bluuebunny/crossref_metadata_2025_split huggingface dataset:
Circulardichroismus von heterodetischen cyclischen Cystinyl‐Tripeptiden
Authors: OTTNAD Michael , HARTTER Peter , JUNG Günther 
Published: March 2005

Circular Dichroism of Heterodetic Cyclic Cystinyl TripeptidesThe circular dichroism of two series of heterodetic cyclic tripeptides, t‐butoxycarbonyl‐l‐cysteinyl‐X‐cysteine disulfide methyl esters [X = Ala, Val, Leu, Phe, Glu(Obut), Lys(Z)] and l‐cysteinyl‐X‐cysteine disulfide methyl ester formates, and a corresponding series of t‐butoxycarbonyl‐l‐cysteinyl‐(glycyl)n‐cysteine disulfide methyl esters (n= 1–5) was investigated. The interaction between the disulfide group and the amide bonds results in a long‐wavelength Cotton effect at 300 nm. which exhibits a strong solvent dependence. In contrast to the long‐wavelength disulfide band and to the n‐π* amide Cotton effect a bathochromic shift is observed from less polar to strongly polar solvents. Within the series of N‐terminal protected tripeptide esters the influence of the central chiral amino acid X is small, but it increases in the non‐protected peptide esters. The circular dichroism is dominated by the influence of the conformation of the disulfide ring, which is essentially the same in both series. Due to interactions of the disulfide group with the amide chromophores, the sign of the long‐wavelength disulfide Cotton effect cannot be used for the determination of the helicity of the CSSC group. Infrared spectra of the protected compounds indicate trans‐peptide bonds in the solid state, which may also be true for the solutions.A pronounced time‐dependence is observed for the circular dichroism of t‐butoxycarbonyl‐l‐cysteinyl‐glycyl‐cysteine disulfide methyl ester after dissolving in ethanol. This is due to a uniform conformational change of the ring structure as shown by spectroscopic analysis using extinction and ellipticity diagrams.In the series of the cyclic cystinyl peptides containing glycyl residues a strong electronic disulfide‐amide interaction is observed for the tripeptide, being still observable for the tetra‐peptide. The comparison of the glycyl tripeptide with the tripeptides containing chiral central amino acids reveals a negative contribution to the circular dichroism at 260 nm due to these l‐residues.
Preparation and Specificity of Endonuclease IV Induced by Bacteriophage T4
Authors: BERNARDI Alberto , MAAT Jan , DE WAARD Adrian , BERNARDI Giorgio 
Published: March 2005

Bacteriophage‐T4‐induced endonuclease IV suitable for DNA sequence analysis has been prepared by a modified and easily reproducible method.The specificity of T4‐induced endonuclease IV has been investigated in order to verify whether this enzyme exhibits a single nucleotide recognition or a short sequence recognition.The 5′‐terminal dinucleotides and 3′‐terminal nucleotides of oligonucleotides released by T4 induced endonuclease IV from three single‐stranded DNAs (from bacteriophages ØX174, fd, M 13) have been analysed. In different DNAs, 74–82% of the 5′‐terminal dinucleotides end in 5′‐deoxycytidylic acid; small but significant levels of several dinucleotides ending in 5′‐deoxyadenylic acid, 5′‐thymidylic acid and 5′‐deoxyguanylic acid are also found. As far as 3′‐terminal nucleotides are concerned all nucleotides are present with a large predominance of thymidylic acid.
Solubilization and Purification of Rat, Liver Microsomal 1,2‐Diacylglycerol: CDP‐choline Cholinephosphotransferase and 1,2‐Diacylglycerol CDP‐ethanolamine Ethanolaminephosphotransferase
Authors: KANOH Hideo , OHNO Kimiyoshi 
Published: March 2005

The procedure, which involved 2‐step sonication of microsomes at pH 7.4 and then at pH 8.5 in the presence of sodium deoxycholate and subsequent dialysis, resulted in 4–5‐fold purification of choline‐phosphotransferase and ethanolaminephosphotransferase with the yield of 40–50%.
Ethanolaminephosphotransferase was further purified 8.5‐fold over microsomes by sucrose density gradient centrifugation of the partially purified preparation, while cholinephosphotransferase activity was considerably lost during this procedure. No separation of the two transferases from each other was achieved at this step.
Cholinephosphotransferase required Mg2+ as cofactor, and microsomal phospholipids for its maximal activity. On the other hand, Mn2+ was more effective than Mg2+ as cofactor for ethanolaminephosphotransferase, and this enzyme was inhibited by microsomal phospholipids.
Both transferases were stimulated several‐fold by sodium deoxycholate and also showed similar optimal pH ranging from pH 8.0 to 8.5.
Km values for 1,2‐diacylglycerol emulsion were 81.0 μM for cholinephosphotransferase and 63.0‐μM for ethanolaminephosphotransferase, respectively. CDP‐choline and CDP‐ethanolamine competitively inhibited, with the‐ same Ki value (both 350 μM), ethanolaminephosphotransferase and cholinephosphotransferase, respectively. The Ki values obtained were much greater than the corresponding Km values for the cytidine substrates (36.4 μM for CDP‐choline and 22.0 μM for CDP‐ethanolamine).
The partially purified enzymes were further treated with Triton X‐100. When enzyme activities were assayed with Mg2+, cholinephosphotransferase, although considerably inactivated, was partially separated from ethanolaminephosphotransferase by sucrose density gradient centrifugation of Triton‐treated preparations. Furthermore, cholinephosphotransferase (but not ethanolaminephosphotransferase) itself was partially separated into Mg2+‐requiring and Mn2+‐requiring components. In contrast, ethanolaminephosphotransferase assayed with either Mg2+ or Mn2+ formed a single peak together with Mn2+‐requiring cholinephosphotransferase.
Immunochemical Studies on Lipopolysaccharides from Wild‐Type and Mutants of Escherichia coli K‐12
Authors: MAYER Hubert , RAPIN Anette M. C. , SCHMIDT Günter , BOMAN Hans G. 
Published: March 2005

Lipopolysaccharides from a number of mutants of Escherichia coli K‐12 were investigated by means of chemical and serological methods. Inhibition of passive hemagglutination and inhibition of precipitation show that l‐rhamnose is the immunodominant sugar in the lipopolysaccharide from wild‐type E. coli K‐12, The disaccharide rhamnosyl‐KDO (where KDO is 3‐deoxy‐d‐manno‐octulosonic acid) was isolated and characterized after mild acid hydrolysis of the lipopolysaccharide. It is concluded that rhamnose is present in the innermost part of the core as a side‐chain substituent on KDO.From crosses between an E. coli K‐12 donor and E. coli O8, hybrids were obtained which contained either one or both of the donor rna and rfb clusters. Serum absorption studies with lipopolysaccharides from these hybrids indicated that the histidine‐linked rfb cluster is responsible for the presence of rhamnose in the K‐12 core oligosaccharide.Using paper chromatography of 32P‐labelled lipopolysaccharides we have found heterogeneous lipopolysaccharide in two strains as well as some differences between two wild‐type strains. The latter difference is believed to be due to varying contents of KDO‐linked ethanolamine phosphate. The overall results presented together with those described in the companion paper clearly show that the core oligosaccharide in E. coli K‐12 has a structure different from the types previously described for other strains of E. coli (designed coli R 1 to coli R4).
Functional Relationships between a Reticulocyte Polypeptide‐Chain‐Initiation Factor (IF‐MP) and the Translational Inhibitor Involved in Regulation of Protein Synthesis by Haemin
Authors: CLEMENS Michael J. 
Published: March 2005

The rate of initiation of protein synthesis in rabbit reticulocyte lysates is regulated by a translational inhibitor protein which is activated in the absence of added haemin. The effects of this inhibitor on amino acid incorporation are overcome by the protein synthesis initiation factor IF‐MP which binds Met‐tRNAf in a ternary complex with GTP and which can transfer this complex to small ribosomal subunits. Addition of this factor to haemin‐deficient lysates prevents loss of polysomes and regenerates polysomes from 80‐S single ribosomes, thus confirming an effect at the level of polypeptide initiation. The ability of the initiation factor to overcome the effects of various concentrations of the translational inhibitor suggests that the inhibitor inactivates the factor catalytically rather than stoichiometrically.In a system in vitro consisting of salt‐washed 40‐S ribosomal subunits, initiator Met‐tRNAf and GTP, the initiation factor IF‐MP transfers Met‐tRNAf to the subunits in the absence of any other factor or mRNA. Equilibrium buoyant density gradient analysis in CsCl shows that formaldehyde‐fixed subunits carrying Met‐tRNAf bound under these conditions have a buoyant density approximately 0.02 g/cm3 lower than the bulk of salt‐washed subunits, suggesting that approximately 100000 daltons of additional, protein are associated with these subunits. This is in marked contrast to the amounts of protein bound to subunits incubated with Met‐tRNAf and GTP in the presence of a crude ribosomal salt‐wash fraction. The translational inhibitor has no effect on formation of the ternary complex IF‐MP. Met‐tRNAf. GTP but does impair the factor‐catalysed transfer of Met‐tRNAf to washed subunits. The possible mechanisms of action of the inhibitor on polypeptide chain initiation are reviewed in the light of these results.
Intracellular Degradation and Deamidation of α‐Crystallin Subunits
Authors: VAN KLEEF Frans S. M. , WILLEMS‐THIJSSEN Willy , HOENDERS Herman J. 
Published: March 2005

The subunits AA2 and N2, present in α‐crystallin from the nucleus of young and old bovine lenses, were isolated and characterized. It was found that AA2 is identical to a shorter A‐chain and hence it was designated as A1–101. The subunit N2 turned out to be identical with a shorter B‐chain and‐was designated as B1/2−170. Characterization of the subunit N1, present only in α‐crystallin from the cortex of old bovine lenses, suggested that this subunit is a modified B‐chain, probably a deamidation product; it was designated as Bo.Comparison of the appearance of degraded and deamidated chains in relation to the age of Uhc fiber cells in old bovine lenses with that in calf lenses revealed that the observed specific limited degradation of the subunits of α‐crystallin increased with older age of the tissue. The deamidation process was found not to be related to the aging of the tissue. Eventually, a clear picture concerning the heterogeneity and fickleness of theα‐crystallin subunit structure was obtained.
Sulfated Bile Acids in Germ‐Free and Conventional Mice
Authors: EYSSEN Hendrik J. , PARMENTIER Guido G. , MERTENS Jan A. 
Published: March 2005

Sulfated and non‐sulfated bile acids were determined in the intestines and in the feces of 7‐month‐old germ‐free and conventional male mice.

The bile acid pools in the gall bladder and small intestine were 21.13 mg/100 g body weight in germ‐free and 11.50 mg in conventional mice. The bile acid pools in the cecum and large intestine of germ‐free mice were 3.03 mg/100 g body weight as compared to 1.24 mg in conventional mice. Fecal bile acid excretion was 2.93 mg and 4.12 mg/100 g body weight in 24 h in germ‐free and conventional mice respectively.
The major bile acids from germ‐free mice were cholic acid, α‐muricholic acid and β‐muricholic acid. Small amounts of chenodeoxycholic and allocholic acid were also present. In addition to these primary bile acids the following secondary bile acids were identified in conventional mice: lithocholic, deoxycholic and ω‐muricholic acid.
In both germ‐free and conventional animals significant amounts of chenodeoxycholic and cholic acid were present as the 7‐monosulfate esters. The sulfate esters of these bile acids did not exceed 2% of the total bile acids in the small intestine, but accounted for approximately 50 of the bile acids in the cecum and the large intestine. In contrast, the muricholic acids were nearly exclusively found in the non‐sulfate fraction.
Alkaline hydrolysis without prior solvolysis of the sulfate esters resulted in loss of bile acids and production of artifacts. Hence, the bile acids of the mouse cannot be analysed by methods involving alkaline deconjugation unless a solvolysis step is included in the procedure.
The Use of Mercurated Nucleoside Triphosphate as a Probe in Transcription Studies in vitro
Authors: BEEBEE Trevor J. C. , BUTTERWORTH Peter H. W. 
Published: March 2005

Purified form A RNA polymerase and the endogenous, nuclear form A RNA polymerase are shown to incorporate 5‐mercuri‐uridine 5′‐triphosphate (Hg‐UTP) into RNA in vitro. The Km for both Hg‐UTP and UTP are in the region of 10 μM for the purified enzyme.
The RNA products formed in nucleoli by endogenous RNA polymerase A have similar base compositions (G + C‐rich) whether UTP or Hg‐UTP is provided as the substrate in vitro.
Sulphydryl‐Sepharose chromatography of RNA synthesised in vitro by nucleoli allows separation of this material from the endogenous RNA, when the former is synthesised in the presence of Hg‐UTP.
In‐vitro‐synthesised nucleolar RNA hybridises with cot profiles similar to 28‐S ribosomal RNA, when made with either Hg‐UTP or UTP.
Hybridisation studies using DNA excess suggest that little competition occurs between the in vitro transcripts and the endogenous nucleolar RNA.
Size analysis of in vitro transcripts show that although some degradation occurs during isolation, purification and hybridisation of the RNA species, most of the RNA remains larger than 5 S throughout.
Studies on the Formation of Acetoin from Acetaldehyde by the Pyruvate Dehydrogenase Complex and Its Regulation
Authors: ALKONYI Istvái , BOLYGÓ Elek , GYÓCSI László , SZABÓ Dénes 
Published: March 2005

Pyruvate dehydrogenase complex was isolated from pigeon breast muscle involving steps of isoelectric precipitation, poly(ethyleneglycol) fractionation and separation on a glycerine gradient in the ultracentrifuge.
Arsenite, a potent inhibitor of the dihydrolipoyl transacetylase, did not affect the formation of acetoin from acetaldehyde, indicating that the pyruvate dehydrogenase component was operative in this reaction.
Production of acetoin by the pyruvate dehydrogenase complex is subject to regulation by phosphorylation and dephosphorylation, the dephosphorylated form only being active.
The inhibition by acetaldehyde of the pyruvate dehydrogenase complex could be partly explained by the formation of acetoin as an alternative reaction.
Synthesis, Separation, Identification and Interconversion of Riboflavin Phosphates and Their Acetyl Derivatives: A Reinvestigation
Authors: SCOLA‐NAGELSCHNEIDER Gisela , HEMMERICH Peter 
Published: March 2005

A reinvestigation of flavin phosphate synthesis, separation, identification, and interconversion was made in view of contradictory results in the literature. It has been confirmed that monochlorophosphoric acid is the best agent for selective 5′‐monophosphorylation of riboflavin and derivatives. This reaction yields, however, invariably up to 20% of an isomer, which has been separated by preparative thick‐layer chromatography and shown to be the 4′‐monophosphate. All the earlier authors failed to detect this isomer which does not bind to flavodoxin. It equilibrates in dilute mineral acid to yield an 8:2 mixture of 5′‐phosphate to 4′‐phosphate by phosphate migration.The formation of 2′,3′,4′‐triacetyl‐flavin mononucleotide, according to Christie, S. M. H., Kenner, G. W. & Todd, A. R. (1954) J. Chem. Soc., 46–52, upon acid‐catalysed acetylation of pure FMN, was confirmed. The same reaction under base catalysis, however, does not yield 2′,3′,4′‐triacetyl‐flavin mononucleotide as claimed by Khomutova, E. D., Shapiro, T. A., Mezentseva, M. V. & Berezovskii, V. M. (1965) Otd. Ohshch. i Tekhn. Khim., 241–244, Chem. Absir. 65, 5516a, but in fact up to 80% 2′,3′‐diacetyl‐flavin 4′: 5′‐cyclophosphate as the main product, which is stable under neutral and weak acidic conditions and does not hydrolyse to 2′,3′‐diacetyl‐flavin 5′‐monophosphate as claimed by McCormick, D. B. (1974) J. Heterocycl. Chem. 11, 969–974.The various flavin phosphates and their acetyl derivatives have been identified by pH‐titration, electrophoresis, and proton magnetic resonance spectrometry, which permit direct analyses of crude reaction products as well as a rapid purity check of commercial FMN.
Purification and Properties of 3‐Hexulosephosphate Synthase from Methylomonas M 15
Authors: SAHM Hermann , SCHÜTTE Horst , KULA Maria‐Regina 
Published: March 2005

3‐Hexulosephosphate synthase, the first enzyme of the ribulose monophosphate cycle, was purified 15‐fold from methanol‐grown Methylomonas M 15. The purification procedure involved chromatography on DEAE‐cellulose, Sephadex G‐75, and DEAE‐Sephadex A‐50. The purified enzyme was more than 95% pure as judged by analytical polyacrylamide gel electrophoresis. The molecular weight was calculated to be 43000 from sedimentation equilibrium experiments. Electrophoresis in sodium dodecylsulfate gels gave a single band corresponding to a molecular weight of 22000.The enzyme catalyzes specifically the condensation of formaldehyde with ribulose 5‐phosphate to yield d‐arabino‐3‐hexulose 6‐phosphate. The Km values were found to be 1.1 mM for formaldehyde and 1.6 mM for ribulose 5‐phosphate. A bivalent cation is essential for activity and stability of the enzyme, Mg2+ and Mn2+ serve best for this purpose. The optimum of pH for enzyme activity is 7.5–8.0.
Photolysis of Desmosine and Isodesmosine by Ultraviolet Light
Authors: BAURAIN Roger , LAROCHELLE Jean‐François , LAMY François 
Published: March 2005

Desmosine and isodesmosine were separated by ion‐exchange and paper chromatography, after acid hydrolysis of purified elastin from beef ligamentum nuchae. The fractions obtained by ion‐exchange chromatography were clearly mixtures of related compounds. The desmosine fraction could be resolved into seven compounds and the isodesmosine into four by paper chromatography.
Destnosine was maximally degraded by irradiation at 274 nm and isodesmosine at 285 nm. These wavelengths did not correspond to the absorption maxima of the cross links, but to shoulders of the main absorption peaks.
When irradiated at their optimum wavelengths, but at various pH, both desmosine and isodesmosine seemed quite stable at pH greater than 8.5. Between pH 8 and 5, the photolytic rate was maximum and decreased slightly at more acidic pH. Below pH 4.0, one of the products of photolysis was free lysine.
In analogy to the mechanism of the photolytic degradation of N‐methyl pyridinium chloride, it appears that the (iso)desmosines were degraded via the formation of an open amino aldehyde, which was hydrolysed at acid pH to give free lysine and a substituted glutaconic aldehyde.
Photosynthetic Activities of Membrane Preparations of the Blue‐Green Alga Phormidium luridum
Authors: BINDER Andres , TEL‐OR Elisha , AVRON Mordhay 
Published: March 2005

Techniques have been developed for the isolation of whole spheroplaste and membrane preparations from the blue‐green alga Phormidium luridum which are active in photosystem‐II‐sensitized and photosystem‐I‐sensitized electron‐transport reactions and in photophosphorylation. Attention is called to the necessity of supplying magnesium and phosphate throughout the preparative and assay procedures for obtaining active membranes.Selective washing of a proper membrane preparation is shown to remove a new protein factor, termed protein factor 1, which restores the ability of the preparation to catalyze photosystem‐I‐sensitized reactions.
Coenzyme Properties of NAD+ Bound to Different Matrices through the Amino Group in the 6‐Position
Authors: SCHMIDT Hanns‐Ludwig , GRENNER Gerd 
Published: March 2005

A method for the synthesis of N6‐(2‐aminoethyl)‐NAD+ is given. The binding of this NAD+ derivative to different soluble and insoluble supports and the direct coupling of NAD+ to epoxyactivated Sepharose are described. Proofs are given that NAD+ is bound through the amino group in 6‐position and the NAD+ derivative through the aliphatic amino group of the side chain.Non‐enzymic reduction of the bound coenzyme to an almost quantitative extent is possible in all cases, but the enzymic reduction is largely influenced by the support. While N6‐(2‐aminoethyl)NAD+ coupled to soluble dextran is nearly completely reducible by different dehydrogenases with a velocity of about 40% of that for free NAD+, the coenzyme bound to different insoluble matrices is very slowly reduced. Only 5% of the coenzyme derivative bound to BrCN‐activated Sepharose are reducible, but 40% when it is bound through a spacer. From capacity determinations evidence is given that, even in this coenzyme gel, only those coenzyme molecules are useful in affinity chromatography which are on the surface of the gel grains; it is supposed that this may be due to the slow diffusion of an enzyme into the inner parts of an affinity gel.
Pyridoxal 5′‐Phosphate and the Regulation of Ornithine Decarboxylase Activity and Stability
Authors: CLARK Jeffrey L. , FULLER Joyce L. 
Published: March 2005

There are two forms of ornithine decarboxylase with respect to pyridoxal 5′‐phosphate (pyridoxal‐P) affinity in exponentially‐growing Swiss 3T3 mouse fibroblasts: form I (Km∼ 10 μM) accounts for 30% of the total activity, and form II (Km∼ 0.4 μM) the remainder.Each form of the enzyme is in rapid equilibrium with ornithine and pyridoxal‐P; neither form recognizes the Schiff base between ornithine and pyridoxal‐P as a substrtate.Total pyridoxal‐P concentrations indicate that both forms may normally be at least partially active in vivo.Upon stimulation of 3T3 cells by pituitary growth factors, form I becomes undetectable within 4 h. As total activity increases over 10‐fold during this time and continues to increase thereafter, a possible conversion of form I to form II could account for this increase only if the Km change reflects other changes in preexisting enzyme.The rates of cofactor dissociation are apparently the same for each form and neither rate changes with the growth state. Since rapid equilibrium kinetics apply, the forms apparently differ in their rate of cofactor association.The half‐lives of the two forms in vivo are the same in unstimulated cells when measured concurrently. Also, the half‐life of total activity decreases markedly upon stimulation as form II becomes dominant. These and other observations are not consistent with pyridoxal‐P serving a major protective function for the enzyme in vivo.