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H Dugas

Publications and source records attributed to H Dugas.

At least 19 recordsLinked to original sources
PubMed1993

Synthesis, biological activity, conformational analysis by NMR and molecular modeling of N-formyl-L-Met-L-Pro-L-Phe-OMe, a proline analogue of the chemotactic peptide N-formyl-L-Met-L-Leu-L-Phe-OH.

The tripeptide N-formyl-Met-Pro-Phe-OMe (f-MPF-OMe), an analogue of the signal peptide N-formyl-Met-Leu-Phe-OH (f-MLF-OH), was synthesized and its chemotactic activity evaluated; it showed no activity in either superoxide production or calcium mobility with human neutrophils. However, the corresponding acid f-MPF-OH retained about 25% activity in the production of superoxide. The conformation of the f-MPF-OMe analogue was evaluated by NMR spectroscopy and molecular simulation and shown to predominate in a gamma-turn with a hydrogen bond between Met CO and Phe NH. Since this analogue is not chemotactic, it is suggested that for recognition the receptor prefers a peptide with a flexible backbone, favoring an extended conformation in the binding site.

Amino Acid Sequence↗
PubMed1991

Structure of a psoralen derivative of a monosubstituted 18-crown-6 ether.

N-[(2,5,9-Trimethyl-7-oxo-7H-furo[3,2-g]-[1] benzopyran-3-yl)methyl]-1,4,7,10,13,16-hexaoxacyclooctadecane- 2,3-dicarboximide, C29H35NO11, Mr = 573.60, monoclinic, P21, alpha = 18.877 (9), b = 6.888 (6), c = 24.488 (10) angstrom, beta = 119.90 (4) degrees, V = 2760.2 angstrom3, Z = 4, Dx = 1.380 Mg m-3, lambda (Cu K a) = 1.54178 angstrom, mu = 0.85 mm-1, F(000) = 1216, T = 170 K, R = 0.065, wR = 0.057 for 3720 observed reflections. There are two molecules (I and II) in the asymmetric unit and the 18-crown-6 part of molecule II is disordered. The psoralen moieties of the two molecules are nearly centrosymmetrically related.

Bleomycin↗
PubMed1989

Catalase activity measurement with the disk flotation method.

Activity measurements of catalase are usually performed with a spectrophotometric method by monitoring the decrease of H2O2 at 240 nm. A different method presented here uses an instrument named Catalase-meter and yields flotation time data which are expressed in tenth of seconds. For the first time, solutions of catalase of different concentrations were tested simultaneously with the two methods, and the Catalase-meter's flotation data were submitted to correlation with international units calculated from spectrophotometric data. The corresponding calibration curve correlates flotation time data to international units. The r2 values thus obtained for the two calibration curves were 0.960 and 0.929, for a range of activity varying from 9.3 to 144.5 international units/ml.

Animals↗
PubMed1985

Dopaminergic activity of four analogs of butaclamol.

The displacing potency of four analogs of the neuroleptic drug butaclamol were evaluated using dog striatal tissue and [3H]-Spiroperidol as ligand. Although significantly less powerful than the parent compound, two of them (N-isobutyl butaclamol equatorial; N-methyl butaclamol equatorial) could be used for dopaminergic receptor studies.

Animals↗
PubMed1981

Interaction of selectively spin-labeled 70S ribosomes with tRNAPhe. An electron paramagnetic resonance study.

70S ribosomes from Escherichia coli, selectively spin labeled on the SH groups of proteins S18, S12, S21, S17, and L27, were used to study the formation of the tertiary complex ribosome-poly(U)-tRNAPhe. Most of these ribosomal proteins are located in the region of binding of tRNA. The electron paramagnetic resonance observable structural change suggests a loosening of the ribosome structure upon binding of the tRNA molecule.

Ascorbic Acid↗
PubMed1980

Interaction of 70 S ribosomes from Escherichia coli with spin-labeled N-Cbz-Phe-tRNAPhe. An electron paramagnetic resonance study.

Two selectively spin-labeled Cbz-Phe-tRNAsPhe, one at position s4U8 and the other at position U33, have been used to study the dynamics of tRNA-ribosome interaction in the presence of poly(U) and factors washable from ribosomes. Upon binding to the ribosome, the correlation time of the spin label at position s4U8 decreases markedly while the same parameter for the label in the anticodon increases. The presence of poly(U) is not a prerequisite condition for the EPR spectral changes observed but larger variation occurs in the presence of factors washable from ribosomes. No variation in the correlation time is observed if uncharged spin-labeled tRNAPhe (on the s4U8 residue) is used in these experiments. Most of the ribosome-bound spin-labeled Cbz-Phe-tRNAPhe are puromycin-reactive, and consequently, the observed effect is manifested mainly at the ribosomal P site. These observations seem to suggest that the interaction between the N-blocked aminoacyl residue on the tRNA and the ribosome results in a conformational change on the tRNA, possibly involving tertiary interactions in a region close to s4U8. The role that the amino acid at the 3'-end can possibly play on this structural change is discussed.

Electron Spin Resonance Spectroscopy↗
PubMed1979

Selective spin-labeling of the ribosomal proteins of 70S ribosomes from Escherichia coli.

We have used a series of N-(1-oxyl-2,2,5,5-tetramethyl-3-pyrrolidinyl) maleimide spin labels of different length to label, covalently and selectively, the most reactive sulfhydryl groups of 70S ribosomal proteins of Escherichia coli. Under short periods of labeling (1--2 min), less than two spin labels per ribosome are incorporated and were shown to be distributed mainly on five ribosomal proteins in the following order: S18 greater than S21, L27 greater than S17, and S12. With a long period of labeling (3 h) up to 13 spin labels are attached to the ribosome, and protein S1 is the most labeled. The shape of the electron paramagnetic resonance (epr) signal shows two components with a predominance for the strongly immobilized orientation, and the percentage of these components in each spectra has been evaluated. When the distance between the nitroxide group and the maleimide-attaching group exceeds 6 A (1 A = 0.1 nm) the strongly immobilized orientation disappears. The effect of magnesium ions on these selectively spinlabeled ribosomes shows that the dissociation into subunits does not affect the epr signal, but more spin labels are incorporated into the subunits if labeling is performed under conditions of dissociation.

Bacterial Proteins↗
PubMed1978

Structural studies of staphylococcal protease. III. Binding of anions to the spin-labeled enzyme.

The staphylococcal protease was coupled at the active-site serine residue with a spin-labeled analog of diisopropyl fluorophosphonate and the interaction of competitive inhibitors such as chloride and acetate anions, as well as N-carbobenzoxy-L-glutamic acid (Z-L-Glu), was investigated by electron paramagnetic resonance spectroscopy. It was observed that the addition of chloride ions to the spin-labeled enzyme increased the freedom of motion of the spin label while the presence of acetate ions and Z-L-Glu resulted in an increase in the immobilization of the spin label. These results suggest that these ions bind to the active site region in different ways.

Binding Sites↗
PubMed1976

Evidence for a conformational change in tRNAPhe upon aminoacylation.

A conformational difference between the structure of tRNAPhe and Cbz-Phe-tRNAPhe from Escherichia coli was detected using the spin label method. A comparison of the respective rotational correlation time (tau c) values of three differently located spin labels, indicates that upon aminoacylation of tRNAPhe, the 4-thiouridine residue region and the miniloop region become more flexible, while the environment of the anticodon loop is not affected. These observations suggest that the main difference in structure between charged and uncharged tRNA resides in the release and exposure of the TpsiC loop for eventual binding to the ribosomes.

Escherichia coli↗
PubMed1976

Specific spin-labeling of transfer ribonucleic acid molecules.

The spin labels anhydride (ASL), bromoacetamide (BSL) and carbodiimide (CSL) were used to label selectively tRNAGlu, tRNA fMet and tRNAPhe from E. coli. The preparation and characterization of the sites of labeling of eight new spin-labeled tRNAs are described. The sites of labeling are: s2U using ASL, BSL and CLS and tRNAGlu; s4U using ASL and BSL on tRNAfMet and tRNAPhe; U-37 with CSL on tRNfMet; U-33 with CSL on tRNAPhe. The rare base X at position 47 of tRNAPhe has been acylated with a spin-labeled N-hydroxysuccinimide (HSL). The 3'end of unfractionated tRNA molecules has been chemically modified to a morpholino spin-labeled analogue (MSL). Their respective e.s.r. spectra are reported and discussed.

Binding Sites↗
PubMed1976

A spin label study of the thermal unfolding of secondary and tertiary structure in E. colic transfer RNAs.

The molecular mechanism of thermal unfolding of E. coli tRNAGlu, tRNAfMet and tRNAPhe (in 0.02M Tris-HC1, pH 7.5. 10 MM Mg C12) has been examined by the spin-labeling technique. The rate of tumbling of the spin label has been measured as a function of temperature for ten different selectively spin-labeled tRNAs. Only spin labels at position s4U-8 were able to probe the tertiary structure. Evidences are presented which support the hypothesis that the thermal denaturation of the three species of tRNAs studied is sequential. The unfolding process occurs in three discrete stages. The first step (30 degrees-32 degrees) could either be assigned to a localized reorganization of the cold-denatured structure or to a "transient" melting, followed by the simultaneous disruption of the tertiary structure and part of the hU helix. This transition is observed even in the absence of magnesium. The second step (50 degrees-54 degrees) involves the melting of the anticodon and miniloop regions. The last step occurs above 65 degrees where the t psi c and amino acid acceptor stems, forming one continuous double helix, melt. A simple dynamic model is considered for tRNA function in protein biosynthesis.

Binding Sites↗
PubMed1975

Proton magnetic resonance titration curves of the three histidine residues of staphylococcal protease.

Proton magnetic resonance spectra of staphylococcal protease, a serine protease from Staphylococcus aureus, strain V8, are presented. Initial proton spectra were obtained at 220 MHz, and more detailed studies of the aromatic region were carried out by correlation spectroscopy at 250 MHz. The overall spectrum bears a close resemblance to one calculated from the sum of spectra of the component amino acids. Chemical shifts of the three tyrosine, four phenylalanine, and three histidine residues appear to be equivalent at pH 3.7 and 8.5 indicating that they are all in normal chemical environments in the enzyme. The staphylococcal protease contains a large number of slowly exchanging protons. In fact, interpretable spectra of the aromatic region were obtained only after extensive exchange of N-H groups with deuterium from the D2O solvent. Proton magnetic resonance titration studies of the three histidine residues indicate that these have normal chemical shifts and pK' values. When the data are fitted to single noninteracting titration curves, the histidine pK' values are 7.19 plus or minus 0.02, 6.85 plus or minus 0.03, and 6.69 plus or minus 0.02. The titration curves of two of the histidine residues indicate negative cooperativity. A possible explanation for this is a direct electrostatic interaction between the two histidines. The titration data for these histidines give a significantly better fit to such a mutual interaction model than to noninteracting titration curves. The component microscopic dissociation constants have been calculated. Mutual interaction leads to pK' displacements of 0.31 unit; which indicates a distance of approximately 7 angstrom between the two interacting histidine rings according to the model of Tanford and Roxby. The proton resonances of the two interacting histidines are doubled in the pH region 6.7-7.0 suggesting the presence of two forms of the enzyme having lifetimes in excess of 30 msec.

Amino Acids↗
PubMed1975

Structural studies of staphylococcal protease. I. Spin labelling of the active site and a comparison with other proteases.

Staphylococcus aureus protease has been spin-labelled at the active-site serine residue with the monocyclic-phosphorus spin label (MSL), 1-oxyl-2,2,6,6-tetramethyl-4-peperi-dinylethylphosphorofluoridate. The electron paramagnetic resonance (E.P.R.) sbectra of the protease in different buffers at various pH's have been analyzed and compared with those of trypsin, subtilisin BPN', and alpha-chymotrypsin under identical conditions. In a given buffer, the shape of E.P.R. signals of spin-labelled staphylococcal protease is unaffected by pH changes except below pH 4.0, at which a gradual loss of conformational integrity of the active site occurs. In bicarbonate buffer and particularly in acetate buffer, the mobility of the label is much more restricted than in phosphate buffer or in potassium chloride solution. The implications of this finding are discussed in terms of a model whereby the label is able to orient towards two different but adjacent regions of the active site. The relative population of the label in each of these orientations is believed to be buffer-dependent. An attempt to correlate the shape of the te.p.r. signals with the pH values of maximal proteolytic avtivity of the enzyme is also presented. These results show that to obtain meaningful information from a comparative spin label study of the geometry of the active site of serine proteases, particular care should be exercised to assure that the different proteases experience identical conditions of pH, buffer, and temperature.

Binding Sites↗