bluuebunny/crossref_metadata_2025 huggingface dataset:
Phosphodeoxyribomutase from Escherichia coli
Authors: Hammer‐Jespersen Karin , Munch‐Petersem Agnete
Published: March 2005

Phosphodeoxyribomutase has been purified 67‐fold from a thymine starved dra−thy− mutant of Escherichia coli K12. The ratio of activities towards Rib‐1‐P and dRib‐1‐P did not change during the purification procedure, indicating that the same protein catalyzes the reaction with both substrates. The activity was dependent on the addition of Mn++, Co++ or Ni++ ions. Activity was stimulated 10‐fold by addition of ribo‐ or deoxyribose‐1,5‐diphosphate; the addition of glucose‐1,6‐diphosphate enhanced the activity 3‐fold. A molecular weight of 32000 ± 3000 was estimated by gel‐filtration.
Interactions entre l'oestradiol et des sites récepteurs utérins
Authors: Best‐Belpomme Martin , Friés Jeanne , Erdos Tamas
Published: March 2005

The oestradiol‐binding protein of calf uterine cytosol forms a slowly reversible complex with oestradiol. Kinetic studies on the formation of this complex, carried out with crude or partially purified uterine cytosol preparations, indicate a reaction of the second order. The rate constant shows an inverse relationship with protein concentration. Dissociation of the complex appears to occur in two stages, both following first order reaction rate kinetics. Affinity constants calculated from association‐dissociation kinetics, decrease with increasing temperature. The affinity constant measured under equilibrium conditions shows an inverse relationship with protein concentration, and does not appear to vary at temperatures between 0° and 18°. The treatment of the data from binding studies at equilibrium suggests a cooperative effect.

These findings indicate that the interaction between the oestradiol‐binding protein, and its ligand cannot be described by simple reaction schemes; work with purified material, now under progress, could simplify interpretation by eliminating ambiguities due to eventual specific or unspecific protein‐protein interactions in our present impure system.
Interaction of Haemoglobin with Ions
Authors: Jänig Gerd‐Rüdiger , Gerber Gerhard , Ruckpaul Klaus , Rapoport Samuel , Jung Friedrich
Published: March 2005

Binding of adenosine triphosphate to human methaemoglobin was investigated at pH 6.2 and 7.2 by the use of an ultracentrifugation method. Two classes of binding sites regarding their affinity were found. The association constant of the class with lower affinity will be diminished by temperature elevation from 5° to 21°. The heat of binding which was evaluated to be – 6.8 kcal/ mole supports the hypothesis derived from previous experiments on pH dependence of binding, that imidazole groups might be involved.
Structural Comparison of Ovalbumins from Nine Different Species
Authors: Fothebgill Linda A. , Fothergill John E.
Published: March 2005

The structures of nine avian ovalbumins were compared by starch gel electrophoresis, ultracentrifugation, tryptic peptide mobilities, and amino acid composition.

The results showed that the ovalbumins belonged to two structurally different groups which corresponded to the two taxonomic orders studied. Relationships within the orders were also indicated.
Direct Spectrophotometric Determination of the Structure of Dinucleoside Monophosphates
Authors: Feulicoli Romano A. , Cervelli Stefano , Ipata Piero L. , Rossi Carlo A.
Published: March 2005

The identification of the nucleoside liberated by the action of spleen or snake venom phos‐phodiesterase on dephosphorylated dinucleosidemonophosphates directly reveals whether the nucleoside participates in the phosphodiester bond with its 3′‐ or 5′‐hydroxyl. We report a sensitive direct Spectrophotometric procedure for the detection and estimation of the liberated nucleoside, based on the following specific enzyme systems: guanosine phosphorylase, guanase and xanthine oxidase for guanosine at 290 nm, adenosine deaminase for adenosine at 265 run; cytidine deaminase for cytidine at 280 nm and uridine phosphorylase for uridine at 282 nm.

The structure of dinucleotides is determined in single experiments: the liberated nucleoside bearing the free 3′‐ or 5′‐hydroxyl is identified and quantitatively measured by its response to one of the ancillary systems; the remaining 5′‐ or 3′‐mononucleotide is identified as nucleoside after addition of phosphomonoesterase.

The use of T1 and pancreatic ribonucleases, along with the proposed principles and techniques, in studying the sequence of trinucleotides and tetranucleotides is discussed in the Appendix.
Comparative Electron‐Nuclear Double Resonance Study of Two Flavoproteins
Authors: Eriksson L. E. Göran , Ehrenberg Anders , Hyde James S.
Published: March 2005

The proton electron‐nuclear double resonance (ENDOR) of flavin radicals in NADPH dehydrogenase and a flavoprotein from Azotobacter vinelandii have been studied at −160 and −120°. By combination of the apoprotein of NADPH dehydrogenase with [8‐Me‐2H3]FMN, it was demonstrated that the CH3(8) group of the coenzyme gives an ENDOR signal. The ENDOR signals from CH3(8) of the two proteins showed different hyperfine couplings which are consistent with the assignments of a radical anion, or a neutral radical enolized at 0(4), in NADPH dehydrogenase and a neutral radical protonated at N(5) in the Azotobacter flavoprotein. Analysis of the matrix‐ENDOR signals in H2O and 2H2O yields information concerning the exchangeability of protons both in the constituent water and on the apoprotein in the vicinity of the radical.
Stereochemical Aspects of Antigenic Specificity in Polysaccharide Determinants
Authors: Simmons D. Alastair R.
Published: March 2005

The structural changes that occur during the biosynthesis of Shigella flexneri type specific factors from precursor group antigen have revealed some of the stereochemical factors that are important in determining the antigenic specificity of complex polysaccharide determinants.
Regulation of Pyruvate Kinase by Fructose 1,6‐diphosphate in Saccharomyces cerevisiae
Authors: Barwell Clive J. , Woodward Brian , Brunt Rodney V.
Published: March 2005

The concentrations of glycolysis intermediates, citric acid and adenine nucleotides were measured in resting cells of Saccharomyces cerevisiae during aerobic glucose metabolism. Addition of glucose resulted in a decrease in the intracellular content of phosphoglyceric acids and phosphoenolpyruvate. As extracellular glucose became depleted the level of these intermediates increased. Thus changes in their intracellular content were regulated by factors other than the supply of extracellular substrate.

Cross‐over behaviour of phosphoenolpyruvate and pryruvic acid together with the inverse relationship which existed between the concentration of phosphoenolpyruvate and the rate of the pyruvate kinase reaction, indicated that regulatory changes in the activity of pyruvate kinase occurred. These activity changes were compared with those of the intracellular content of reported in vitro effectors of the enzyme: fructose 1,6‐diphosphate, citric acid, ATP and ADP. ATP and ADP did not appear to be involved in regulating changes in the activity of the enzyme. Although citric acid may have contributed towards some of the changes in enzyme activity, fructose 1,6‐diphosphate appeared to be the primary regulator, under the conditions of these experiments.

Comparison of the results with the properties reported for the isolated enzyme, suggested that, in vivo, fructose 1,6‐diphosphate affects the affinity of pyruvate kinase for phosphoenolpyruvate. This, in turn, appears to regulate the intracellular content of phosphoenolpyruvate and phosphoglyceric acids. The significance of the results is discussed in relation to both glycolysis and gluconeogenesis.
Reaction of Formaldehyde and of Methanol with Xanthine Oxidase
Authors: Pick Frances M. , McGartoll Mary A. , Bray Robert C.
Published: March 2005

Earlier studies on inactivation of xanthine oxidase induced by methanol and accompanied by development of a specific electron paramagnetic resonance signal (the Inhibited signal) have been extended. The same reaction takes place when enzyme is treated with formaldehyde. With this, time‐courses for signal development and disappearance of enzymic activity cannot be distinguished. Under a variety of conditions, disappearance of the signal accompanied by partial restoration of activity could be achieved. Side reactions also take place in the presence of formaldehyde. Electron paramagnetic resonance parameters of the Inhibited signal are reported. In the signal‐giving species a single non‐exchangeable proton interacts with Mo(V) in the enzyme active site. Experiments with C2H3OH and with 2HC2HO show that this proton is derived from the inactivating agent. It is proposed that a formyl residue, –CHO, becomes attached to a group in the active centre and that interaction of molybdenum with this residue stabilizes the five‐valent state of the metal, thus preventing enzyme turnover. Methanol is presumed to be oxidized to formaldehyde at the active centre before taking part in the reaction.
The Dissociation and Reassociation of Rat Liver Mitochondrial Monoamine Oxidase
Authors: B. H. Youdim Moussa , G. S. Collins Godfrey
Published: March 2005

Ultracentrifugation of solubilised purified rat liver mitochondrial monoamine oxidase gave an s20, w value of 6.8 and a molecular weight of 155 000 whereas the molecular weight determined by gel filtration was about 300 000. Using polyacrylamide gel electrophoresis, five enzyme forms have been separated; the molecular weights of four of these forms have been found by gel filtration to be about 300 000 whereas after exposure to 8 M urea or 1% (w/v) sodium dodecyl sulphate containing 0.1 M 2‐mercaptoethanol, a single non‐enzymatically active band of protein was isolated having a molecular weight of about 75 000. Reassociation was achieved by dialysis against 0.1 M phosphate buffer pH 7.4 containing 0.1 M potassium chloride. When dissociated for 10 min in the presence of 8 M urea and 0.2 M mercaptoethanol followed by dialysis, each isoenzyme form reassociated to form all five isoenzyme. The nature of the multiplicity of form of the isoenzymes is discussed.
Reinigung und Kristallisation der NAD‐Glykohydrolase aus C‐Streptokokken
Authors: Fehrenbach Franz J.
Published: March 2005

The purification and concentration of NAD‐glycohydrolase from group‐C streptococci was achieved by zone precipitation, adsorption chromatography and re‐chromatography on hydroxylapatite. Subsequently, the enzyme could be isolated as a microcristalline product showing double refraction in the polarization microscope.

Analytical gel permeation chromatography, analytical gradient elution chromatography on hydroxylapatite, density gradient centrifugation, sodium dodecylsulfate–urea electrophoresis and agar gel diffusion have been used to demonstrate the homogeneity of the purified enzyem. The polyacrylamide gel electrophoresis shows the enzyme as an uniformly migrating band. Furthermore the electrophoretic mobility of NAD‐glycohydrolase corresponded approximately to that of hemoglobin. This finding provides first evidence for the existence of identical subunits of the enzyme protein. Besides the demonstration of its catalytic function and antibody specificity the crystallized enzyme has been shown to possess hemolytic activity which could be activated by thiol groups. Though a variety of efficient purification procedures were employed they failed to remove Streptolysin‐O from the enzyme. Considering analytical data and criteria of homogeneity it may be assumed that a chemical correlation exists between the crystallized enzyme from group‐C streptococci and Streptolysin‐O previously described as two different substances.
Pyruvate Dehyrogenase Component Subunit Structure of the Escherichia coli K 12 Pyruvate Dehydrogenase Complex
Authors: Vogel Otto , Henning Ulf
Published: March 2005

The pyruvate dehydrogenase component of the Escherichia coli pyruvate dehydrogenase complex is shown to be a dimeric protein consisting of identical or very nearly identical polypeptide chains with a molecular weight 90 000 to 110 000.

In reasonable agreement with data obtained by Reed and collaborators the native enzyme was found to be a species with a sedimention coefficient s°20, w= 9.8 S, a diffusion coefficent D°20, w= 4.4 × 10−7 cm2/sec, and a molecular weight of 200 000 to 220 000. Amino acid analyses and analyses of tryptic peptides showed that the minimum chemical molecular weight is around 100 000. Serine was found as an N‐terminal residue but with poor yields. One carboxyl terminal sequence…Arg‐Leu‐Ala‐CO2H per 105 daltons protein was shown by carboxypeptidase degradation.

The protein could be dissociated by a variety of methods into only one species of subunit. The molecular weight of this subunit was found, by dodecylsulfate polycrylamide‐gel electrophoresis and sedimentation equilibrium in various solvents, to be 90 000 to 110 000.
Interaction of Hemoglobin with Ions
Authors: Klinger Reinhard G. , Zahn Dieter P. , Brox Dieter H. , Frunder Horst E.
Published: March 2005

Methods are described for the rapid determination of free ATP and ATP loosely‐ and firmlybound to oxygenated hemolobin using anion exchange resin and ultrafiltration. In hemolyzates of human erythrocytes under simulated in vivo conditions about 20% of total ATP is firmly‐bound to hemoglobin with an association constant of about 1 × 109 (M−1).

Depending on the total ATP concentration used, equilibrium between free and firmly‐bound ATP is reached within 0.2 and 0.5 sec after ATP addition to “stripped” hemolyzates. In equilibrium the exchange between these two fractions is very slow, not reaching readioactive equilibrium until several days after labelling the free ATP fraction. On the other hand, exchange between free and loosely‐bound ATP is very fast.

In addition to the known two binding sites for ATP in oxy‐ and deoxyhemoglobin there exists a third binding site which firmly binds ATP and other anions independet of the oxygenation state of the hemoglobin molecule.
Bovine Erythrocyte Cupro‐Zinc Protein
Authors: Bannister Joseph , Bannister William , WOOD Edward
Published: March 2005

Bovine erythrocyte cupro‐zinc protein was obtained by ethanol‐chloroform fractionation of red blood cells and purified by chromatography on fibrous DEAE‐cellulose, gel filtration on Sephadex G‐75, and rechromatography on microgranular DEAE‐cellulose. The protein has a molecular weight of abotu 33 000 by gel filtration, and contains 2 gram atoms of cupric copper and 2 gram atoms of zinc per mole. The amino acid composition of the protein has been determined. The protein has an isoelectric point at pH 4.95 by isoelectric focussing and is resolved into two ‘size’ isomers, one being a minor component, in polyacrylamide gel electrophoresis. The apoprotein behaves as a molecule of smaller ‘size’ than the native protein components in polyacrylamide gel electrophoresis. The ultraviolet spectra of the native and apoproteins show unusual features. These spectra together with the ultraviolet spectra of partially reconstituted apoprotein indicate a stabilizing effect of the zinc atoms on the structure of the chromophoric system. It is suggested that the copper and zinc atoms are not mutually replaceable at their respective sites in the protein. The near ultraviolet and visible spectra of the native protein have been fitted with Gaussian curves which indicate absorption bands at 340, 440 and 680 nm. The value of ɛM/Cu2+ for the 680 nm band has been found to be 120, indicating a cupric complex of high symmetry.
Étude du transfert enzymatique de D‐[14C]mannose á partir de GDP‐[14C]mannose, sur un accepteur endogène dans les microsomes d'Aspergillus niger
Authors: Letoublon Robert , Richard Michel , Louisot Pierre , Got René
Published: March 2005

A glycoprotein‐mannosyltransferase, utilsing an endogenous acceptor found in Aspergillus niger is herein characterized. Data presented indicate that among the subcellulr fractions, only microsomers are active in the mannose transfer reactions.

The enzyme catalising the transfer requires Mn++ or Mg++ (1–2 mM) and has an optimum pH of 6.8 and an optimum temperature of 22°.

It exibits a Km of 0.2 μM for GDP‐mannose. The transfer is inhibited by ATP, CTP and also bt GTP or its structural analogue β‐γ‐methlene‐guanosine triphosphate, but AMP or GDP have only a slight inhibitory action.

Triton X‐100 causes of 2‐ to 3‐fold enhancement of incorporation of mannose when a suitable detergent: protein ratio is applied.

The efficiency of this enzyme‐aceptor system is discussed.
Propertis of Phosphatidate Phosphohydrolase
Authors: Mitchell Martin P. , Brindley David N. , Hübscher Georg
Published: March 2005

A study was made of factors affecting the activity of the soluble phosphatidate phosphohydrolase which acts preferentially on membrane‐bound phosphatidate. The requirement for Mg++ was demonstrable only after exhaustive treatment of the membrane‐bound substrate with EDTA and sonication. An analysis of the reaction products indicated that the enzyme did not act on lysophosphtidate.

An indication of the substrate specificity of the soluble phosphatidate phosphohydrolase was obtained with preparations from cat intestinal mucosa by determining its stimulating effect on microsomal glyceride formation from glycerolphosphate and various fatty acids ranging from C6:0 to C18:0 and including C18:1. Maximum stimulation was obtained with oleate incorporation while the stimulation of incorporation of all saturated acid was far lower. Of these, the incorporations of palmitate and myristate were stimulated to the greatest extent.

A more direct assessment of the substrate specificities of soluble and particulate phosphatidate phosphohydrolases was obtained with preparations from rat liver by using membranebound phosphatidates of various fatty acid compositions as substrates. The soluble enzyme had the greatest activity with dimyristoyl and palmitoyl‐oleoyl phosphatidates, whilst dilauroyl and distearoyl phosphatidates were poor substrates. The particulate enzyme had in general much lower activities which were highest with distearoyl and dioleoyl phosphatidates.

Microsomal biosynthesis of phosphatidylcholine as determined by the incorporation of sn‐[1,3‐3H2]glycerol 3‐phosphate was enhanced by the addition o9f particle‐free supernatant indicating that the soluble phosphatidate phosphohydrolase is also important for the conversion of phosphatidate to that diglyceride which is subsquently incorporated into phosphatidylcholine.
Conductimetric Enzyme Assays
Authors: Lawrence Anthony J.
Published: March 2005

A direct reading, six channel conductivity recording apparatus has been constructed and used to assay enzymes, including some of those found in muscle. This paper gives the essential theory of the method, and indicates that it is a highly versatile assay system. It may be applicable whenever ionic substrates are involved, but this is more easily tested by experiment than by calculation. The manipulation involved in an assay is probably less than in any other method.

Conductimetry could be used as a kinetic method for some enzymes (for example amidases), but suffers from restrictions in the amounts of non‐reacting ionic species which can be tolerated.

Improvements in the temperature control of the apparatus could allow much higher sensitivity to be used.
Sur la structrue chimique des cires D, peptidoglycolipides macromoléculaires des souches humaines de Mycobacterium tuberculosis
Authors: Markovits Judit , Vilkas Erna , Lederer Edgar
Published: March 2005

A sample of Wax D, a macromolecular peptidoglycolipid of Mycobacterium tuberculosis, was purified by column chromatography on silicic acid and then analysed. It contains about 90% of a mycolate of arabinogalactan and 8% of a phosphorylated mucopeptide.

The structure of the mycolate of arabinogalactan shown in Fig.2 is based on the isolation of the following products of partial hydrolysis: (a) mycolates of arabinofuranose and arabinobiose, in which mycolic acids are esterified to the 5‐OH of arabinofuranose; (b) a 1→4 diagalactopyranoside; (c) a 1→4 arabinogalactoside; (d) an arabinodigalactoside having two 1→4 linkages.

The structure of the phosphorylated mucopeptide is shown in Fig. 3 B, as compared to that proposed by Migliore and Jollès [30] (Fig. 3 A). The discrepancy between these two structures might be due to different methods of purification of Wax D and of isolation of the mucopeptide.

These results as well as previous work of Azuma et al. [21], Migliore and Jollès [30], Kanetsuna and San Blas [32] lead to a hypothetical structure of Wax D shown in Fig.4.

The origin of Wax D is discussed; as already suggested by work of Kanetsuna and San Blas [32] and David et al. [34], Wax D is thought to be the result of partial enzymatic degradation of the cell wall.
Involvement of 1‐Methyladenosine and 7‐Methylguanosine in the Three‐Dimensional Structure of (yeast)tRNAPhe
Authors: Igo‐Kemenes Tibor , Zachau Hans G.
Published: March 2005

The role 1‐methyladenosine and 7‐methylguanosine in the structure and function of (yeast)tRNAPhe was investigated by studying the NaBH4‐reduction of this RNA, of fragments containing 1‐methyladenosine and 7‐methylguanosine and of the nucleosides themselves.

Treatment of 1‐methyladenosine with NaB3H4 yielded 1‐methyl‐6[3H]hydroadenosine which was fairly stable when kept under a nitrogen atmosphere in a weekly acidic solution. 1‐Methyladenosine was converted to the reduction product when it was part of an octanucleotide or of a fragment comprising the CCA‐half of (yeast)tRNAPhe, but it was not reduced when it was part of intact (yeast)tRNAPhe.

The reduction product of 7‐methylguanosine, 7‐methyl‐8‐hydroguanosine, was too unstable to be isolated under the conditions of oligonucleotide analysis. The reduction of 7‐methyl‐guanosine and the reoxidation could be followed, however, by measuring the absorption of the reduction product at 310 nm. It turned out that also 7‐methylguanosine was reduced when part of an oligonucleotide or a larger fragment of the tRNA while it was protected in the intact (yeast)tRNAPha.

The finding that in intact tRMA 1‐methyladenosine and 7‐methylguanosine are inaccessible to NaBH4 demands a revision of parts of the current three‐dimensionalmodels of tRNA. Our results support the hypothesis that it is the function of the positively charged odd nucleosides, as 1‐methyladenosine and 7‐methylguanosine to stabilize certain regions of the tRNA structure.

The amino acid acceptance of mixtures of CCA‐and pG‐halves of (yeast)tRNAPhe was not altered when 1‐methyladenosine was reduced. 1‐Methyladenosine is therefore probably not an essential part of the synthetase recognition site of this tRNA.
kinetic Studies on the Reaction Mechanism and the Citrate Activation or Liver Acetyl Coenzyme A Carboxylase
Authors: Hashimoto Tkashi , Numa Shosaku
Published: March 2005

Kinetoc studies were carried out on the reaction mechanism of rat liver acetyl coenzyme A carboxylase. The results of initial velocity and product inhibition studies on the forward and reverse reaction indicate that the carboxylase reaction is most likely to proceed through the “bi bi uni uni ping pong” mechanism. The order of the addition of substrates to the enzyme is as follos: ATP, HCO3‐ and acetyl‐CoA in the forward reaction, and malonyl‐CoA, Pi and ADP in the reverse reaction.

Kinetic analysis of the stimulatory effect of citrate on the initial velocities of the forward and reverse reaction indicate that the site of citrate action during catalysis lies predominantly onthe earboxylated enzyme. It is proposed that there exists and equilibrium between active and inactive forms of the carboxybiotinyl enzyme and that this equilibrium is shifted toward the active form in the presence of citrate.
Studies on the Structure of T1 Lipopolysaccharides
Authors: Berst Mireille , Lüuderitz Otto , Westphal Otto
Published: March 2005

Lipopolysaccharides of Salmonella T1 forms contain ribose and galactose both present as furanosides. Mild periodate oxidation of the T1 lipopolysaccharide followed by treatment with alkali resulted in the formation of a main fragment which was identified as poly‐ribose linked to the core polysaccharide. Poly‐ribose was also identified as a degradation product of periodate oxidized T1 lipopolysaccharide which had been treated with mild acid.Another fragment formed under these conditions was identified as being derived from ‐6‐Galf‐1,3‐Galf‐1,3‐Galf‐1,6‐Galf‐1‐. These results indicated that the T1‐specific polysaccharide of T1 lipopolysaccharides contain poly‐ribose and poly‐galactose regions. Galactose‐trisaccharide units may be present as repeating units of a poly‐galactose chain.
DNA Polymerase, Triphosphates and Deoxyribonuclease in a Systemof Synchronized L Cells
Authors: Madreiter Hermann , Kaden Peter , Mittermayer Christian
Published: March 2005

In a system of synchronously dividing mouse fibroblasts (L cells), the activity of several enzymes was determined in relation to initiation of DNA synthesis. Synchrony of DNA synthesis and cell division was obtained by selective mechanical detachment of cells in mitosis, thereby avoiding the use of blocking agents. The activity of DNa polymerase under assay condition spermitting terminal or replicative activity of DNA polymerase under assay conditions permitting terminal or replicative activity to occur shows characterstic fluctuations throughout the life cycle of the cell: whereas the terminal enzyme activity exhibits a peak at about 12 h, the replicative form of theenzyme has its maximum 16 h after mitosis. DNA synthesis starts at about 9 h and reaches its peak at 16 h after mitosis. The two enzyme fractions examined (particular and supernatant) prefer native DNA as primer. The supernatant enzyme exhibits exclusively terminal activity with heat‐denatured DNA as primer. The degradation of TTP by TTPase activity of the particular enzymes fraction in the S‐phase (phase of DNA synthesis) may contribute to a preservation of the TTP pool, thus acting as an additional regulatory emchanism. DNase activity nearly constant throughout the mitotic cycle.
Porcine Luteinizing Hormone and its Subunits
Authors: Hennen Georges , Prusík Zdeněk , Maghuin‐Rogister Guy
Published: March 2005

A new method is described for the isolation of the subunits of porcine luteinizing hormone which have not been characterized before. By chromatography on SE‐Sephadex C‐25 in 8 M urea, the two subunits LHα and β were obtained in good yield and without apparent cross contamination. The physicochemical and immunological properties of these subunits were studied. Unlike the bovine or ovine LHβ subunits described earlier, porcine LHβ subunit doest not contain lysine.

Porcine LHα subunit is electrophoretically heterogeneous and two components represented by two homogeneous electrophoretical zones were isolated and characterized. The amino acid composition of these componetns is very similar; they differ, however, slightly in peptide maps of their tryptic digests.

Whole LHα subunit was submitted to specific cleavage at methionine residues by cyanogen bromide and a homogeneous peptide not containing homoserine was isolated. The latter represents most probably the C‐terminal fragment of LHα subunit. Its partial amino‐acid sequence and composition is: Gly‐Asn‐Ala‐Arg‐Val‐Glu‐(His3,Lys,CM‐Cys3‐4,Asp,Thr2,Ser,Glu,Tyr2)‐Ser.

The fragment contains 34% of sugar (by weight). In view of the similarity in peptide maps and the unambiguous sequence described above, the primary structure of the different components of LHα subunit must be very similar.
Immunological Studies of Plant Hormones
Authors: Fuchs Sara , Haimovich Joseph , Fuchs Yoram
Published: March 2005

Specific antibodies to two plant hormones, namely indole‐3‐acetic acid and gibberellic acid, were producd in rabbits which were immunized with conjugates of either hormone to hemolocyanin. The Specific antiboddies obtained were used for assaying the hormones by several immunological assays. The most sensitive assay reported in this paper, for the determination of indole‐3‐acetic acid and gibberellic acid, is the inhibition of inactivation of modifed bacteriphage. For this assay conjugates of either hormone with bacteriophage T4 were prepared and characterized. Such chemically modified bacteriophahges were completely inactivated by the respective specific antiserum and this inactivation was inhibiteid by the free hormone.

The specificity of the antibodies produced was tested by a series of cross‐reactions. There was no immunological cross‐reaction between the two hormones which can, therefore, be determined independently. However, antibodies to indole‐3‐acetic acid cross‐reacted with chemically related compounds such as indoelcetyl‐ɛ‐amino carporic acid, tryptamine and truyptophan. The immunological specificity of different gibberellins tested was found to bne in good correlation with the biological specificity.
Mise en évidence dans les mitochondries des cellules de Mammifères de particules ribosomales se différenciant des ribosomes cytoplasmiques par leur taille et leurs acides nucléiques
Authors: Houssais Jean‐François
Published: March 2005

Evidence showing the presence of 61 S, 46 S and 35 S ribosome‐like particles in mitochondria of mammalian cells (a permanent line of mouse cells (LMTK‐) CI.1d, and mycoplasma‐free HeLa cells, is described.

The two known techniques for isolating the mitochondira were used and compared: method I involves the use of EDTA and prevents large contamination by the cytoplasmic ribosomes; method II is based on the use of high concentration of magnesium and NH4Cl, and is thought to give better protection of the mitochondrial ribosomes. Method II applied to mammalian cells gives larege contaimination by cytoplasmic ribosomes, but it is shown that it does not prevent the study of the mitochondiral particles.

The 61 S, 46 S and 35 S particle exhibit a ribosomalbehaviour at different magnesium conentrations: the 46 S and 35 S peaks increase at the expense of the 61 S peak at low magnesium concentration. The 80 S peak obtained by method II is shown to contain exclusively cytoplasmic ribosomes as indicated by base composition.

RNA was extracted and purified from whole mitochondira isolated by method I and II. The RNA species obtained by method I were the already known 16 S, 12 S, and 4 S mitochondiral RNA species. The RNA species obtaiend by method II were the 28 S cytoplasmic RNA and 18 S RNA. The proporation of the latter was however large than expected. No clear 16 S and 12 S RNA were seen between the large 18 S and the 4–5 S RNA. Base composition of the 18 S RNA proved to be different from that the 18 S cytoplasmic RNA, with a higher content of cytosine and adenine, and lower of guanine. On the other hand, the RNA extracted from the mixture of 61 S and 46 S particles obtained from mitochondria isolated by method I, gave the 16 S, 12 S and 4 S species, the 12 S species predominating. In order to define more precisely the RNA molecules of the 61 S and 46 S particles (obtained from mitochondira isolated by method II) their RNAs were extractd using the RNase inhibitors polyvinylsulfate and diethylpyrocarbonate. The base compositions of these RNAs are very similar. They differ both from the 28 S and 18 S cytoplasmic RNAs and the 16 S and 12 S mitochondrial RNAs by the proportions of cytosine, adenine and guanine. The analysis on surcrose grdient of the RNA of the 61 S particle has shown that it comprises to RNA molecules: an 18.6 S molecule migrating a little farther than the 18 S cytoplasmic RNA, and a 16 S molecule. Essentially a single RNA species was found in the 46 S particle which is 18.6 S.

Since a very strong nuclese activity in the mitochondrial lysates in the absence of nuclease inhibitors is observed the possible effect of nucleases on the measured size of ribosome‐like particles, and on the RNA species characterized, are discussed. Our data suggest that the 16 S and 12 S mitochondiral RNAs might be party degraded products of larger molecules.
Excision of Pyrimidine Dimers from Ultraviolet‐Irradiated DNA by Exonucleases from Mammalian Cells
Authors: Lindahl Tomas
Published: March 2005

Two DNA specific exonucleases, DNase III and Dnase IV, are present in mammalian cell nuclei. The activity of the purified enzymes with ultaviolet‐irradiated polydeoxynucleotides and DNA as substrates has been investigated. DNase III shows a very poor ability to release pyrimidine dimers, althought excision can be demonstrated by using high concentrations of enzyme. In contrast, DNase IV liberates dimers in an effective fashion in vitro, and is a likely candidate to serve in this function also in vivo. The pyrimidine dimers are relesed by DNase IV as parts of oligonucleotides containing 5–8 residus. Neither of the two exonucleases is markedly inhibited by caffeine.
The Action Pattern of Mammalian Deoxyribonuclease IV
Authors: Lindahl Tomas
Published: March 2005

The mode of action of an exonuclese that probably is involved in DNA repair, mammalian DNase IV, has been studied. By kinetic measurements with both uniformly and terminally labeled polynucleotides it is shown that enzyme breaks severa adjacent phosphodiester bonds each time it binds to its substrate, but tht the number of hydrolytic events is small compared to the degree of polymerization of the substrate. At 37° the enzyme releases 50‐200 nucelotides in an exonucleolytic fashion at each encounter with native DNA. The significance of these observations in relation to DNA repair in vivo is discussed.
Subribosomal Particles Separated from Liver and Hepatoma Homogenates with a Zonal Rotor
Authors: Mullock Barbara M. , Hinton Richard H. , Dobrota Miloslav , Froomberg Dorrit , Reid Eric
Published: March 2005

Native subribosomal cytoplasmic particles, labelled in vivo with [14C]orotic acid or with [32P]phosphate, have been isolated in quantity from rat liver and hepatoma by centrifugation in a zonal rotor. The particles were concentrated by membrane filtration and examined by resedimentation through sucrose gradients. The RNA contained in the particles was extracted and examined by electrophoresis and base‐ratio analysis. In liver, the 40 S region contained two distinct ribonucleoprotein particles, which are almost certainly two forms of the small ribosomal subunit. In addition, particles containing rapidly labelled, polydisperese, DNA‐like RNA were concentrated in this region. The 60 S region also contained some RNA not attributable to ribosomal subunits and was deficient in the latter. Hepatoma differed from liver in that its 40 S region was homogeneous on resedimentation and contained much less polydisperese RNA. The tumour also showed a much higher proportion of large to small ribosomal subunits.
Partial Nuclease Digestion of Transfer Ribonucleic Acids and Aminoacylated Transfer Ribonucleic Acids
Authors: Hänggi Urs J. , Zachau Hans G.
Published: March 2005

Charged and uncharged tRNAPhe and tRNASer from yeast, tRNAPhe from Escherichia coli, and acetyl‐Phe‐tRNAPhe (yeast) were exposed, under partial digestioin conditions, to four endonucleases and one exonuclease. The digests were analysed by disc electrophoresis. There were no qualitative or quantitative differences between the degradation patterns of the charged and uncharged forms of the two yeast tRNAs. Between the two forms of the E. coli tRNA or between acetyl‐Phe‐tRNAPhe (yeast) and Phe‐tRNAPhe (yeast) no qualitative differences were found. On the basis of the partial nuclease digestions together with some gel filtratioin experiments, major changes of conformation upon aminoacylation can be excluded for the investigated tRNAs. Small changes in conformation which do not result in the formation of a new exposed region or in the disappearnace of a previously exposed region cannot be excluded by our methods. The findings are discussed in relation to published experiments on the conformation of charged and uncharged tRNAs.
The Alignment of Cyanogen Bromide Fragments from the Flagellin of Salmonella adelaide
Authors: Davidson Barrie E.
Published: March 2005

The alignment of the polypeptide fragments obtained by digestion of flagellin with cyanogen bromide has been established from (a) the amino acid sequence of the methionine‐containing, tryptic peptides and (b) the results of end group analyses on the cyanogen bromide fragments. A partial amino acid sequence of flagellin is presented.
Polymorphism of Serine Specific Transfer Ribonucleic Acid
Authors: Paradies H. Hasko
Published: March 2005

A method is described for the crystallizatino of serine specific transfer ribonucleic acid (tRNAI+IISer) in the prsence of magnesium as a divalent counter ion. The crystallization conditions in relation to different temperatures and various tRNA concentrations were investigated in detail were and three crystal forms were found. Tentative values for the unit cell parameters for two forms determined from X‐ray powder photographs. For the first form a= 120.3 Å, b= 54.2 Å and c= 68.6 Å and the space group is probably P222; for the second form a= 213.2 Å and c= 115.7 Å and the space group is either P6 or P622. From the thrid crystal form a still X‐ray photograph with reflections out to 3.5 Å from a single crystal was obtained using a precession camera. This phtograph together with a precession photograph suggests monolinic symmetry with the cell dimensions of a= 63 Å, b= 41 Å, c= 112 Å and β= 102.30, with the possible space group P21.
A Comparison of Five Genetically Distinct Invertases from Saccharomyces
Authors: Ottoleghi Paul
Published: March 2005

The β‐D‐fructofuranoside fructohydrolases (invertases) of Saccharomyces under the control of the non‐alleic genes SU‐1, SU‐2, SU‐3, SU‐4, and SU‐5 have been examined with respect to the effect of pH on the Michaelis contstants and maximum vlocities of their hydrolysis of sucrose.

In every case, the two molecular forms of each invertase (the large extracellular form and the small intracellular form) appeared to be identical as far as the reaction kinetics, but between the different invertases both similarities and differences exist.

If quasi‐equilibrium is assumed to be operative, the reaction of all the invrtases fits a model according to which the enzyme‐substrate complexes of all five invertases behave, kinetically, in identical manner. The unsubstrated enzymes, on the other hand, show small but definite differences: SU‐1 and SU‐4 invrtases and SU‐2, SU‐3 and SU‐5 invertases form two groups within which the enzymes respond in essentially identical manner to variations in the environmental pH. SU‐2 invertase can be separated from the second group as it is unique among the five invertases studied in being inhibited by o‐dianisidine.
Isolement et identification de 4 UDP‐trisaccharides dans la glande mammaire et le colostrum de brebis
Authors: Denamur Robert E. , Gaye Pierre J.‐B.
Published: March 2005

4 UDP‐trisaccharides present in the mammary gland and colostrum of the ewe were isolated and identified. The nucleotides, extracted by means of trichloracetic acid, were purified in the following way : 2 ion‐exchange chromatographies on Dowex 1 (formate), filtration on Sephadex G‐25, paper electrophoresis, pH 5, followed by paper chromatography in 3 different solvents. Using these fractionation techniques, 4 uridine derivatives (X1′, X2′, Y1′, Y2′,) with positive reaction to the Warren neuraminic acid test, were obtained.

UDP‐5′, which constitutes the nucleotidic part of the 4 derivatives, was defined more accurately after various acid or enzymatic analyses. The isolation of UDP‐5′ was made by ionexchange or paper chromatography and the total and labile phosphate was estimated. The Warren positive substances released from X1′, X2′, Y1′, Y2′, by neuraminidase or mild acid hydrolysis were identified as N‐glycolyl‐neuraminic acid for the X1′, Y2′ nucleotides, and N‐acetyl‐neuraminic acid for the X2′, Y2′ nucleotides.

The behaviour of asialylated nucleotides to ion‐exchange and paper chromatography is described as well as that of asialylated oligosaccharides. The 4 asialylated oligosaccharides contain galactose and an N‐acetyl‐D‐hexosamine, the latter being N‐acetyl‐D‐glucosamine in the case of X1′ and X2′, and N‐acetyl‐D‐galactosamine in the case of Y1′ and Y2′. Total analysis of UDP‐oligosaccharides shows a 1:1:1:1 ratio between uridine, N‐acetyl‐D‐hexosamine, galactose, and neuraminic acid.

The 1‐P‐trisaccharides and the trisaccharides were identified after phosphodiesterase hydrolysis followed by phosphatase hydrolysis. The N‐acetyl‐D‐hexosamine carried the reducing group (sodium borohydride). Some structural characteristics of the 4 nucleotides were determined using β‐galactosidase, neuraminidase, periodate, and periodate borohydride. Two of the nucleotides contain N‐acetyl‐glucosamine and are identical to those previously isolated from the goat colostrum. As for the two others, containing N‐acetyl‐galactosamine and appearing as new trisaccharide nucleotides, we may, on the basis of the results obtained, suggest the following sequences:

a) UDP‐N‐acetyl‐D‐galactosamine (3←1) D‐galactose (6←2) N‐acetyl‐neuraminic acid;

b) UDP‐N‐acetyl‐D‐galactosamine (3←1) D‐galactose (6←2) N‐glycolyl‐neuraminic acid. The possible role of these new nucleotides is discussed.
The Atomic Structure of Erythrocruorin in the Light of the Chemical Sequence and its Comparison with Myoglobin
Authors: Huber Robert , Epp Otto , Steigemann Wolfgang , Formanek Helmut
Published: March 2005

The atomic structure of the monomeric insect haemoglobin is closely similar to the structure of whale myoglobin, although only about 20% of the amino acids are identical in both protein molecules.

A quantitative comparison of both structures, however, showed that in the vicinity of some amino acid substitutions considerable changes occur. The comparison also revealed unexpected long range structural changes induced by amino acid deletions. In particular a deletion of four amino acids in the A B corner shifts the whole helix B considerably.

The general distribution of amino acids in erythrocruorin is analogous to that which has been found in other haemoglobin molecules. The large proportion of phenylalanines in the hydrophobic core of the molecule is remarkable. Also the haem pocket is lined with seven phenylalanine side chains. Four of these are unknown in vertebrate haemoglobins.

The distal histidine E 7, which is invariant in all known vertebrate haemoglobins, is replaced by a glutamic acid, which has no haem contact, but protrudes into the surrounding solution. This proves that a histidine or another polar residue is not essential for the function of a haemoglobin molecule as a reversible oxygen carrier.
Spécificité de la protéase neutre de Micrococcus caseolyticus
Authors: Desmazeaud Michel J. , Hermier Jean H.
Published: March 2005

The specificity of an exocellular neutral protease produced by Micrococcus caseolyticus has been investigated with amides, dipeptides mono‐and di‐substituted dipeptides and glucagon. This protease does not exhibit exopeptidase activity: it splits neither amides : Leu(NH2), Phe(NH2), Ala(NH2), Val(NH2), Ileu(NH2), nor Gly‐Leu, Gly‐Phe, Gly‐Ala, nor Z‐Gly‐Leu, Z‐Gly‐Phe, Z‐Gly‐Phe, Z‐Gly‐Ala. There is no free amino acid in glucagon digests. Therefore this protease has to be considered as an endopeptidase. Moreover it splits neither AcTyrOEt or BzTyrOEt (chymotrypsin substrates) nor TosArgOMe or BzArgOEt (trypsin substrates).

The results obtained with disubstituted dipeptides Z‐Gly‐X(NH2) indicate that the peptide bonds which are hydrolyzed by the neutral protease were those involving amino groups of hydrophobic amino acids (Phe, Leu, Ala, Val, Ileu but not Trp) or of tyrosine. The highest rate of hydrolysis was obtained with Z‐Gly‐Phe (NH2); it was 1.48 micromoles per minute per mg of protease at 37°C. The second residue (X′) involved in the bond by the carboxyl group in the series Z‐X′‐Phe(NH2) has also an important effect on enzymic activity. So the rates of hydrolysis (in micromoles per min per mg of protease, at 37°), were: 2.01 for Z‐Ala‐Phe(NH2), 1.48 for Z‐Gly‐Phe(NH2) 0.25 for Z‐Leu‐Phe(NH2), 0.146 for Z‐Ser‐Phe(NH2), 0.140 for Bz‐Gly‐Leu(NH2) and 0.122 for Z‐Gly‐Val(NH2).

This specificity has been confirmed with glucagon. Peptide bonds in which phenylalanine, leucine or alanine are involved by their amino group are rapidly split.

The M. caseolyticus neutral protease displays a substrate specificity which is similar to that found for Pseudomonas aeruginosa or Streptomyces griseus. Thus the hydrophobic nature of the specificity of the protease of M. caseolyticus is not as absolute as that for the megateriopeptidase of Bacillus megaterium.
The Elastase‐Like Enzymes from Streptomyces griseus (Pronase)
Authors: Gertler Arieh , Trop Moshe
Published: March 2005

Three elastase‐like enzymes, designated Enzyme I, II and III, were purified from a commercial preparation of Streptomyces griseus (pronase) by successive chromatography on DEAE‐cellulose at pH 9.5 and CM‐cellulose at pH 5.5.

Electrophoresis on cellulose acetate at pH 6.9, on polyacrylamide gels at pH 4.5 and 9.5 and exclusion chromatography on Sephadex G‐75 indicated that all three fractions were essentially homogenous.

The amino acid composition of the three enzymes was determined and their molecular weights were estimated by gel‐chromatographic methods.

All three enzymes possess strong esterolytic activity when assayed on a specific elastase substrate N‐acetyl‐L‐alanyl‐L‐alanyl‐L‐alanine methyl ester (AcAla3OMe) and are active, to a smaller extent, on N‐acetyl‐L‐tyrosine ethyl ester. Their activity on α‐N‐benzoyl‐L‐arginine ethyl ester was negligible. All the enzymes have strong proteolytic activity at slight alkaline pH and Enzyme II and III possess also an elastolytic activity on congo red elastin.

All three enzymes are inhibited by diisopropylfluorophosphate. Enzymes II and III were slowly inhibited by N‐tosyl‐L‐phenylalanine chloromethane. Enzyme I was inhibited by the esterolysis products of AcAla3OMe, thus indicating that the same active site controls both esterolytic activities.
A Comparison of Nucleotide Sequences from Mitochondrial and Cytoplasmic Ribosomal RNA of Aspergillus nidulans
Authors: Verma Inder M. , Edelman Marvin , Littauer Uriel Z.
Published: March 2005

The nucleotide fragments arising from digestion of Aspergillus nidulans mitochondrial and cytoplasmic ribosomal RNAs with T1 ribonuclease have been compared by means of two dimensional ionophoresis. Major qualitative and quantitative differences were found to exist between the two ribosomal RNA species. In addition, distinct differences were observed between the fingerprint patterns of the heavy and light components within each ribosomal RNA sample. The larger component of mitochondrial ribosomal RNA presented a more complex ionophoretic pattern than the smaller one, implying that the former was not a simple dimer of the latter.

It is concluded that the nucleotide sequence of the mitochondrial rRNA differs from that of the homologous cytoplasmic rRNA species.