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J Choay

Publications and source records attributed to J Choay.

At least 73 records · Page 4Linked to original sources
PubMed1986

Synthesis of heparin fragments. A chemical synthesis of the pentasaccharide O-(2-deoxy-2-sulfamido-6-O-sulfo-alpha-D-glucopyranosyl)-(1-4 )-O-(beta-D-glucopyranosyluronic acid)-(1-4)-O-(2-deoxy-2-sulfamido-3,6-di-O-sulfo-alpha-D-glu copyranosyl)-(1-4)-O-(2-O-sulfo-alpha-L-idopyranosyluronic acid)-(1-4)-2-deoxy-2-sulfamido-6-O-sulfo-D-glucopyranose decasodium salt, a heparin fragment having high affinity for antithrombin III.

Known allyl 4,6-O-benzylidene-alpha-D-glucopyranoside was first converted into methyl (prop-1-enyl 2,3-di-O-benzyl-4-O-chloroacetyl-alpha-D-glucopyranosid)-uronate. Acid hydrolysis, followed by treatment with (bromomethylene)dimethyl-ammonium bromide, gave methyl (2,3-di-O-benzyl-4-O-chloroacetyl-alpha-D-glucopyranosyl bromide)uronate. Condensation of this bromide with 3-O-acetyl-1,6-anhydro-2-azido-2-deoxy-4-O-(methyl 2,3-di-O-benzyl-4-O-chloroacetyl-beta-D-glucopyranosyluronate)-bet a-D-glucopyranose. Acetolysis, followed by treatment with titanium tetrabromide, then gave 3,6-di-O-acetyl-2-azido-2-deoxy-4-O-(methyl 2,3-di-O-benzyl-4-O-chloroacetyl-beta-D-glucopyranosyluronate)-alp ha-D-glucopyranosyl bromide. Condensation of this bromide with benzyl 6-O-acetyl-3-O-benzyl-2-benzyloxy- carbonylamino-2-deoxy-4-O-(methyl 2-O-acetyl-3-O-benzyl-alpha-L- idopyranosyluronate)-alpha-D-glucopyranoside provided benzyl O-(methyl 2,3-di-O-benzyl-4-O-chloroacetyl-beta-D-glucopyranosyluronate)-(1- ---4)-O-(3,6-di-O-acetyl- -2-azido-2-deoxy-alpha-D-glucopyranosyl)-(1----4)-O-(methyl 2-O-acetyl-3-O-benzyl-alpha-L-idopyranosyluronate)-(1----4)-6-O-ac etyl-3-O- acetyl-3-O-benzyl-2-benzyloxycarbonylamino-2-deoxy-alpha-D-gluc opyranoside. O-Dechloroacetylation followed by condensation with 6-O-acetyl-2-azido-3,4-di-O-benzyl-2-deoxy-alpha-D-glucopyranosyl bromide provided benzyl O-(6-O-acetyl-2-azido-3,4-di-O-benzyl-2-deoxy-beta-D-glucopyranosyl)- (1----4)-O-(methyl 2,3-di-O-benzyl-beta-D-glucopyranosyluronate)-(1----4)- O-(3,6-di-O-acetyl-2-azido-2-deoxy-alpha-D-glucopyranosyl)-(1----4)-O-(m ethyl 2-O-acetyl-3-O-benzyl-alpha-L-idopyranosyluronate)-(1----4)-6-O-ac etyl-3-O- benzyl-2-benzyloxycarbonylamino-2-deoxy-alpha-D-glucopyranoside in 70% yield. O-Deacetylation followed by re-esterification, O-sulfation, saponification, catalytic hydrogenolysis, and N-sulfation gave the decasodium salt of O-(2-deoxy-2-sulfamido-6-O-sulfo-alpha-D- glucopyranosyl)-(1----4)-O-(beta-D-glucopyranosyluronic acid)-(1----4)-O-(2-deoxy-2-sulfamido-3,6-di-O-sulfo-alpha-D-gl ucopyranosyl)-(1----4)-O-(2-O-sulfo-alpha-L-idopyranosyluronic+ ++ acid)-(1----4)-2-deoxy-2-sulfamido-6-O-sulfo-D-glucopyranose. This synthetic pentasaccharide binds to antithrombin III with an association constant similar to that of high-affinity heparin and elicits a potent anti-factor Xa activity in plasma.

Antithrombin III↗
PubMed1986

Structural determinants of the capacity of heparin to inhibit the proliferation of vascular smooth muscle cells. II. Evidence for a pentasaccharide sequence that contains a 3-O-sulfate group.

Earlier work from our laboratory demonstrated that heparin inhibited the proliferation of vascular smooth muscle cells in vivo and in vitro. Both anticoagulant and non-anticoagulant heparin species were equally effective as antiproliferative agents. Previous structure-function studies indicated that hexasaccharide and larger fragments retained antiproliferative activity, whereas tetra- and disaccharides were inactive. These experiments also suggested that both N- and O-sulfates of heparin were necessary for growth inhibitory capacity. In this paper, we have further analyzed the structural determinants of the antiproliferative activity of heparin. These experiments were done using synthetically prepared and therefore chemically defined heparin oligosaccharides. We present evidence that a pentasaccharide fragment retains antiproliferative activity, and that the 3-O-sulfate on the internal glucosamine residue is critical for growth inhibitory capacity of the pentasaccharide. We also show that heparins obtained from different manufacturers differ significantly in their ability to suppress smooth muscle cell proliferation.

Animals↗
PubMed1985

Contribution of monosaccharide residues in heparin binding to antithrombin III.

The importance of 3-O- and 6-O-sulfated glucosamine residues within the heparin octasaccharide iduronic acid(1)----N-acetylglucosamine 6-O-sulfate(2)----glucuronic acid(3)----N-sulfated glucosamine 3,6-di-O-sulfate(4)----iduronic acid 2-O-sulfate(5)----N-sulfated glucosamine 6-O-sulfate(6)----iduronic acid 2-O-sulfate(7)----anhydromannitol 6-O-sulfate(8) was determined by comparing with synthetic tetra- and penta-saccharides its ability to bind human antithrombin. The octasaccharide had an affinity for antithrombin of 1 X 10(-8) M (10.2 kcal/mol) measured by intrinsic fluorescence enhancement at 6 degrees C. The synthetic pentasaccharide, consisting of residues 2-6, had an affinity of 3 X 10(-8) M (9.6 kcal/mol). The same pentasaccharide, except lacking the 3-O-sulfate on residue 4, had an affinity of 5 X 10(-4) M (4.5 kcal/mol) measured by equilibrium dialysis. The tetrasaccharide, consisting of residues 2-5, bound antithrombin with an affinity of 5 X 10(-6) M (6.8 kcal/mol). The tetrasaccharide, consisting of residues 3-6, had an affinity of 5 X 10(-5) M (5.5 kcal/mol). Since the loss of either the 6-O-sulfated residue 2 or the 3-O-sulfate of residue 4 results in a 4-5 kcal/mol or a 40-50% loss in binding energy of the pentasaccharide, these two residues must be the major contributors to the binding and must be linked to the biologic activity of the octasaccharide.

Animals↗
PubMed1985

Mono- and bidimensional 500 MHz 1H-NMR spectra of a synthetic pentasaccharide corresponding to the binding sequence of heparin to antithrombin-III: evidence for conformational peculiarity of the sulfated iduronate residue.

1H-NMR spectra of the synthetic pentasaccharide (N-sulfate-6-0-sulfate-alpha-D-glucosamine) 1----4 (beta-D-glucuronic acid) 1----4 (N-sulfate-3,6-di-0-sulfate-alpha-D-glucosamine) 1----4 (2-0-sulfate-alpha-L-iduronic acid) 1----4 (N-sulfate-6-0-sulfate-alpha-D-glucosamine), corresponding to the active site of heparin for antithrombin (AT-III), have been resolved at 500 MHz and assigned by mono- and bidimensional techniques. Vicinal proton coupling constants of the D-glucosamine residues are similar to those in the regular sequences of heparin, indicating that the 4C1 conformation of the ring, and preference for the g,g conformation of the sulfated hydroxymethyl groups of these residues are neither affected by the unique 3-0-sulfo group nor by sequence effects. By contrast, an unusually large coupling between H-2 and H-3 of the sulfated L-iduronic acid residue suggests a greater departure from the 1C4 conformation of this residue. when present in the binding sequence to AT-III than in the regular sequences. Such a departure, leading to different orientation and spacing of essential sulfate groups, may have implications for high-affinity binding to AT-III.

Antithrombin III↗
PubMed1985

Biologic studies on chemically synthesized pentasaccharide and tetrasaccharide fragments.

We have synthesized the pentasaccharide representing the binding site of heparin to AT III. This pentasaccharide contains a 3-0-sulfate group on one of its glucosamine units. A pentasaccharide analogue lacking this 3-0-sulfate group and two different tetrasaccharides, each being part of the pentasaccharide sequence with AT III affinity, have also been synthesized. The pentasaccharide with high affinity for AT III also shows high antifactor Xa activity and, in vivo, antithrombotic activity. Removal of glucosamine at the nonreducing end maintains partial binding capacity as well as partial antifactor Xa activity.

Animals↗
PubMed1983

Structure-activity relationship in heparin: a synthetic pentasaccharide with high affinity for antithrombin III and eliciting high anti-factor Xa activity.

The structures of the tetrasaccharide (beta-D-glucuronic acid)1 leads to 4 (N-sulfate-3,6-di-0-sulfate-alpha-D-glucosamine)1 leads to 4(2-0-sulfate-alpha-L-iduronic acid)1 leads to 4(N-sulfate-6-0-sulfate-D-glucosamine) and of the pentasaccharide (N-sulfate-6-0-sulfate-alpha-D-glucosamine)1 leads to 4(beta-D-glucuronic acid)1 leads to 4(N-sulfate-3,6-di-0-sulfate-alpha-D-glucosamine)1 leads to 4(2-0-sulfate-alpha-L-iduronic acid)1 leads to 4(N-sulfate-6-0-sulfate-D-glucosamine), both prepared for the first time, by chemical synthesis from D-glucose and D-glucosamine, have been confirmed by nuclear magnetic resonance. The synthetic tetrasaccharide neither binds to AT-III nor induces anti-factor Xa activity enhancement of this inhibitor. In contrast, the synthetic pentasaccharide strongly binds to AT-III (Ka: 7.10(6)M-1) forming an equimolar complex and also enhances the AT-III inhibitory activity towards factor Xa. These results confirm that the synthetic pentasaccharide with the above structure corresponds to the actual minimal sequence required in heparin for binding to AT-III.

Antithrombin III↗
PubMed1982

[Enhancement of the activity of hepatitis B virus vaccine by association with murabutide].

Murabutide (N-acetyl-muramyl-L-alanyl-D-glutamine-alpha-butylester), an MDP analogue, is a potential adjuvant for Human immunization. High levels of specific antibodies were obtained in Mouse and Guinea-Pig following administration with murabutide of low dosages of anti-hepatitis B viral vaccine containing the surface antigen (HBs). The effect of murabutide was enhanced without increasing the level of specific IgE by association with suboptimal dosages of aluminium hydroxide.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1982

Studies on a highly active anticoagulant fraction of high molecular weight isolated from porcine sodium heparin.

We have studied heparin fractionation using gel filtration and ion-exchange chromatographic methods. The starting material was commercial grade porcine mucosal sodium heparin (PSH). The fractionation was monitored employing synthetic substrates for assaying both antithrombin (with H-D-Phe-Pip-Arg-pNA ; S-2238) and anti-FXa (with Bz-Ileu-Glu-Gly-Arg-pNA ; S-2222) activities. The resulting fractions were evaluated in different amidolytic and coagulation methods used to determine heparin potency by comparison with PSH. By gel filtration of PSH on Ultrogel Aca 54, both strong anti-FXa and antithrombin activities were associated with the fractions eluted in the high molecular weight range (MW congruent to 20 x 10(3)). These fractions also had potent anticoagulant action when assayed by conventional clotting methods. PSH was also subjected to fractionation by an ion-exchange technique (DEAE-Sephacel) with increasing salt molarity. The patterns for antithrombin and anti-FXa activities were again closely related, if not identical. Four fractions were usually distinguished, with respectively negligible, intermediate, high and very high activities when compared to PSH. The very highly active fraction (HAF), approximately 15% by weight, was eluted at high salt molarity (greater than 0.8 M NaCl). On a weight basis its anticoagulant activity was congruent to 2-3 times that of PSH as determined by amidolytic as well as clotting methods. Intravenous injection of HAF to rabbits and dogs (1.0 and 2.5 mg/kg) produced a much stronger anticoagulant response than PSH, also showing an effect which persisted for a longer duration.

Animals↗
PubMed1982

Contribution of zinc and other metals to the biological activity of the serum thymic factor.

The serum thymic factor (FTS) utilized in its synthetic or natural form loses its biological activity in a rosette assay after treatment with a metal ion-chelating agent, Chelex 100. This activity is restored by the addition of Zn salts and, to a lesser extent, certain other metal salts. FTS activation is secondary to the binding of the metal to the peptide. The metal-to-peptide molar ratio of 1:1 provides the best activation. These data indicate the existence of two forms of FTS. The first one lacks Zn and is biologically inactive; the second one contains Zn and is biologically active, for which we propose the name of "thymulin" (FTS-Zn). The presence of Zn in synthetic FTS was confirmed by atomic absorption spectrometry. The interaction between Zn and FTS was further suggested by microanalysis demonstrating the presence of this metal in thymic reticuloepithelial cells.

Animals↗
PubMed1981

The structure of heparin oligosaccharide fragments with high anti-(factor Xa) activity containing the minimal antithrombin III-binding sequence. Chemical and 13C nuclear-magnetic-resonance studies.

The chemical composition and the 13C n.m.r. spectra of heparin oligosaccharides (essentially octasaccharides), having high affinity for antithrombin III and high anti-(Factor Xa) activity, prepared by three independent approaches (extraction, partial deaminative cleavage with HNO2 and partial depolymerization with bacterial heparinase), leading to different terminal residues, have been studied and compared with those of the corresponding inactive species. Combined wit chemical data, the spectra of the active oligosaccharides and of their fragmentation products afforded information on composition and sequence. The three types of active oligosaccharides were shown to have the common hexasaccharide core I-Aa-G-As*-Is-As, where I and alpha-L-idopyranosyl-uronic acid, Aa = 2-acetamido-2-deoxy-alpha-D-glucopyranose, G = beta-D-glucopyranosyl-uronic acid, Is = alpha-L-idopyranosyluronic acid 2-O-sulphate, As = 2-deoxy-2-sulphamino-alpha-D-glucopyranose 6-O-sulphate. The fourth residue (As*) is an unusually substituted amino sugar resistant to mild deamination. The 13C spectra of the active species are characterized by signals from the above atypical amino sugar, the most evident of which is at 57.7 p.p.m. These signals, compared with those of appropriate synthetic model compounds, are compatible with the recently proposed 3-O-sulphation of the residue As* [Lindahl, Bäckström, Thunberg & Leder (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 6551-6555].

Animals↗