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V Fleury

Publications and source records attributed to V Fleury.

15 recordsLinked to original sources
PubMed2001

[A fundamental symmetry between morphogenesis and function of branched organs].

It is generally difficult to find any relationship between the morphogenesis of an organ and its final function. A priori, such a relationship has no reason to exist, since organs do not actually function during their formation. I will show in this article that, for a very large class of organs--the branched organs--there exists a hidden relationship between their morphogenesis and their function. This class of organs comprises: the lungs, the salivary mammary and lacrymal glands, the kidneys, the pancreas, and possibly other organs, such as testes. For all these organs, a fundamental fact that comes from recent developments in physics explains at the same time how they form, and why they work. This suggests, first, that complex organs are not the result of gradual and long selection processes, and, second, that this specific structure for the organs is imposed by the laws of physics. The growth process, as described here, is possibly the only one that allows both to build a fluid-secreting organ, and make it work.

Animals↗
PubMed1998

Human thrombopoietin structure-function relationships: identification of functionally important residues.

Thrombopoietin (TPO) is a haematopoietic growth factor responsible for megakaryocyte progenitor proliferation and differentiation. It belongs to the four-helix-bundle cytokine family and exerts its biological effects through binding to a specific receptor, c-Mpl. With the use of site-directed mutagenesis we have generated 20 TPO mutants. Each of the TPO mutants was produced in a eukaryotic expression system and the mutants' ability to induce the proliferation of factor-dependent c-Mpl-expressing megakaryoblastic M-O7e cells was compared with that of wild-type TPO. Among the mutations studied, 10 lead to a significant decrease in TPO bioactivity. Of these ten residues, three are located in helix A of the protein (Arg10, Lys14 and Arg17) and four in helix D (His133, Gln132, Lys138 and Phe141), indicating that in TPO, as in other cytokines, these two helices are important for functional cytokine/receptor interactions. Surprisingly, mutant Arg10-->Ala (R10A) lacked any proliferative activity, despite the fact that this mutation was recently reported to have no effect on TPO/c-Mpl binding in a TPO phage ELISA [Pearce, Potts, Presta, Bald, Fendly and Wells (1997) J. Biol. Chem. 272, 20595-20602]. The lack of M-O7e proliferation is probably due to an inability of R10A mutant to promote receptor dimerization and thus receptor activation. Moreover we found that the Arg10 and Arg17 residues of TPO seem to be specific determinants for TPO/c-Mpl recognition. We also demonstrate that the O-glycosylation site located at position 110 of TPO is not necessary for the bioactivity of the cytokine.

Amino Acid Sequence↗
PubMed1993

Mechanism of the enhanced intrinsic activity of single-chain urokinase-type plasminogen activator during ongoing fibrinolysis.

The activation of plasminogen at the surface of fibrin by single-chain urokinase-type plasminogen activator (scu-PA) was investigated using recombinant forms of a plasmin-resistant mutant of scu-PA, rscu-PA-Glu158, and an inactive catalytic site mutant of human plasminogen, rPg-Ala741. Conversion of cleavable 125I-labeled single-chain proteins to their two-chain forms, was quantitated by radioisotope counting of protein bands on reduced SDS-polyacrylamide gels. The efficiency of the activation (moles of plasmin generated per mol of plasminogen activator) of native Glu-plasminogen bound to degraded fibrin was comparable for scu-PA and its two-chain form (tcu-PA) and approximately 4-fold lower for rscu-PA-Glu158. The corresponding values with rPg-Ala741 were 4-fold or 9-fold lower for scu-PA or rscu-PA-Glu158, as compared to tcu-PA. In contrast, in solution in the absence of fibrin, the efficiency of scu-PA for activation of rPg-Ala741 was 100-fold lower than that of tcu-PA. Initial activation rates of rPg-Ala741 (32.7 fmol/well containing 50 microliters of solution) with 4 nM tcu-PA were comparable in solution and bound to degraded fibrin (v(o) = 1.01 and 1.16 fmol/min, respectively). In contrast, with 4 nM scu-PA the corresponding values when rPg-Ala741 was bound to degraded fibrin were 20-fold higher as compared to the soluble phase (v(o) = 0.23 and 0.012 fmol/min, respectively). Comparable results were obtained when using Glu- or Lys-forms of rPg-Ala741. Furthermore, in the presence of normal human plasma, activation of Glu-plasminogen bound to degraded fibrin was found to be about 2.5-fold more efficient with scu-PA than with tcu-PA. These findings indicate that the fibrin specificity of scu-PA does not require its conversion to tcu-PA, nor conversion of Glu- to Lys-plasminogen, but appears to be due to the additional binding of plasminogen to partially digested fibrin; scu-PA may thus represent a physiological functional form of u-PA in plasma.

Alanine↗
PubMed1993

Molecular assembly of plasminogen and tissue-type plasminogen activator on an evolving fibrin surface.

A well characterized model of an intact and a degraded surface of fibrin that represents the states of fibrin during the initiation and the progression of fibrinolysis was used to quantitatively characterize the molecular interplay between tissue-type plasminogen activator (t-PA), plasminogen and fibrin. The molecular assembly of t-PA and plasminogen on these surfaces was investigated using combinations of proteins that preclude complications due to side reactions caused by generated plasmin: native plasminogen with di-isopropylphosphofluoridate-inactivated t-PA, and a recombinant human plasminogen with the active-site Ser741 mutagenized to Ala which renders the catalytic site inactive. Under these conditions, neither the affinity nor the maximal number of binding sites for plasminogen were modified by the presence of t-PA, indicating that binding sites for plasminogen pre-exist in intact fibrin and are not dependent on the presence of t-PA. In contrast, when plasminogen activation is allowed, increasing binding of plasminogen to the progressively degraded fibrin surface is directly correlated (r = 0.98) to the appearance of the fibrin E-fragment as shown using a monoclonal antibody (FDP-14) that has its epitope in the E domain of fibrin. t-PA was shown to bind with a high affinity to both the intact (Kd = 3.3 +/- 0.6 nM) and the degraded surface of fibrin (Kd = 1.2 +/- 0.4 nM). Binding of t-PA to carboxy-terminal lysine residues of degraded fibrin was shown to be efficiently competed by physiological concentrations of plasminogen (2 microM), indicating that the affinity of t-PA for these residues was lower than that of plasminogen (Kd = 0.66 +/- 0.22 microM) and unrelated to the high affinity of t-PA for specific binding sites on intact fibrin. These data confirm and establish that the generation of carboxy-terminal lysine residues on fibrin during ongoing fibrinolysis, and the binding of plasminogen to these sites, is an important pathway in the acceleration of clot dissolution.

Amino Acid Sequence↗
PubMed1992

Apolipoprotein(a) and plasminogen interactions with fibrin: a study with recombinant apolipoprotein(a) and isolated plasminogen fragments.

Lipoprotein(a) [Lp(a)], but not low-density lipoprotein (LDL), was previously shown to impair the generation of fibrin-bound plasmin [Rouy et al. (1991) Arterioscler. Thromb. 11, 629-638] by a mechanism involving binding of Lp(a) to fibrin. It was therefore suggested that the binding was mediated by apolipoprotein(a) [apo(a)], a glycoprotein absent from LDL which has a high degree of homology with plasminogen, the precursor of the fibrinolytic enzyme plasmin. Here we have evaluated this hypothesis by performing comparative fibrin binding studies using a recombinant form of apo(a) containing 17 copies of the apo(a) domain resembling kringle 4 of plasminogen, native Lp(a), and Glu-plasminogen (Glu1-Asn791). Attempts were also made to identify the kringle domains involved in such interactions using isolated elastase-derived plasminogen fragments. The binding experiments were performed using a well-characterized model of an intact and of a plasmin-digested fibrin surface as described by Fleury and Anglés-Cano [(1991) Biochemistry 30, 7630-7638]. Binding of r-apo(a) to the fibrin surfaces was of high affinity (Kd = 26 +/- 8.4 nM for intact fibrin and 7.7 +/- 4.6 nM for plasmin-degraded fibrin) and obeyed the Langmuir equation for adsorption at interfaces. The binding to both surfaces was inhibited by the lysine analogue AMCHA and was completely abolished upon treatment of the degraded surface with carboxypeptidase B, indicating that r-apo(a) binds to both the intrachain lysines of intact fibrin and the carboxy-terminal lysines of degraded fibrin. As expected from these results, both r-apo(a) and native Lp(a) inhibited the binding of Glu-plasminogen to the fibrin surfaces.(ABSTRACT TRUNCATED AT 250 WORDS)

Apolipoproteins↗
PubMed1991

Characterization of the binding of plasminogen to fibrin surfaces: the role of carboxy-terminal lysines.

In the present study we have quantitatively characterized the interaction of purified human Glu- and Lys-plasminogen with intact and degraded fibrin by ligand-binding experiments using a radioisotopic dilution method and antibodies against human plasminogen. A fibrinogen monolayer was covalently linked to a solid support with polyglutaraldehyde and was treated with thrombin or with thrombin and then plasmin to respectively obtain intact and degraded fibrin surfaces. Under these conditions, a well-defined surface of fibrin is obtained (410 +/- 4 fmol/cm2) and, except for a 39-kDa fragment, most of the fibrin degradation products remain bound to the support. New binding sites for plasminogen were detected on the degraded surface of fibrin. These sites were identified as carboxy-terminal lysine residues both by inhibition of the binding by the lysine analogue 6-aminohexanoic acid and by carboxy-terminal end-group digestion with carboxypeptidase B. The binding curves exhibited a characteristic Langmuir adsorption isotherm saturation profile. The data were therefore analyzed accordingly, assuming a single-site binding model to simplify the analysis. Equilibrium dissociation constants (Kd) and the maximum number of binding sites (Bmax) were derived from linearized expression of the Langmuir isotherm equation. The Kd for the binding of Glu-plasminogen to intact fibrin was 0.99 +/- 0.17 microM and for degraded fibrin was 0.66 +/- 0.22 microM. The Kd for the binding of Lys-plasminogen to intact fibrin was 0.41 +/- 0.22 microM and for degraded fibrin was 0.51 +/- 0.12 microM.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗
PubMedDate not supplied

Betaine use by rhizosphere bacteria: genes essential for trigonelline, stachydrine, and carnitine catabolism in Rhizobium meliloti are located on pSym in the symbiotic region.

Rhizobium meliloti is known to use betaines synthesized by its host, Medicago sativa, as osmoprotectants and sources of energy. It is shown in the present report that the symbiotic megaplasmid (pSym) of R. meliloti RCR2011 encodes functions essential to the catabolism of three betaines, trigonelline (nicotinic acid N-methylbetaine), stachydrine (proline betaine or dimethylproline), and carnitine (gamma-trimethyl-beta-hydroxybutyrobetaine). Preliminary evidence is presented showing that functions on pSym also influence the catabolism of choline and its oxidative product, glycine betaine. Genes implicated in betaine catabolism are found in the symbiotic region of pSym. Trigonelline catabolism functions lie between two clusters of symbiotic genes, nifKDH and nok/fixVI'. Stachydrine and carnitine functions lie to the right of trigonelline catabolism functions, immediately to the right of fixVI'. Information necessary to choline and glycine betaine catabolism is probably encoded to the right of stachydrine catabolism functions.

Alkaloids↗