bluuebunny/crossref_metadata_2025_split huggingface dataset:
Studies on Human Erythrocyte IMP: Pyrophosphate Phosphoribosyltransferase
Authors: Craft John A. , Dean Betty M. , Watts Richard W. E. , Westwick Wendy J. 
Published: March 2005

Some kinetic properties of the enzyme, IMP: pyrophosphate phosphoribosyltransferase have been investigated.
The effect of varying the concentration [S] of the purine substrate (hypoxanthine or guanine) on the initial velocity v of the reaction was studied at constant concentrations of P‐Rib‐PP and Mg2+. The velocity curves (plots of v against [S]) were hyperbolic.
The velocity curves were sigmoid when the P‐Rib‐PP concentration was varied at constant concentrations of purine and Mg2+.
Sigmoid velocity curves also resulted when the Mg2+ concentration was varied in the presence of constant concentrations of P‐Rib‐PP and purine.
Mn2+ could not replace Mg2+ in the system, but low concentrations of Mn2+ were stimulatory and high concentrations were inhibitory in the presence of low concentrations of Mg2+ and P‐Rib‐PP.
The shape of the curves obtained by plotting the P‐Rib‐PP concentration against the initial velocity at constant concentrations of purine and Mg2+ was pH dependent, being hyperbolic at pH 8.5 and sigmoid at pH 7.5 or less.
Inorganic orthophosphate had a slight activating effect when the P‐Rib‐PP concentration was varied at constant concentrations of purine and high concentrations of Mg2+ (6 mM). However, if the Mg2+ concentration was low (1 mM), high concentrations of inorganic phosphate (12 mM) were inhibitory.
The activity of the enzyme was reduced disproportionately with respect to the amount of protein present when it was highly diluted.
The possibility that these findings are due to the enzyme having allosteric properties, and their bearing on the possible function of IMP: pyrophosphate phosphoribosyltransferase in the regulation of de novo purine biosynthesis are discussed.
Étude comparative des DNA de phages 2C, SP 8*, SP 82, φe, SP 01 et SP 50
Authors: Truffaut Nicole , Revet Bernard , Soulie Marie‐Odile 
Published: March 2005

The following Bacillus subtilis phages: SP8*, SP82, φe, SP01, 2C and SP50, were studied and compared for morphological characteristics as well as for physico‐chemical properties of their DNA. All the determinations were effected under identical conditions for the six different phages. The aspect and variety of plaques formed by them with susceptible strain B. subtilis 168–2 (as indicator host), on Difco nutrient agar plates, are slightly different from phage to phage, but do not allow differenciation.As shown by electron microscopy measurements, the five phages SP8*, SP82, SP01, φe, and 2C are morphologically identical and easily distinguishable from SP50, but not from each other.As for the physico‐chemical properties of their DNA, the first five phages are clearly different from SP50 DNA which was taken as a reference: (a) their analytical base composition is consistently the same, all of them containing hydroxymethyluracil instead of thymine in their DNA, and the analyzed percentage guanine plus cytosine is 39%, as compared to 41.6% for SP50 DNA which has no unusual bases; (b) sedimentation coefficients for all the DNA (including SP50 DNA) are about 51 S, at a concentration of 20 μg/ml (s20, w); (c) the melting point in 0.15 M NaCl, 0.015 M Na citrate is 77.5°, without any significant variation among the first group (as compared to 87.3°, for SP50, which is approximately the same as the host Bacillus subtilis 168–2); (d) in a cesium chloride gradient, the buoyant densities of the native DNA of the first group are all 1.742 g/cm3. Since the complementary strands differ in average base composition, two bands of buoyant density 1.752 and 1.762 g/cm3 are obtained following denaturation; (e) all denatured DNA become associated with polyriboguanylic acid [poly(G)], which is preferentially bound to the heavy component, although a slight binding to the light component also occured. Preparative centrifugation in CsCl density gradients in the presence of poly G results in good separation of the two components; (f) the extent of homologous base sequence among the six phages has been determined by DNA/DNA hybridization: hybridization of 2C DNA, SP01 DNA and SP82 DNA with 2C DNA is experimentally identical and assumed to represent 100% homology. However for SP8 DNA/2C DNA and φe DNA/2C DNA, the percentage is lower, amounting to 92% and 83%, respectively. With SP50 and B. subtilis DNA it is only 7%.On the whole, there are no apparent properties, peculiar to one of the 2C, SP8*, SP82, φe, and SP01 phages, allowing its differentiation from the others, whereas SP50 is morphologically distinct, particularly by the length of its tail. It also differs by diverse physical and biological properties. All the common characteristics possessed by the first group of phages, particularly the high extent of DNA/DNA hybridization among them, and the fact that the phages and their DNA are serologically related, demonstrate their close relationship.
Identification of Sulfur‐Containing Bases in Transfer Ribonucleic Acids by Phosphorescence Spectroscopy
Authors: HÉLÈNE Claude , Yaniv Moshe 
Published: March 2005

The phosphorescence spectra and phosphorescence excitation spectra of tRNAVal, tRNATyr, tRNAPhe and total tRNA from Escherichia coli have been rcorded at 77° K in the presence and absence of magnesium ions. The main phosphorescence of these tRNA species originates from sulfur‐containing nucleosides which have very high phosphorescence quantum yields as compared to their oxygenated analogs.The phosphorescence of 4‐thiouridine excited at 345 nm has a very short lifetime (about) 3 msec). The phosphorescence spectrum and quantum yield of 4‐thiouridine in tRNATyr are shown to differ from those of 4‐thiouridine in tRNAVal and tRNAPhe (lower quantum yield and red‐shift). These differences are attributed to interactions between two adjacent 4‐thiouridine residues in tRNATyr. The phosphorescence spectrum of 2‐methylthio‐N6‐(Δ2‐isopentenyl)‐adenosine excited at 280 nm has a characteristic shape which is observed for many adenine on the phosphorescence spectra of the tRNA species used in this study. The phosphorescence quantum yields of thionucleosides in tRNATyr are increased in the presence of Mg2+ions.Phosphorescence measurements provide a convenient method to determine the presence of and identify sulfur‐containing nucleosides in pure species of tRNA. The possible use of thionucleoside phosphorescence to study conformational changes in tRNA and interactions involving tRNA molecules is suggested.
The Diagram, a Method for Comparing Sequences
Authors: Gibbs Adrian J. , Mcintyre George A. 
Published: March 2005

A simple method for detecting similarities in sequences is described. It is used:

To provide similarity measures for classifying 25 cytochromes by their amino acid sequences,
to detect repetitions in the amino acid sequences of various proteins,
to detect regions of possible base‐pairing in the nucleotide sequence of a nuclei acid.
Coupled Endogenous Respiration in Brown Adipose Tissue Mitochondria
Authors: Ppedersen Jan I. 
Published: March 2005

Carnitine + ATP were required for oxidation of endogenous substrates in brown adipose tissue mitochondria from newborn guinea‐pigs when the reaction medium lacked serum albumin. There was no evidence of phosphorylation linked to this oxidation.
When the reaction medium contained 2% serum albumin endogenous substrates were oxidized provided that phosphate and ADP were present. Carnitine did not influence the respiration. This oxidation was coupled to phosphorylation, P/O ratios between 2.4 and 2.8 being found.
Evidence is presented that the substrates responsible for the coupled endogenous respiration are mainly fatty acids, probably derived from an internal pool and activated in an internal mitochondrial compartment.
Brown adipose tissue mitochondria from newborn animals respiring on endogenous substrates could rapidly be brought into the controlled state (state 4) provided that high energy nucleotides were added to the reaction medium. In their absence, addition of ADP was followed by a slower and more gradual transition into state 4.
With brown adipose tissue from foetuses and weaned animals the endogenous respiration was under phosphate acceptor control in absence of high energy compounds.
The results suggest that the highly loose coupled state of brown adipose tissue mitochondria in thermogenic phases is related to lowered amounts of endogenous high energy phosphates.
Horse Liver Alcohol Dehydrogenase
Authors: Jørnvall Hans 
Published: March 2005

The [14C]carboxymethylated protein of the ethanol‐active isoenzyme of horse liver alcohol dehydrogenase has been treated with trypsin (with and without previous maleylation of the substrate), chymotrypsin, pepsin and cyanogen bromide, respectively. Peptide mixtures obtained have been fractionated and analysed.
Data are given for those peptides originating from a C‐terminal region comprising about 60% of the protein chain, and the amino acid sequence of this segment, containing the last 234 residues, is deduced. The N‐terminal part was known previously and the primary structure of the whole protein chain is therefore established.
The ethanol‐active isoenzyme is found to be a dimer of two completely identical protein chains, each 374 residues long. The N‐terminal part of the molecule contains the reactive cysteine residue, six out of the seven histidine residues in the whole chain, the majority of the mutations between the isoenzyme chains of different substrate specificities, and a region homologous to another dehydrogenase.
Serine residues that reacted during maleylation of the protein were found to undergo O → N maleyl shift when they became N‐terminal residues in peptides. Evidence was obtained suggesting that carboxymethylmethionyl peptide bonds are labile.
Horse Liver Alcohol Dehydrogenase
Authors: Jörnvall Hans 
Published: March 2005

Tryptic digests of the [14C]carboxymethylated derivatives of the three isoenzymes EE, ES and SS of horse liver alcohol dehydrogenase have been compared in “fingerprint” experiments.
Eight peptide spots present in the digest of the carboxymethylated EE enzyme were not detected in the digest of the carboxymethylated SS enzyme; and seven spots found in the latter were not discovered in the former. No other differences were noticed. The ES derivative yielded both types of spots but in reduced amounts. It is concluded that the E‐ and S‐types of subunits are very similar and that the ES isoenzyme is a hybrid molecule.
From the carboxymethylated SS and ES isoenzymes the S‐chain peptides that differ from their counterparts in the E‐chain were prepared and their structures analysed. They were compared to the known structures [1–3] of the corresponding E‐chain peptides. It is concluded that all the differences between the two sets of peptides are accounted for by amino acid changes at only six positions along the protein chains of the E‐ and S‐types of subunits, and an ancestral geneduplication is suggested. The differences at five positions (17, 94, 101, 110 and 366) are amino acid exchanges compatible with one‐base mutations, while the nature of the sixth difference (position 115) is not fully established.
The six differences make the S‐chain more hydrophobic and three units of charge more positive than the E‐chain. These properties fit the solubilities and electrophoretic mobilities of the three isoenzymes. The difference in substrate specificity between the E‐ and S‐chains might be explained by a direct participation in the substrate binding site of some of the residues exchanged, but other explanations cannot be excluded.
β‐Methylcrotonyl‐CoA‐Carboxylase
Authors: Apitz‐Castro Rafael , Rehn Kurt , Lynen Feodor 
Published: March 2005

β‐Methylcrotonyl‐CoA‐carboxylase was purified 120‐fold from an Achromobacter species. When the cells were grown on isovaleric acid, the enzyme was formed mainly during exponential growth. The enzyme can be labeled by addition of radioactive biotin to growing cells. A former procedure of purification was improved by using adsorption to calcium phosphate gel and chromatography on Sephadex G‐200. Enzyme homogeneous by both sedimentation and Tiselius electrophoresis was thus obtained in a yield of 40–70% and was crystallized in a form resembling crystalline propionyl‐CoA‐carboxylase.
The molecular weight of the enzyme as determined by sedimentation velocity is 760000 daltons. The biotin content of 1,27 μmoles/g indicates 4 moles of biotin are bound to 1 mole of protein as is also the case for propionyl‐CoA‐carboxylase and pyruvate carboxylase.
Glutamic and aspartic acid comprise 23% of the amino acids in the enzyme. Extrapolation of electrophoretic mobility as a function of pH indicates the isoelectric point of the enzyme is in the vicinity of pH 3.5. Such a value would mean that most of the carboxyl groups of the acidic residues are free.
The usual methods of preparations for electron microscopy, even with preceding fixation steps, cause degradation of the enzyme since only particles of about 1/4 of the expected size could be seen. However, ultracentrifugation did not indicate any dissociation of the enzyme into subunits.
Hemmung der Trypsinkatalyse durch kationische Carbonsäureester und Amide
Authors: Hartmann Hermann , Holler Eggehard 
Published: March 2005

In paper [5] a simplified model for the hydrophobic binding sites of trypsin was proposed, showing where 1‐amino‐n‐alkanes and α,ω‐diamino‐n‐alkanes are bound while they competitively inhibit the catalyzed hydrolysis of a substrate ester. In the model, the enzyme has two hydrophobic binding sites separated by an aqueous region which presumably contains the catalytic important residues. The first site binds up to 4 methylene groups of the aliphatic side chain adjacent to the ω‐amino group; additional –CH2 groups in the chain extend into the aqueous region. If the chain is larger than 7 carbon atoms, the additional methylene or methyl groups are bound to the second hydrophobic site. This site is found to bind less strongly than the first site.This paper presents a thermodynamic investigation of the binding properties of trypsin in the formation of complexes with cationic esters and amids which bear an ammonium group on either the end of the acid side chain or the end of the alcohol or amine component. Free energy and in addition enthalpy and entropy of binding were measured as a function of the length of the inhibitor molecule and as a function of the position of the ester or amide carbonyl group. It was found that the stability of a complex increases linearly with length of the molecule, and that the magnitude of increase depends on the position of the carbonyl group. Amides always exhibit a smaller increase than esters. Inhibitors with the ammonium group on the alcohol component were bound with about the same stability than the corresponding ω‐aminoacidesters. The results are consistent with the model. Positioning of the carbonyl group within the first or second binding site reduces stability due to restricted interaction of the hydrophobic residues on enzyme and inhibitor. Only when the carbonyl group falls into the aqueous region, the free energy for an inhibitor containing the ester group is similar to that for a 1‐amino‐n‐alkane with identical chain length.Enthalpy and entropy as functions of the position of the carbonyl group show minima for the position within the aqueous region. This observation is correlated with the jump of enthalpy and entropy of complex formation of trypsin in the series of 1‐amino‐n‐alkanes reported earlier.It is shown that the magnitude of free energy of complex formation with specific substrates is well estimated on basis of the model. As a consequence, the alternative binding of either the 2‐N‐acyl group or the alcohol or amine component to the second hydrophobic site may imply a mode of less productive binding. It is emphasized that unproductive binding should be reflected in the Michaelis Menten constants for substrate amides whereas the Michaelis Menten constant for a substrate ester is normally not perturbed.
A Kinetic Study of Saccharopine Dehydrogenase Reaction
Authors: Fujioka Motoji , Nakatani Yoichi 
Published: March 2005

Kinetic studies were carried out on saccharopine [ɛ‐N‐(l‐glutaryl‐2)‐l‐lysine] dehydrogenase which catalyzed a reversible reaction according to the following equation.Saccharopine + NAD++ H2O ⇌ lysine +α‐ketoglutarate + NADH + H+.Initial velocity kinetics of the reverse direction showed that the saccharopine dehydrogenasecatalyzed reaction was consistent with a mechanism that the addition and release of reactants to and from the enzyme follow an obligatory order. The product inhibition studies in the forward direction also revealed kinetic patterns consistent with the ordered mechanism where the sequence of addition of substrates being NAD+ and saccharopine, and that of release of products being lysine, α‐ketoglutarate and NADH. The various kinetic parameters determined at pH 6.8 and 22° are also described.
Concentration of Cyclic 3′: 5′‐Adenosine Monophosphate and Glycogen Metabolism in the Liver
Authors: Van den Berghe Georges , De Wulf Henri , Hers Henri‐Géry 
Published: March 2005

The increase in the concentration of cyclic adenosine 3′: 5′‐phosphate in the liver after the administration of a small dose of glucagon precedes the inactivation of glycogen synthetase by 1–2 min. This sequence of events is in agreement with the role of the nucleotide as second messenger in the hormonal action. The increase is dose dependent and is the same in control and prednisolone treated mice.Treatment of mice with glucose or prednisolone causes a slight decrease in the concentration of cyclic adenosine 3′: 5′‐phosphate in the liver. The minuteness of this effect is in contrast with the large activation of glycogen synthetase that occurs under the same conditions.
Cinétique des réactions catalysées par la polynucléotide phosphorylase de Escherichia coli
Authors: Bon Suzanne , Godefroy Thérèse , Grunberg‐Manago MARIANNE 
Published: March 2005

A systematic study of the effect of deoxy‐ADP on the reactions catalyzed by polynucleotide phosphorylase from Escherichia coli confirmed its behavior as a monofunctional analog of ADP.The polymerization reaction is limited to the addition of one (or two at the most) residues of dAMP to a primer oligonucleotide. The Km of oligonucleotides in this reaction have been studied as a function of chain length, from (pA)2 to (pA)6, and for (pA)40. The Km and Vmax decrease with the chain length, as in the phosphorolysis reaction, but the variation is quantitatively different. (pA)2pdA is not a good substrate for the limited elongation, due to its poor Vmax, while it is a substrate for phosphorolysis.The dADP/Pi exchange occurs only in the presence of oligonucleotide. Comparison of ADP/Pi exchange and dADP/Pi exchange in the presence of (pA)2 seems to indicate an interaction between binding sites of the oligonucleotide and the nucleoside diphosphate. This interaction is larger with ADP than with dADP.The inhibitory properties of dADP in both polymerization of ADP and phosphorolysis were studied. In the absence of oligonucleotide, dADP increases the lag phase of polymerization, and also increases the concentration of oligonucleotide necessary to remove the lag phase. In the presence of saturating oligonucleotide, dADP behaves like a competitive inhibitor of ADP. Similarly, dADP is a competitive inhibitor of both phosphate and oligonucleotide in the phosphorolysis reaction. In the phosphorolysis of poly A it is only competitive with phosphate.The results presented here are consistent with a complete reversibility of polymerization and phosphorolysis. Namely, the oligonucleotide, when used in the phosphorolysis reaction, covers the nucleoside diphosphate binding site. The importance of the 2′OH in the different positions of the substrates is discussed.
Oxidation of 5β‐Cholestane‐3α, 7α, 12α‐triol by Rat Liver Microsomes
Authors: Cronholm Tomas , Johansson Gunnar 
Published: March 2005

The oxidation of 5β‐cholestane‐3α, 7α, 12α‐triol by different subcellular fractions of rat liver homogenate was studied. The most efficient oxidation was observed with the microsomal fraction fortified with the 100000 ×g supernatant fluid. Efficient oxidation occurred also with the microsomal fraction fortified with NADPH. The main products were identified as a number of cholestanetetrols. 5β‐Cholestane‐3α, 7α, 12α,25‐tetrol and 5β‐cholestane‐3α, 7α, 12α, 26‐tetrol accounted for about 60% of the cholestanetetrols. The oxidation of 5β‐cholestane‐3α, 7α, 12α‐triol by the microsomal fraction fortified with NADPH was stimulated about five‐fold by administration of phenobarbital. Phenobarbital treatment stimulated hydroxylations at C‐23, C‐24 and C‐25 to a much greater extent than that at C‐26. Carbon monoxide inhibited all the side‐chain hydroxylations and the most marked inhibition was observed for the 26‐hydroxylation.5β‐Cholestane‐3α, 7α, 12α, 26‐tetrol was found to be a much more efficient precursor of cholic acid than either 5β‐cholestane‐3α, 7α, 12α, 25‐tetrol or the two C‐24 epimers of 5β‐cholestane‐3α, 7α, 12α, 24‐tetrol.
Immunochemistry of R Lipopolysaccharides of Escherichia coli
Authors: Schmidt Günter , Jann Barbara , Jann Klaus 
Published: March 2005

Acapsular (K−) forms were isolated from Escherichia coli strain E56b (serotype O8 :K27(A): H−). A number of different R strains were obtained from the K− forms by selection of spontaneous mutants and by phage selection.
The lipopolysaccharides of these R strains (consisting of core and lipid A) were subjected to mild acid degradation and, after removal of insoluble lipid A, the mixtures were fractionated by gel chromatography. Results of chemical analyses showed that the core oligosaccharides of different R mutants differed with respect to their sugar constituents as well as to the molar ratios of these constituents. A group of R mutants did not contain phosphate in their core oligosaccharides (P− mutants).
Enzyme determinations indicated that one R mutant had a block involving UDP glucose pyrophosphorylase, whereas the enzymes of activation and interconversion of component sugars were intact in the other R mutants.
By allelic exchange (mating with S‐form Hfr strains) it could be demonstrated that with one exception the chromosomal site of the SR mutation of all mutants maps close to mtl (mannitol utilization). In Salmonella the rfa locus maps in the same region. All R mutants had an unimpaired (his‐linked) rfb locus.
The groups of mutants which differed in the sugar composition of their core oligosaccharides could also be differentiated serologically and by their sensitivity to phages. There was serological cross reactivity between the lipopolysaccharides of some R mutants with R mutants of Salmonella minnesota which also have a very incomplete core.

It is concluded that the lipopolysaccharides of the R mutants described in this paper have incomplete core structures which result from blocks at successive stages in the synthesis of the coli R1 core. In analogy to Salmonella these E. coli R mutants should, with one exception, be termed rfa type mutants.