosa were cloned and sequenced for structural comparison of their gene products with the homologous product of the nosZ gene from Pseudomonas stutzeri [Viebrock, A. & Zumft, W. G. (1988) J. Bacteriol. 170, 4658–4668] and the subunit II of cytochrome‐c oxidase (COII). Both types of enzymes possess the CuA binding site. The nosZ genes were identified in cosmid libraries by hybridization with an internal 1.22‐kb PstI fragment (NS220) of nosZ from P. stutzeri. The derived amino acid sequences indicate unprocessed gene products of 70084 Da (A. eutrophus) and 70695 Da (P. aeruginosa). The N‐terminal sequences of the NosZ proteins have the characteristics of signal peptides for transport.A homologous domain, extending over at least 50 residues, is shared among the three derived NosZ sequences and the CuA binding region of 32 COII sequences. Only three out of nine cysteine residues of the NosZ protein (P. stutzeri) are invariant. Cys618 and Cys622 are assigned to a binuclear center, A, which is thought to represent the CuA site of NosZ and is located close to the C terminus. Two conserved histidines, one methionine, one aspartate, one valine and two aromatic residues are also part of the CuA consensus sequence, which is the domain homologous between the two enzymes. The CuA consensus sequence, however, lacks four strictly conserved residues present in all COII sequences. Cys165 is likely to be a ligand of a second binuclear center, Z, for which we assume mainly histidine coordination. Of 23 histidine residues in NosZ (P. stutzeri), 14 are invariant, 7 of which are in regions with a degree of conservation well above the 50% positional identity between the Alcaligenes and Pseudomonas sequences. Conserved tryptophan residues are located close to several potential copper ligands. Trp615 may contribute to the observed quenching of fluorescence when the CuA site is occupied.
Molecular dynamics study of a complex between the human histocompatibility antigen HLA‐A2 and the IMP58‐66 nonapeptide from influenza virus matrix protein
Authors: ROGNAN Didier , ZIMMERMANN Norbert , JUNG Günther , FOLKERS Gerd 
Published: March 2005

The structure of the influenza‐virus‐matrix‐protein (IMP) 58‐66 nonapeptide, bound to the major‐histocompatibility‐complex‐encoded human leukocyte antigen (HLA) A2 protein was studied by molecular dynamics simulation. Starting from the extra electron density map of peptides co‐crystallized with HLA‐A2, the nonapeptide IMP58‐66 was docked residue by residue in the protein binding cleft. The complex was simulated for 100 ps in a shell of 1372 water molecules. The averaged simulated HLA‐A2 conformation was found to be similar to the crystal structure (0.182 nm RMS deviation, for the backbone atoms of the α1‐α2 domain). Nine out of the 14 hydrogen bonds observed in the antigen‐binding site were reproduced in the simulation.The IMP58‐66 peptide exhibits an extended conformation with kinks at positions 3 and 5. The side chains of residues 2, 3 and 9 develop van der Waals' interactions with hydrophobic pockets of HLA‐A2, corresponding to polymorphic residues of the major‐histocompatibility‐complex‐encoded proteins. Both the N‐terminus and C‐terminus of the nonapeptide were anchored in the antigen‐binding groove by hydrogen bonds with conserved amino acids. The N‐terminus was more flexible and contacts four HLA‐A2 conserved tyrosines (Tyr7, Tyr59, Tyr159 and Tyr171) and Glu63 by direct or water‐relayed hydrogen bonds. Water intercalation occurred only around the N‐terminus of the peptide, the C‐terminal carboxylate forming strong hydrogen bonds with polar residues (Tyr84 and Thr143) and a salt bridge with Lys146 all over the molecular dynamics simulation.This model is fully compatible with the recently published crystal structure of the HLA‐B27 protein, complexed by a mixture of self nonapeptides [15]
7‐Substituted pterins in humans with suspected pterin‐4a‐carbinolamine dehydratase deficiency
Authors: ADLER Carmen , GHISLA Sandro , REBRIN Igor , HAAVIK Jan , HEIZMANN Claus W. , BLAU Nenad , KUSTER Thomas , CURTIUS Hans‐Christoph 
Published: March 2005

A recently described new form of hyperphenylalaninemia is characterized by the excretion of 7‐substituted isomers of biopterin and neopterin and 7‐oxo‐biopterin in the urine of patients. It has been shown that the 7‐substituted isomers of biopterin and neopterin derive from L‐tetrahydrobiopterin and D‐tetrahydroneopterin and are formed during hydroxylation of phenylalanine to tyrosine with rat liver dehydratase‐free phenylalanine hydroxylase.We have now obtained identical results using human phenylalanine hydroxylase. The identity of the pterin formed in vitro and derived from L‐tetrahydrobiopterin as 7‐(1′,2′‐dihydroxypropyl)pterin was proven by gas‐chromatography mass spectrometry. Tetrahydroneopterin and 6‐hydroxymethyltetrahydropterin also are converted to their corresponding 7‐substituted isomers and serve as cofactors in the phenylalanine hydroxylase reaction. Dihydroneopterin is converted by dihydrofolate reductase to the tetrahydro form which is biologically active as a cofactor for the aromatic amino acid monooxygenases. The 6‐substituted pterin to 7‐substituted pterin conversion occurs in the absence of pterin‐4a‐carbinolamine dehydratase and is shown to be a nonenzymatic process. 7‐Tetrahydrobiopterin is both a substrate (cofactor) and a competitive inhibitor with 6‐tetrahydrobiopterin (Ki∼ 8 μM) in the phenylalanine hydroxylase reaction. For the first time, the formation of 7‐substituted pterins from their 6‐substituted isomers has been demonstrated with tyrosine hydroxylase, another important mammalian enzyme which functions in the hydroxylation of phenylalanine and tyrosine.