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Biomedical subjects

C Michaud

Publications and source records attributed to C Michaud.

At least 37 records · Page 2Linked to original sources
PubMed1990

A study of financial resources devoted to research on health problems of developing countries.

The Commission on Health Research for Development is an independent, international commission composed of 12 leaders from the fields of health research, social science research and development policy. Chaired by John Evans, the other Commissioners are Gelia Castillo, vice-chair, F. H. Abed, Sune Bergstrom, Doris Calloway, Esmat Ezzat, Demissie Habte, Walter Kamba, Adetokumbo Lucas, Adolfo Martinez-Palomo, Saburo Okita and V. Ramalingaswami. The Commission began its work in November 1987, charged with analysing the strengths, weaknesses and gaps in current research on health problems of developing countries, and making proposals and promoting action for improvement. The Commission is sponsored by a variety of foundations, bilateral donor agencies, and international organizations. The Commission's report was published in the spring of 1990. As part of the work of the Commission Secretariat, we have undertaken a study of financial resources devoted to research on the health problems of developing countries. The study which began in May 1988 has three objectives: (i) To describe the current pattern of funding for research on the health problems of developing countries. (ii) To provide a baseline against which future trends and/or new programmes can be measured. (iii) To explore the possibility of an ongoing information system on research on health problems of developing countries.

Developing Countries↗
PubMed1989

Pituitary multicatalytic proteinase complex. Specificity of components and aspects of proteolytic activity.

The 700-kDa multicatalytic proteinase complex from bovine pituitaries separates in polyacrylamide gel electrophoresis under dissociating and reducing conditions into 11 components with molecular masses ranging from 21 to 32 kDa. No higher molecular mass components were detected. A rabbit polyclonal antibody raised against the complex recognizes five immunoreactive components. As reported previously, the complex exhibits three distinct proteolytic activities designated as chymotrypsin-like, trypsin-like, and peptidylglutamyl-peptide hydrolyzing activities. All three activities are rather rapidly inactivated by 3,4-dichloroisocoumarin, a general serine protease inhibitor, however, the pseudo-first-order rate constants of inactivation of the three components differ within a wide range, with the chymotrypsin-like activity being most sensitive to inhibition. The peptidylglutamyl-peptide hydrolyzing activity is greatly activated by low concentrations of sodium dodecyl sulfate and fatty acids and seems to constitute the main component responsible for degradation of protein substrates. In addition to cleaving bonds on the carboxyl side of glutamyl residues, this activity also cleaves, albeit at a slower rate, bonds on the carboxyl side of hydrophobic residues; however, the secondary specificity of this component is clearly different from the chymotrypsin-like activity. Heparin selectively activates the chymotrypsin-like activity. The complex cleaves rapidly both native and dephosphorylated beta-casein in a reaction greatly accelerated by low concentrations of sodium dodecyl sulfate. The nature of proteolytic products, and also the rate of formation of acid-soluble, ninhydrin-reactive products, is different for the phosphorylated and dephosphorylated form of beta-casein, indicating that the degree of phosphorylation influences the rate and pattern of proteolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1989

[Nutrition behavior in adolescent students (15-19 years of age) in the Nancy metropolitan area. A comparison with the recommended nutritional intake of the French population].

This paper describes the food behavior of 495 adolescents (15-19 years old) sample living in Nancy (France). The results are compared with the Recommended Dietary Allowances (RDA) for French adolescent population. The RDA are not satisfied for important percentages of girls for the daily energy intake (26.3% of girls have allowances less than 30% RDA), for the calcium intake (28.7% of girls have allowances less than 40% RDA), and for the protein intake (17.6% of girls have allowances less than 20% RDA). Lipids consumption is excessive (19.9% of boys and 23.4% of girls have allowances greater than 20% RDA) and energy intake from breakfast are low (25% of adolescents provide less 10% of daily energy intake by breakfast) for all these teen-agers. These results confirm the data observed in two others studies concerning the food behavior of French adolescents.

Adolescent↗
PubMed1988

Substrate-related potent inhibitors of brain metalloendopeptidase.

Rat brain metalloendopeptidase (EC 3.4.24.15) generates Leu- and Met-enkephalin from several larger opioid peptides and is capable of degrading a number of neuropeptides. Substrate-related N-(1-carboxy-3-phenylpropyl) peptide derivatives were synthesized and tested for enzyme inhibition. The best of these derivatives, N-[1(RS)-carboxy-3-phenylpropyl]-Ala-Ala-Tyr-p-aminobenzoate, inhibited the enzyme in a competitive manner with a Ki of 16 nM. The data indicate that the carboxyl group of the N-(1-carboxy-3-phenylpropyl) moiety coordinates with the active site zinc atom and that the remaining part of the inhibitor is necessary for interaction with the substrate recognition site of the enzyme. Replacement of the 1-carboxy-3-phenylpropyl group by a carboxymethyl group decreased the inhibitory potency by more than 3 orders of magnitude, emphasizing the importance of the hydrophobic phenyl group for inhibitor binding to a hydrophobic pocket at the S1 subsite. Replacement of the Tyr residue by an Ala residue decreased the inhibitory potency by more than 20-fold. Changes in the structure of the residue interacting with the S1' subsite could cause a more than 60-fold change in inhibition. The inhibitors were either ineffective or only weakly inhibitory against membrane-bound metalloendopeptidase ("enkephalinase", EC 3.4.24.11), an enzyme highly active in rabbit kidney but also present in brain. The data indicate the presence of an extended binding site in the enzyme with residues interacting with S1, S1', and S3' subsites largely determining inhibitor binding.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Endopeptidase-24.15 is the primary enzyme that degrades luteinizing hormone releasing hormone both in vitro and in vivo.

The concentration of luteinizing hormone releasing hormone (LHRH) (pGlu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2), which reaches the anterior pituitary via the hypothalamo-hypophyseal portal system, appears to be controlled in part by the rate of LHRH degradation within the hypothalamus and/or pituitary. Specific, active site-directed endopeptidase inhibitors synthesized in our laboratory were used to identify the enzyme(s) involved in LHRH degradation by hypothalamic and pituitary membrane preparations, and by an intact anterior pituitary tumor cell line (AtT20). Incubation of LHRH with pituitary and hypothalamic membrane preparations led to the formation of pGlu-His-Trp (LHRH1-3) as the main reaction product. Under the same conditions, addition to the incubation mixtures of captopril, an inhibitor of the angiotensin converting enzyme, led to accumulation of pGlu-His-Trp-Ser-Tyr (LHRH1-5) and, to a lesser extent, pGlu-His-Trp-Ser-Tyr (LHRH1-6). The degradation of LHRH and the formation of the N-terminal tri- and pentapeptides was blocked by N-[1-(R,S)-carboxy-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoate (cFP-AAF-pAB), a specific, active site directed inhibitor of endopeptidase-24.15. Some inhibition of LHRH degradation and formation of the N-terminal hexapeptide was also obtained in the presence of N-[1-carboxy-2-phenylethyl]-Phe-p-aminobenzoate (cFE-F-pAB), an inhibitor of endopeptidase-24.11. Similar results were obtained with AtT20 cell membranes and with intact AtT20 cells in monolayer culture. Following cleavage by endopeptidases the C-terminal part of LHRH was rapidly degraded by aminopeptidases. Superactive analogs of LHRH in which Gly6 was replaced by a D-amino acid are resistant to degradation by both endopeptidase-24.11 and -24.15. In vivo, when LHRH was injected directly into the third ventricle of rats, the presence of cFP-AAF-pAB inhibited LHRH degradation. It is concluded that LHRH degradation is primarily initiated by the membrane-bound form of endopeptidase-24.15 to yield pGlu-His-Trp-Ser-Tyr and to a lesser extent by endopeptidase-24.11 to yield pGlu-His-Trp-Ser-Tyr-Gly.

Animals↗
PubMed1988

Incorporation of LL-diaminopimelic acid into peptidoglycan of Escherichia coli mutants lacking diaminopimelate epimerase encoded by dapF.

Recently a dapF mutant of Escherichia coli lacking the diaminopimelate epimerase was found to have an unusual large LL-diaminopimelic acid (LL-DAP) pool as compared with that of meso-DAP (C. Richaud, W. Higgins, D. Mengin-Lecreulx, and P. Stragier, J. Bacteriol. 169:1454-1459, 1987). In this report, the consequences of high cellular LL-DAP/meso-DAP ratios on the structure and metabolism of peptidoglycan were investigated. For this purpose new efficient high-pressure liquid chromatography techniques for the separation of the DAP isomers were developed. Sacculi from dapF mutants contained a high proportion of LL-DAP that varied greatly with growth conditions. The same was observed with the two DAP-containing precursors, UDP-N-acetylmuramyl-tripeptide and UDP-N-acetylmuramyl-pentapeptide. The limiting steps for the incorporation of LL-DAP into peptidoglycan were found to be its addition to UDP-N-acetylmuramyl-L-alanyl-D-glutamate and the formation of the D-alanyl-DAP cross-bridges. The Km value of the DAP-adding enzyme for LL-DAP was 3.6 x 10(-2) M as compared with 1.1 x 10(-5) M for meso-DAP. When isolated sacculi were treated with Chalaropsis N-acetylmuramidase and the resulting soluble products were analyzed by high-pressure liquid chromatography, the proportion of the main peptidoglycan dimer was lower in the dapF mutant than in the parental strain. Moreover, the proportion of LL-DAP was higher in the main monomer than in the main dimer, where it was almost exclusively located in the donor unit. There are thus very few D-alanyl-LL-DAP cross-bridges, if any. We also observed that large amounts of LL-DAP and N-succinyl-LL-DAP were excreted in the growth medium by the dapF mutant.

Amino Acid Isomerases↗
PubMed1987

Partial purification and specificity studies of the D-glutamate-adding and D-alanyl-D-alanine-adding enzymes from Escherichia coli K12.

The D-glutamate-adding and D-alanyl-D-alanine-adding enzymes from Escherichia coli were partially purified by fast protein liquid chromatography on an anion exchanger. Their relative molecular masses, determined by gel filtration on Superose 12, were 54,000 +/- 2000 and 51,000 +/- 2000, respectively. In order to investigate the specificity of these ligases, several compounds derived from their respective nucleotide substrates were prepared. In the case of the D-Glu-adding enzyme, DDP-MurNAc-L-Ala (DDP = dihydrouridine 5'-diphosphate) and P1-MurNAc-L-Ala were substrates of the reaction. In the case of the D-Ala-D-Ala-adding enzyme, only DDP-MurNAc-L-Ala-D-Glu(-meso-A2pm) was a substrate; P1-MurNAc-L-Ala-D-Glu(-meso-A2pm) was neither a substrate nor an inhibitor. Concerning the amino acid site of the D-Glu-adding enzyme, even closely related analogues of D-glutamate hardly inhibited the reaction.

Chromatography↗
PubMed1986

Substrate and inhibitor studies of thermolysin-like neutral metalloendopeptidase from kidney membrane fractions. Comparison with bacterial thermolysin.

The inhibitory constants of a series of synthetic N-carboxymethyl peptide inhibitors and the kinetic parameters (Km, kcat, and kcat/Km) of a series of model synthetic substrates were determined for the membrane-bound kidney metalloendopeptidase isolated from rabbit kidney and compared with those of bacterial thermolysin. The two enzymes show striking similarities with respect to structural requirements for substrate binding to the hydrophobic pocket at the S1' subsite of the active site. Both enzymes showed the highest reaction rates with substrates having leucine residues in this position while phenylalanine residues gave the lowest Km. The two enzymes were also inhibited by the same N-carboxymethyl peptide inhibitors. Although the mammalian enzyme was more susceptible to inhibition than its bacterial counterpart, structural variations in the inhibitor molecules affected the inhibitory constants for both enzymes in a similar manner. The two enzymes differed significantly, however, with respect to the effect of structural changes in the P1 and P2' positions of the substrate on the kinetic parameters of the reaction. The mammalian enzyme showed the highest reaction rates and specificity constants with substrates having the sequence -Phe-Gly-Phe- or -Phe-Ala-Phe- in positions P2, P1, and P1', respectively, while the sequence -Ala-Phe-Phe- was the most favored by the bacterial enzyme. The sequence -Gly-Gly-Phe- as found in enkephalins was not favored by either of the enzymes. Of the substrates having an aminobenzoate group in the P2' position, the mammalian enzyme favored those with the carboxyl group in the meta position while the bacterial enzyme favored those with the carboxyl group in the para position.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1985

Specificity of the uridine-diphosphate-N-acetylmuramyl-L-alanyl-D-glutamate: meso-2,6-diaminopimelate synthetase from Escherichia coli.

To investigate the specificity of the uridine-diphosphate-N-acetylmuramyl-L-alanyl-D-glutamate: meso-2,6-diaminopimelate synthetase, various compounds mimicking more or less different parts of the UDP-MurNAc-L-Ala-D-Glu substrate were prepared. Their size ranged from that of uridine or L-Ala-D-Glu to that of the whole nucleotide substrate. Chemical synthesis led to N alpha-acyl-dipeptides, in which the acyl group mimicked the MurNAc moiety, and to glycopeptides MurNAc(alpha or beta-Me)-L-Ala-D-Glu, in which the anomeric function is blocked. Partial degradation or chemical modification of the substrate UDP-MurNAc-L-Ala-D-Glu afforded: MurOHNAc-L-Ala-D-Glu, P1-MurNAc-L-Ala-D-Glu, and DDP-MurNAc-L-Ala-D-Glu (DDP = dihydrouridine-diphosphate). All these compounds were tested as substrates or (and) inhibitors of the reaction catalyzed by the A2pm-adding enzyme, which, after partial purification, was obtained in two active forms. Among the compounds tested as substrates, only DDP-MurNAc-L-Ala-D-Glu was a good one. The Km for this compound was 97 microM versus 55 microM for the natural substrate. Among the various compounds tested as inhibitors, only P1-MurNAc-L-Ala-D-Glu and MurNAc(alpha or beta-Me)-L-Ala-D-Glu had a significant inhibitory effect at 1mM. Apparently, no particular portion of the molecule is predominantly responsible for its recognition by the enzyme. In other words, multiple sites located over the whole molecule are required for a proper recognition and determine the high specificity of this activity. Therefore, to obtain efficient competitive inhibitors it is necessary to synthesize molecules very similar in size and structure to the natural substrate.

Binding Sites↗
PubMed1983

A soluble metalloendopeptidase from rat brain. Purification of the enzyme and determination of specificity with synthetic and natural peptides.

A metalloendopeptidase, optimally active at a neutral pH, was purified from the soluble fraction of brain homogenates. The enzyme (molecular weight about 67000) is strongly inhibited by metal chelators such as EDTA and o-phenanthroline. An EDTA-treated enzyme can be reactivated by several divalent metal ions including Zn2+, Co2+ and Mn2+. The specificity and kinetic parameters of the enzyme were studied with a series of model synthetic substrates. The enzyme preferentially cleaves peptide bonds in which the carbonyl group is contributed by an aromatic amino acid residue in the P1 position. The lowest Km values and the highest Kcat/Km ratios were obtained with substrates having aromatic residues in the P'3 and P1 position or in the P'3 and both the P1 and P2 positions. Lower kcat/Km ratios were obtained with substrates having arginine residues in position P1, and even lower values with those substrates having a glycine or aspartyl residue in this position. Introduction of a D-amino acid residue in either position P1 or P'1 renders the substrate totally resistant to hydrolysis. The specificity studies suggest that the active site of the metalloendopeptidase can accommodate at least five amino acid residues, with two of those residues binding on the N-terminal side and three binding on the C-terminal side of the hydrolyzed bond. Several biologically active peptides are cleaved by the enzyme at sites consistent with the specificity deduced from studies with model synthetic substrates.

Animals↗
PubMed1980

[Determination of Lewis and P blood group phenotypes with the aid of the Autogrouper 16C].

For more than one year, we are using the Autogrouper 16 C, at the beginning the prototype machine according to Rechsteiner, for ABO, Rhesus and Kell determinations and now Lewis and P grouping increases its possibilities, using free chanels. The connection to a PDP 11 computer enables us to sophisticate the interpretation of results and to use, for example, unadsorbed reagents.

Autoanalysis↗
PubMed1979

[Use of the BG 15 autoanalyzer for automatic recording and determination of ABO, Rhesus and Kell blood groups].

The use of the 15 channels blood grouping autoanalyzer with automated recording and interpretation proved its fiability through more than 30 000 tests for ABO, Rh and Kell determinations. Misreactions come mostly from the carry-over phenomenom. However no discrepancy was noticed for ABO determination. Only few ones occured for Rhesus and Kell, leading to false positive interpretations which were always corrected by checking the event recorder track. New improvements are expected from the automatic phasing using a microprocessor and further extents are setting up with the connection to a computer.

ABO Blood-Group System↗