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Hiromu Yawo - One of the best experts on this subject based on the ideXlab platform.

  • μ opioid receptor inhibits n type ca2 channels in the calyx presynaptic terminal of the embryonic chick ciliary ganglion
    The Journal of Physiology, 2000
    Co-Authors: Katsuaki Endo, Hiromu Yawo
    Abstract:

    1A study was made on the mechanisms by which Enkephalins inhibit synaptic transmission at calyx-type presynaptic terminals in the ciliary ganglion of chick embryos at stages 39–40. 2Excitatory postsynaptic currents (EPSCs) were recorded by nystatin-perforated patch clamp at low [Ca2+]o and high [Mg2+]o. [Leu5]Enkephalin (L-ENK, 1–10 μM) reduced the quantal content (m) without changing the quantal size (q). This effect was antagonized by naloxone (1 μM). Similar results were observed under conventional whole-cell clamp of the postsynaptic neuron. 3A specific agonist of the μ-opioid receptor, [D-Ala2, M-Me-Phe4,Gly5]Enkephalin-ol (DAMGO) reduced m without changing q. A specific agonist of the δ-opioid receptor, [d-Pen2, d-Pen5]Enkephalin (DPDPE) also reduced m without changing q. 4Both L-ENK and [Met5]Enkephalin (M-ENK) reduced the stimulus-dependent increment of the intraterminal Ca2+ concentration (Δ[Ca2+]t) without affecting the decay time constant of the intraterminal Ca2+ concentration and basal Ca2+ level. This effect was antagonized by naloxone. DAMGO reduced Δ[Ca2+]t more effectively than DPDPE. 5When extracellular Ca2+ was replaced by Ba2+, the stimulus-dependent increment of the intraterminal Ba2+ concentration (Δ[Ba2+]t) was also reduced by L-ENK or DAMGO. 6L-ENK reduced Δ[Ca2+]t even in the presence of 4-aminopyridine (4-AP), which blocks the transient K+ conductance during the falling phase of the presynaptic action potential. When N-type Ca2+ channels were blocked by ω-conotoxin GVIA (ω-CgTxGVIA), the Δ[Ca2+]t was no longer sensitive to L-ENK and DAMGO. 7It is suggested that Enkephalins reduce the transmitter release through presynaptic opioid receptors. The μ-opioid receptor may suppress presynaptic Ca2+ influx by selectively inhibiting N-type Ca2+ channels.

  • μ opioid receptor inhibits n type ca2 channels in the calyx presynaptic terminal of the embryonic chick ciliary ganglion
    The Journal of Physiology, 2000
    Co-Authors: Katsuaki Endo, Hiromu Yawo
    Abstract:

    A study was made on the mechanisms by which Enkephalins inhibit synaptic transmission at calyx-type presynaptic terminals in the ciliary ganglion of chick embryos at stages 39-40. Excitatory postsynaptic currents (EPSCs) were recorded by nystatin-perforated patch clamp at low [Ca2+]o and high [Mg2+]o. [Leu5]Enkephalin (L-ENK, 1-10 microM) reduced the quantal content (m) without changing the quantal size (q). This effect was antagonized by naloxone (1 microM). Similar results were observed under conventional whole-cell clamp of the postsynaptic neuron. A specific agonist of the mu-opioid receptor, [D-Ala2, M-Me-Phe4,Gly5]Enkephalin-ol (DAMGO) reduced m without changing q. A specific agonist of the delta-opioid receptor, [d-Pen2, d-Pen5]Enkephalin (DPDPE) also reduced m without changing q. Both L-ENK and [Met5]Enkephalin (M-ENK) reduced the stimulus-dependent increment of the intraterminal Ca2+ concentration (Delta[Ca2+]t) without affecting the decay time constant of the intraterminal Ca2+ concentration and basal Ca2+ level. This effect was antagonized by naloxone. DAMGO reduced Delta[Ca2+]t more effectively than DPDPE. When extracellular Ca2+ was replaced by Ba2+, the stimulus-dependent increment of the intraterminal Ba2+ concentration (Delta[Ba2+]t) was also reduced by L-ENK or DAMGO. L-ENK reduced Delta[Ca2+]t even in the presence of 4-aminopyridine (4-AP), which blocks the transient K+ conductance during the falling phase of the presynaptic action potential. When N-type Ca2+ channels were blocked by omega-conotoxin GVIA (omega-CgTxGVIA), the Delta[Ca2+]t was no longer sensitive to L-ENK and DAMGO. It is suggested that Enkephalins reduce the transmitter release through presynaptic opioid receptors. The mu-opioid receptor may suppress presynaptic Ca2+ influx by selectively inhibiting N-type Ca2+ channels.

Istvan Toth - One of the best experts on this subject based on the ideXlab platform.

  • A Drug Delivery Strategy: Binding Enkephalin to Asialoglycoprotein Receptor by Enzymatic
    2016
    Co-Authors: Michelle P Christie, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Lauren E. Hartley-tassell, Christopher J. Day, Freda -c. E. Jen, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-59-diphosphogalactose 4-epimerase and lipopolysaccharyl a-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60 % remained stable after a 2 hr incubation at 37uC). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.161028 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 mM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemente

  • a drug delivery strategy binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PLOS ONE, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Lauren E Hartleytassell, Michael P Jennings, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5'-diphosphogalactose 4-epimerase and lipopolysaccharyl alpha-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37 degrees C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono-and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1x10(-8) cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (K-D=91 mu M). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.

  • A drug delivery strategy: binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PloS one, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Freda E.-c. Jen, Lauren E. Hartley-tassell, Christopher J. Day, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5′-diphosphogalactose 4-epimerase and lipopolysaccharyl α-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37°C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1×10−8 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 µM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.

  • determination of transport in the caco 2 cell assay of compounds varying in lipophilicity using lc ms enhanced transport of leu Enkephalin analogues
    European Journal of Pharmaceutical Sciences, 2002
    Co-Authors: Allan Wong, Benjamin P Ross, Yiu Ngok Chan, Per Artursson, Lucia Lazorova, Alun Jones, Istvan Toth
    Abstract:

    Purpose: To synthesize a number of analogues of Leu-Enkephalin with different lipophilicities and to develop an LC–MS method for determining the Caco-2 cell permeability values of these compounds. Methods: A number of sugar and sugar plus lipoamino acid analogues of Leu-Enkephalin were synthesized by solid-phase and solution methods. An LC–MS method was developed for analyzing the Caco-2 cell assay samples and validated against the traditional method using radiolabelled compounds. Results: A sensitive and specific LC–MS assay was developed. Standard curves were linear in the range of 0.025–5 μM. Apparent permeability values determined by LC–MS and liquid scintillation counter were identical, for both a hydrophilic drug, cephalexin and a lipophilic Leu-enkaphalin analogue. Caco-2 permeability values for the analogues of Leu-Enkephalin were determined and it was found that attachment of sugar or sugar and lipoamino acid to the Leu-Enkephalin peptide resulted in an increase in the apparent permeability values compared to the native peptide, which was not transported across the Caco-2 cell monolayers. Conclusions: A rapid, generic LC–MS method for analyzing a range of compounds was developed. Attachment of a sugar or sugar and lipoamino acid to Leu-Enkephalin improves the apparent permeability across Caco-2 cell monolayers.

Katsuaki Endo - One of the best experts on this subject based on the ideXlab platform.

  • μ opioid receptor inhibits n type ca2 channels in the calyx presynaptic terminal of the embryonic chick ciliary ganglion
    The Journal of Physiology, 2000
    Co-Authors: Katsuaki Endo, Hiromu Yawo
    Abstract:

    1A study was made on the mechanisms by which Enkephalins inhibit synaptic transmission at calyx-type presynaptic terminals in the ciliary ganglion of chick embryos at stages 39–40. 2Excitatory postsynaptic currents (EPSCs) were recorded by nystatin-perforated patch clamp at low [Ca2+]o and high [Mg2+]o. [Leu5]Enkephalin (L-ENK, 1–10 μM) reduced the quantal content (m) without changing the quantal size (q). This effect was antagonized by naloxone (1 μM). Similar results were observed under conventional whole-cell clamp of the postsynaptic neuron. 3A specific agonist of the μ-opioid receptor, [D-Ala2, M-Me-Phe4,Gly5]Enkephalin-ol (DAMGO) reduced m without changing q. A specific agonist of the δ-opioid receptor, [d-Pen2, d-Pen5]Enkephalin (DPDPE) also reduced m without changing q. 4Both L-ENK and [Met5]Enkephalin (M-ENK) reduced the stimulus-dependent increment of the intraterminal Ca2+ concentration (Δ[Ca2+]t) without affecting the decay time constant of the intraterminal Ca2+ concentration and basal Ca2+ level. This effect was antagonized by naloxone. DAMGO reduced Δ[Ca2+]t more effectively than DPDPE. 5When extracellular Ca2+ was replaced by Ba2+, the stimulus-dependent increment of the intraterminal Ba2+ concentration (Δ[Ba2+]t) was also reduced by L-ENK or DAMGO. 6L-ENK reduced Δ[Ca2+]t even in the presence of 4-aminopyridine (4-AP), which blocks the transient K+ conductance during the falling phase of the presynaptic action potential. When N-type Ca2+ channels were blocked by ω-conotoxin GVIA (ω-CgTxGVIA), the Δ[Ca2+]t was no longer sensitive to L-ENK and DAMGO. 7It is suggested that Enkephalins reduce the transmitter release through presynaptic opioid receptors. The μ-opioid receptor may suppress presynaptic Ca2+ influx by selectively inhibiting N-type Ca2+ channels.

  • μ opioid receptor inhibits n type ca2 channels in the calyx presynaptic terminal of the embryonic chick ciliary ganglion
    The Journal of Physiology, 2000
    Co-Authors: Katsuaki Endo, Hiromu Yawo
    Abstract:

    A study was made on the mechanisms by which Enkephalins inhibit synaptic transmission at calyx-type presynaptic terminals in the ciliary ganglion of chick embryos at stages 39-40. Excitatory postsynaptic currents (EPSCs) were recorded by nystatin-perforated patch clamp at low [Ca2+]o and high [Mg2+]o. [Leu5]Enkephalin (L-ENK, 1-10 microM) reduced the quantal content (m) without changing the quantal size (q). This effect was antagonized by naloxone (1 microM). Similar results were observed under conventional whole-cell clamp of the postsynaptic neuron. A specific agonist of the mu-opioid receptor, [D-Ala2, M-Me-Phe4,Gly5]Enkephalin-ol (DAMGO) reduced m without changing q. A specific agonist of the delta-opioid receptor, [d-Pen2, d-Pen5]Enkephalin (DPDPE) also reduced m without changing q. Both L-ENK and [Met5]Enkephalin (M-ENK) reduced the stimulus-dependent increment of the intraterminal Ca2+ concentration (Delta[Ca2+]t) without affecting the decay time constant of the intraterminal Ca2+ concentration and basal Ca2+ level. This effect was antagonized by naloxone. DAMGO reduced Delta[Ca2+]t more effectively than DPDPE. When extracellular Ca2+ was replaced by Ba2+, the stimulus-dependent increment of the intraterminal Ba2+ concentration (Delta[Ba2+]t) was also reduced by L-ENK or DAMGO. L-ENK reduced Delta[Ca2+]t even in the presence of 4-aminopyridine (4-AP), which blocks the transient K+ conductance during the falling phase of the presynaptic action potential. When N-type Ca2+ channels were blocked by omega-conotoxin GVIA (omega-CgTxGVIA), the Delta[Ca2+]t was no longer sensitive to L-ENK and DAMGO. It is suggested that Enkephalins reduce the transmitter release through presynaptic opioid receptors. The mu-opioid receptor may suppress presynaptic Ca2+ influx by selectively inhibiting N-type Ca2+ channels.

Michelle P Christie - One of the best experts on this subject based on the ideXlab platform.

  • A Drug Delivery Strategy: Binding Enkephalin to Asialoglycoprotein Receptor by Enzymatic
    2016
    Co-Authors: Michelle P Christie, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Lauren E. Hartley-tassell, Christopher J. Day, Freda -c. E. Jen, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-59-diphosphogalactose 4-epimerase and lipopolysaccharyl a-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60 % remained stable after a 2 hr incubation at 37uC). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.161028 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 mM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemente

  • a drug delivery strategy binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PLOS ONE, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Lauren E Hartleytassell, Michael P Jennings, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5'-diphosphogalactose 4-epimerase and lipopolysaccharyl alpha-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37 degrees C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono-and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1x10(-8) cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (K-D=91 mu M). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.

  • A drug delivery strategy: binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PloS one, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Freda E.-c. Jen, Lauren E. Hartley-tassell, Christopher J. Day, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5′-diphosphogalactose 4-epimerase and lipopolysaccharyl α-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37°C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1×10−8 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 µM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.

Michael P Jennings - One of the best experts on this subject based on the ideXlab platform.

  • A Drug Delivery Strategy: Binding Enkephalin to Asialoglycoprotein Receptor by Enzymatic
    2016
    Co-Authors: Michelle P Christie, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Lauren E. Hartley-tassell, Christopher J. Day, Freda -c. E. Jen, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-59-diphosphogalactose 4-epimerase and lipopolysaccharyl a-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60 % remained stable after a 2 hr incubation at 37uC). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.161028 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 mM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemente

  • a drug delivery strategy binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PLOS ONE, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Lauren E Hartleytassell, Michael P Jennings, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5'-diphosphogalactose 4-epimerase and lipopolysaccharyl alpha-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37 degrees C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono-and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1x10(-8) cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (K-D=91 mu M). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.

  • A drug delivery strategy: binding Enkephalin to asialoglycoprotein receptor by enzymatic galactosylation
    PloS one, 2014
    Co-Authors: Michelle P Christie, Pavla Simerska, Waleed M Hussein, Mohamad F M Rawi, Michael P Jennings, Freda E.-c. Jen, Lauren E. Hartley-tassell, Christopher J. Day, Istvan Toth
    Abstract:

    Glycosylation of biopharmaceuticals can mediate cell specific delivery by targeting carbohydrate receptors. Additionally, glycosylation can improve the physico-chemical (drug-like) properties of peptide based drug candidates. The main purpose of this study was to examine if glycosylation of the peptide Enkephalin could facilitate its binding to the carbohydrate receptor, asialoglycoprotein. Firstly, we described the one-pot enzymatic galactosylation of lactose modified Enkephalin in the presence of uridine-5′-diphosphogalactose 4-epimerase and lipopolysaccharyl α-1,4-galactosyltransferase. Stability experiments using human plasma and Caco-2 cell homogenates showed that glycosylation considerably improved the stability of Enkephalin (at least 60% remained stable after a 2 hr incubation at 37°C). In vitro permeability experiments using Caco-2 cells revealed that the permeability of mono- and trisaccharide conjugated Enkephalins was 14 and 28 times higher, respectively, than that of Enkephalin alone (Papp 3.1×10−8 cm/s). By the methods of surface plasmon resonance and molecular modeling, we demonstrated that the enzymatic glycosylation of Enkephalin enabled binding the asialoglycoprotein receptor. The addition of a trisaccharide moiety to Enkephalin improved the binding of Enkephalin to the asialoglycoprotein receptor two fold (KD = 91 µM). The docking scores from molecular modeling showed that the binding modes and affinities of the glycosylated Enkephalin derivatives to the asialoglycoprotein receptor complemented the results from the surface plasmon resonance experiments.