Arylsulfatase A

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Volkmar Gieselmann - One of the best experts on this subject based on the ideXlab platform.

  • brAin cell type specific endocytosis of ArylsulfAtAse A identifies limitAtions of enzyme bAsed therApies for metAchromAtic leukodystrophy
    Human Molecular Genetics, 2021
    Co-Authors: Debora Kaminski, Frank Matthes, Volkmar Gieselmann, Claudia Yaghootfam, Annika Resing, Ulrich Matzner
    Abstract:

    Enzyme replAcement therApies, Allogeneic bone mArrow trAnsplAntAtion And gene therApies Are treAtment options for lysosomAl storAge diseAses cAused by inherited deficiencies of soluble lysosomAl enzymes. Independent from the ApproAch, the enzyme must be delivered to lysosomes of deficient pAtient cells. Little is known About the disseminAtion of enzyme within A tissue where cells compete for uptAke viA different receptor systems, binding Affinities And endocytic rAtes. To evAluAte disseminAtion And lysosomAl tArgeting of A lysosomAl enzyme in the CNS, we AnAlysed receptor-mediAted endocytosis of ArylsulfAtAse A (ASA) by different types of brAin-derived cell lines And primAry murine brAin cells. For ASA expressed by chinese hAmster ovAry cells for enzyme replAcement therApy of metAchromAtic leukodystrophy, endocytic rAtes decline from microgliA to neurons And Astrocytes And to oligodendrocytes. Only immAture oligodendrocytes endocytose significAnt Amounts of enzyme. UptAke by non-microgliAl cells is due to mAnnose 6-phosphAte receptors, whereAs severAl receptor systems pArticipAte in endocytosis by microgliAl cells. Interestingly, ASA expressed by microgliAl cells cAnnot be tAken up in A mAnnose 6-phosphAte dependent mAnner. The resulting fAilure to correct non-microgliAl cells corroborAtes in vivo dAtA And indicAtes thAt therApeutic effects of Allogeneic bone mArrow trAnsplAntAtion And hemAtopoietic stem cell gene therApy on metAchromAtic leukodystrophy Are independent of metAbolic cross-correction of neurons, Astrocytes And oligodendrocytes by receptor-mediAted endocytosis.

  • evolutionAry redesign of the lysosomAl enzyme ArylsulfAtAse A increAses efficAcy of enzyme replAcement therApy for metAchromAtic leukodystrophy
    Human Molecular Genetics, 2019
    Co-Authors: Heidi Simonis, Claudia Yaghootfam, Volkmar Gieselmann, Marc Sylvester, Ulrich Matzner
    Abstract:

    Protein engineering is A meAns to optimize protein therApeutics developed for the treAtment of so fAr incurAble diseAses including cAncers And genetic disorders. Here we report on An engineering ApproAch in which we successfully increAsed the cAtAlytic rAte constAnt of An enzyme thAt is presently evAluAted in enzyme replAcement therApies (ERT) of A lysosomAl storAge diseAse (LSD). Although ERT is A treAtment option for mAny LSDs, outcomes Are lAgging fAr behind expectAtions for most of them. This hAs been Ascribed to insufficient enzyme Activities AccumulAting in tissues difficult to tArget such As brAin And peripherAl nerves. We show for humAn ArylsulfAtAse A (hARSA) thAt the Activity of A therApeutic enzyme cAn be substAntiAlly increAsed by reversing Activity-diminishing And by inserting Activity-promoting Amino Acid substitutions thAt hAd occurred in the evolution of hominids And non-humAn mAmmAls, respectively. The potentiAl of this ApproAch, here designAted As evolutionAry redesign, wAs highlighted by the observAtion thAt murinizAtion of only 1 or 3 Amino Acid positions increAsed the hARSA Activity 3- And 5-fold, with little impAct on stAbility, respectively. The two kineticAlly optimized hARSA vAriAnts showed no immunogenic potentiAl in ERT of A humAnized ARSA knockout mouse model of metAchromAtic leukodystrophy (MLD) And reduced lysosomAl storAge of kidney, peripherAl And centrAl nervous system up to 3-fold more efficiently thAn wild-type hARSA. Due to their sAfety profile And higher therApeutic potentiAl the engineered hARSA vAriAnts might represent mAjor AdvAnces for future enzyme-bAsed therApies of MLD And stimulAte AnAlogous ApproAches for other enzyme therApeutics.

  • potentiAl of surfActAnt coAted nAnopArticles to improve brAin delivery of ArylsulfAtAse A
    Journal of Controlled Release, 2017
    Co-Authors: Tilman Schuster, Volkmar Gieselmann, Astrid Muhlstein, Claudia Yaghootfam, Olga Maksimenko, E V Shipulo, Svetlana Gelperina, Jorg Kreuter, Ulrich Matzner
    Abstract:

    AbstrAct The lysosomAl storAge disorder (LSD) metAchromAtic leukodystrophy (MLD) is cAused by A deficiency of the soluble, lysosomAl hydrolAse ArylsulfAtAse A (ASA). The diseAse is chArActerized by AccumulAtion of 3-O-sulfogAlActosylcerAmide (sulfAtide), progressive demyelinAtion of the nervous system And premAture deAth. Enzyme replAcement therApy (ERT), bAsed on regulAr intrAvenous injections of recombinAnt functionAl enzyme, is in clinicAl use for severAl LSDs. For MLD And other LSDs with centrAl nervous system (CNS) involvement, however, ERT is limited by the blood-brAin bArrier (BBB) restricting trAnsport of therApeutic enzymes from the blood to the brAin. In the present study, the potentiAl of different types of surfActAnt-coAted biodegrAdAble nAnopArticles to increAse brAin delivery of ASA wAs evAluAted. Three different strAtegies to bind ASA to nAnopArticle surfAces were compAred: (1) Adsorption, (2) high-Affinity binding viA the streptAvidin-biotin system, And (3) covAlent binding. Adsorption Allowed binding of high Amounts of Active ASA. However, in presence of phosphAte-buffered sAline or serum rApid And complete desorption occurred, rendering this strAtegy ineffective for in vivo ApplicAtions. In contrAst, stAble immobilizAtion with negligible dissociAtion wAs Achieved by high-Affinity And covAlent binding. Consequently, we AnAlyzed the brAin tArgeting of two stAbly nAnopArticle-bound ASA formulAtions in ASA−/− mice, An AnimAl model of MLD. CompAred to free ASA, injected As A control, the biodistribution of nAnopArticle-bound ASA wAs Altered in peripherAl orgAns, but no increAse of brAin levels wAs detectAble. The fAilure to improve brAin delivery suggests thAt the ASA glycoprotein interferes with processes required to tArget surfActAnt-coAted nAnopArticles to brAin cApillAry endotheliAl cells.

  • ArylsulfAtAse A overexpressing humAn ipsc derived neurAl cells reduce cns sulfAtide storAge in A mouse model of metAchromAtic leukodystrophy
    Molecular Therapy, 2015
    Co-Authors: Jonas Doerr, Ulrich Matzner, Volkmar Gieselmann, Annika Bockenhoff, Benjamin Ewald, Julia Ladewig, Matthias Eckhardt, Oliver Brustle, Philipp Koch
    Abstract:

    MetAchromAtic leukodystrophy (MLD) is An inherited lysosomAl storAge disorder resulting from A functionAl deficiency of ArylsulfAtAse A (ARSA), An enzyme thAt cAtAlyzes desulfAtion of 3-O-sulfogAlActosylcerAmide (sulfAtide). LAck of Active ARSA leAds to the AccumulAtion of sulfAtide in oligodendrocytes, SchwAnn cells And some neurons And triggers progressive demyelinAtion, the neuropAthologicAl hAllmArk of MLD. SeverAl therApeutic ApproAches hAve been explored, including enzyme replAcement, Autologous hemAtopoietic stem cell-bAsed gene therApy, intrAcerebrAl gene therApy or cell-bAsed gene delivery into the centrAl nervous system (CNS). However, long-term treAtment of the blood-brAin-bArrier protected CNS remAins chAllenging. Here we used MLD pAtient-derived induced pluripotent stem cells (iPSCs) to generAte long-term self-renewing neuroepitheliAl stem cells And AstrogliAl progenitors for cell-bAsed ARSA replAcement. Following trAnsplAntAtion of ARSA-overexpressing precursors into ARSA-deficient mice we observed A significAnt reduction of sulfAtide storAge up to A distAnce of 300 µm from grAfted cells. Our dAtA indicAte thAt neurAl precursors generAted viA reprogrAmming from MLD pAtients cAn be engineered to AmeliorAte sulfAtide AccumulAtion And mAy thus serve As Autologous cell-bAsed vehicle for continuous ARSA supply in MLD-Affected brAin tissue.

  • compArison of five peptide vectors for improved brAin delivery of the lysosomAl enzyme ArylsulfAtAse A
    The Journal of Neuroscience, 2014
    Co-Authors: Annika Bockenhoff, Volkmar Gieselmann, Philipp Wolte, Hansjoachim Galla, Sandra Cramer, Simeon Knieling, Claudia Wohlenberg, Ulrich Matzner
    Abstract:

    Enzyme replAcement therApy (ERT) is A treAtment option for lysosomAl storAge disorders (LSDs) cAused by deficiencies of soluble lysosomAl enzymes. ERT depends on receptor-mediAted trAnsport of intrAvenously injected recombinAnt enzyme to lysosomes of pAtient cells. The blood–brAin bArrier (BBB) prevents efficient trAnsfer of therApeutic polypeptides from the blood to the brAin pArenchymA And thus hinders effective treAtment of LSDs with CNS involvement. We compAred the potentiAl of five brAin-tArgeting peptides to promote brAin delivery of the lysosomAl enzyme ArylsulfAtAse A (ASA). Fusion proteins between ASA And the protein trAnsduction domAin of the humAn immunodeficiency virus TAT protein (TAt), An Angiopep peptide (Ang-2), And the receptor-binding domAins of humAn Apolipoprotein B (ApoB) And ApoE (two versions, ApoE-I And ApoE-II) were generAted. All ASA fusion proteins were enzymAticAlly Active And tArgeted to lysosomes when Added to cultured cells. In contrAst to wild-type ASA, which is tAken up by mAnnose-6-phosphAte receptors, All chimeric proteins were AdditionAlly endocytosed viA mAnnose-6-phosphAte-independent routes. For ASA-Ang-2, ASA-ApoE-I, And ASA-ApoE-II, uptAke wAs pArtiAlly due to the low-density lipoprotein receptor-relAted protein 1. TrAnsendotheliAl trAnsfer in A BBB cell culture model wAs elevAted for ASA-ApoB, ASA-ApoE-I, And ASA-ApoE-II. BrAin delivery wAs, however, increAsed only for ASA-ApoE-II. ApoE-II wAs Also superior to wild-type ASA in reducing lysosomAl storAge in the CNS of ASA-knock-out mice treAted by ERT. Therefore, the ApoE-derived peptide AppeArs useful to treAt metAchromAtic leukodystrophy And possibly other neurologicAl disorders more efficiently.

Nongnuj Tanphaichitr - One of the best experts on this subject based on the ideXlab platform.

  • ArylsulfAtAse A deficiency cAuses seminolipid AccumulAtion And A lysosomAl storAge disorder in sertoli cells
    Journal of Lipid Research, 2011
    Co-Authors: Kessiri Kongmanas, Kym F Faull, Suraj Kadunganattil, Charles E Smith, Tony Rupar, Naoko Gotoinoue, Louis Hermo, Nongnuj Tanphaichitr
    Abstract:

    SulfogAlActosylglycerolipid (SGG) is the mAjor sulfoglycolipid of mAle germ cells. During spermAtogenesis, Apoptosis occurs in >50% of totAl germ cells. Sertoli cells phAgocytose these Apoptotic germ cells And degrAde their components using lysosomAl enzymes. Here we demonstrAted thAt SGG wAs A physiologicAl substrAte of Sertoli lysosomAl ArylsulfAtAse A (ARSA). SGG AccumulAted in Sertoli cells of ArsA−/− mice, And At 8 months of Age, this buildup led to lysosomAl swelling And other cellulAr AbnormAlities typicAl of A lysosomAl storAge disorder. This disorder likely compromised Sertoli cell functions, mAnifesting As impAired spermAtogenesis And production of sperm with neAr-zero fertilizing Ability in vitro. Fecundity of ArsA−/− mAles wAs thus reduced when they were older thAn 5 months. Sperm SGG is known for its roles in fertilizAtion. Therefore, the minimAl sperm fertilizing Ability of 8-month-old ArsA−/− mAles mAy be explAined by the 50% reduction of their sperm SGG levels, A result thAt wAs Also observed in testiculAr germ cells. These unexpected decreAses in SGG levels might be pArtly due to depletion of the bAckbone lipid pAlmitylpAlmitoylglycerol thAt is generAted from the SGG degrAdAtion pAthwAy in Sertoli cells And normAlly recycled to new generAtions of primAry spermAtocytes for SGG synthesis.

  • interAction of ArylsulfAtAse A AsA with its nAturAl sulfoglycolipid substrAtes A computAtionAl And site directed mutAgenesis study
    Glycoconjugate Journal, 2009
    Co-Authors: Matthias Schenk, Daniela Costa Santos, Nongnuj Tanphaichitr, Euridice Carmona, Chaitanya A K Koppisetty, Smita Bhatia, Pergeorg Nyholm
    Abstract:

    ArylsulfAtAse A (ASA) hydrolyzes sulfAte esters with A pH optimum of 5. InterActions between p-nitrocAtechol sulfAte (NCS, ArtificiAl substrAte) And Active site residues of ASA Are reveAled from their co-crystAl structure. Since equivAlent ASA interActions with its nAturAl substrAtes, sulfogAlActosylcerAmide (SGC) And sulfogAlActosylglycerolipid (SGG), Are not known, we computAtionAlly docked SGC/SGG to the ASA crystAl structure. Our dockings suggested thAt Cys69 wAs the Active site residue, And Lys302 & Lys123 As residues Anchoring the sulfAte group of SGC/SGG to the Active site, As observed for NCS. We further confirmed these results using 2 recombinAnt ASA mutAnts: C69A And CKK (Cys69, Lys302 And Lys123-All mutAted to AlA). Both ASA mutAnts fAiled to desulfAte SGC/SGG, And CKK showed minimAl binding to [14C]SGC, Although C69A still hAd Affinity for this sulfoglycolipid. However, our dockings suggested AdditionAl intermoleculAr hydrogen bonding And hydrophobic interActions between ASA And SGC/SGG, thus contributing to the specificity of SGC/SGG As nAturAl substrAtes.

  • sperm surfAce ArylsulfAtAse A cAn disperse the cumulus mAtrix of cumulus oocyte complexes
    Journal of Cellular Physiology, 2007
    Co-Authors: Araya Anupriwan, Daniela Costa Santos, Tony Rupar, Euridice Carmona, Sitthichai Iamsaard, Krittalak Chakrabandhu, Benjamin K Tsang, Nongnuj Tanphaichitr
    Abstract:

    Cumulus cell lAyers of expAnded cumulus oocyte complexes (COCs) Are interlinked with networks of hyAluronic Acid, chondroitin sulfAte B proteoglycAns And link proteins, And they cAn be dispersed by sperm surfAce hyAluronidAses. In this report, we showed thAt ArylsulfAtAse A (AS-A), existing on the sperm heAd surfAce, Also hAd this dispersion Action. Purified AS-A free of proteAse, hyAluronidAse And chondroitinAse Activities could disperse the cumulus mAtrix of expAnded COCs. However, this COC dispersion Action wAs not AssociAted with AS-A desulfAtion Activity, AssAyed by using p-nitrocAtecholsulfAte (ArtificiAl substrAte). COCs incubAted for 1 h with sperm pretreAted with Anti-AS-A IgG in the presence of Apigenin (A hyAluronidAse inhibitor) did not exhibit mAtrix dispersion, whereAs severAl cumulus lAyers were AlreAdy dispersed in COCs incubAted with sperm pretreAted with preimmune IgG. Furthermore, sperm from AS-A null mice showed A significAnt delAy in COC dispersion, compAred with wild-type sperm. Within 1 h of sperm-COC co-incubAtion, the size of COCs incubAted with AS-A null sperm wAs 65% of the originAl dimension, whereAs thAt of COCs inseminAted with wild-type sperm wAs only 17%. A further delAy in COC dispersion by AS-A(-/-) mouse sperm wAs observed when Apigenin wAs present in the co-incubAtion. We Also showed for the first time thAt AS-A hAd A specific Affinity for chondroitin sulfAte B, A component of cumulus mAtrix proteoglycAn networks; this might provide A mechAnism of cumulus mAtrix destAbilizAtion induced by sperm surfAce AS-A.

  • Acquisition of ArylsulfAtAse A onto the mouse sperm surfAce during epididymAl trAnsit
    Biology of Reproduction, 2003
    Co-Authors: Wattana Weerachatyanukul, Araya Anupriwan, Louis Hermo, Euridice Carmona, Michael G Wade, Solange Maria Da Silva, Peter Rippstein, Prasert Sobhon, Prapee Sretarugsa, Nongnuj Tanphaichitr
    Abstract:

    ArylsulfAtAse A (AS-A) is locAlized to the sperm surfAce And pArticipAtes in sperm-zonA pellucidA binding. We investigAted how AS-A, usuAlly known As An AcrosomAl enzyme, trAfficked to the sperm surfAce. Immunocytochemistry of the mouse testis confirmed the existence of AS-A in the AcrosomAl region of round And elongAting spermAtids. However, immunofluorescence And flow cytometry indicAted the Absence of AS-A on the surfAce of live testiculAr sperm. In contrAst, positive AS-A stAining wAs observed in the heAds of live cAudAl epididymAl And vAs deferens sperm. The results suggested thAt Acquisition of AS-A on the sperm surfAce occurred during epididymAl trAnsit. Immunocytochemistry of the epididymis reveAled AS-A in nArrow And ApicAl cells in the initiAl segment And in cleAr cells in All epididymAl regions. However, these epitheliAl cells Are in the minority And Are not involved in secretory Activity. In the cAudAl epididymis And vAs deferens, AS-A wAs Also locAlized to principAl cells, the mAjor epitheliAl cells. BecAuse principAl cells hAve secretory Activity, they mAy secrete AS-A into the epididymAl fluid. This hypothesis wAs supported by our results reveAling the presence of AS-A in the epididymAl And vAs deferens fluid (determined by immunoblotting And ELISA) And An AS-A trAnscript in the epididymis (by reverse trAnscription polymerAse chAin reAction). AlexA-430 AS-A bound to epididymAl sperm with high Affinity (Kd = 46 nM). This binding wAs inhibited by treAtment of sperm with An Antibody AgAinst sperm surfAce sulfogAlActosylglycerolipid. This finding suggests thAt AS-A in the epididymAl fluid mAy deposit onto sperm viA its Affinity to sulfogAlActosylglycerolipid.

  • role of sperm surfAce ArylsulfAtAse A in mouse sperm zonA pellucidA binding
    Biology of Reproduction, 2002
    Co-Authors: Julierut Tantibhedhyangkul, Araya Anupriwan, Wattana Weerachatyanukul, Euridice Carmona, Hongbin Xu, Dominick Michaud, Nongnuj Tanphaichitr
    Abstract:

    We hAve previously described the zonAe pellucidAe (ZP) binding Ability of A pig sperm surfAce protein, P68. Our recent results on peptide sequencing of 3 P68 tryptic peptides And moleculAr cloning of pig testis ArylsulfAtAse A (AS-A) reveAled the identity of P68 As AS-A. In this report, we demonstrAte the presence of AS-A on the mouse sperm surfAce And its role in ZP binding. Using Anti-AS-A Antibody, we hAve shown by immunoblotting thAt AS-A wAs present in A Triton X-100 extrAct of mouse sperm. The presence of AS-A on the sperm plAsmA membrAne wAs conclusively demonstrAted by indirect immunofluorescence, immunogold electron microscopy, And AS-A’s desulfAtion Activity on live mouse sperm. The AS-A remAined on the heAd surfAce of in vivo cApAcitAted sperm, As reveAled by positive immunofluorescent stAining of oviductAl/uterine sperm. SignificAntly, the role of mouse sperm surfAce AS-A on ZP binding wAs demonstrAted by dose-dependent decreAses of sperm-ZP binding on sperm pretreAtment with Anti-AS-A IgG/FAb. Furthermore, AlexA-430 conjugAted AS-A bound to mouse ZP of unfertilized eggs but not to fertilized ones, And this level of binding increAsed And ApproAched sAturAtion with increAsing AlexA430 AS-A concentrAtions. Moreover, in vivo fertilizAtion wAs mArkedly decreAsed when mouse sperm pretreAted with AntiAS-A IgG were ArtificiAlly inseminAted into femAles. All of these results designAted A new function for AS-A in mouse gAmete interAction. fertilizAtion, gAmete biology, sperm

Ulrich Matzner - One of the best experts on this subject based on the ideXlab platform.

  • brAin cell type specific endocytosis of ArylsulfAtAse A identifies limitAtions of enzyme bAsed therApies for metAchromAtic leukodystrophy
    Human Molecular Genetics, 2021
    Co-Authors: Debora Kaminski, Frank Matthes, Volkmar Gieselmann, Claudia Yaghootfam, Annika Resing, Ulrich Matzner
    Abstract:

    Enzyme replAcement therApies, Allogeneic bone mArrow trAnsplAntAtion And gene therApies Are treAtment options for lysosomAl storAge diseAses cAused by inherited deficiencies of soluble lysosomAl enzymes. Independent from the ApproAch, the enzyme must be delivered to lysosomes of deficient pAtient cells. Little is known About the disseminAtion of enzyme within A tissue where cells compete for uptAke viA different receptor systems, binding Affinities And endocytic rAtes. To evAluAte disseminAtion And lysosomAl tArgeting of A lysosomAl enzyme in the CNS, we AnAlysed receptor-mediAted endocytosis of ArylsulfAtAse A (ASA) by different types of brAin-derived cell lines And primAry murine brAin cells. For ASA expressed by chinese hAmster ovAry cells for enzyme replAcement therApy of metAchromAtic leukodystrophy, endocytic rAtes decline from microgliA to neurons And Astrocytes And to oligodendrocytes. Only immAture oligodendrocytes endocytose significAnt Amounts of enzyme. UptAke by non-microgliAl cells is due to mAnnose 6-phosphAte receptors, whereAs severAl receptor systems pArticipAte in endocytosis by microgliAl cells. Interestingly, ASA expressed by microgliAl cells cAnnot be tAken up in A mAnnose 6-phosphAte dependent mAnner. The resulting fAilure to correct non-microgliAl cells corroborAtes in vivo dAtA And indicAtes thAt therApeutic effects of Allogeneic bone mArrow trAnsplAntAtion And hemAtopoietic stem cell gene therApy on metAchromAtic leukodystrophy Are independent of metAbolic cross-correction of neurons, Astrocytes And oligodendrocytes by receptor-mediAted endocytosis.

  • evolutionAry redesign of the lysosomAl enzyme ArylsulfAtAse A increAses efficAcy of enzyme replAcement therApy for metAchromAtic leukodystrophy
    Human Molecular Genetics, 2019
    Co-Authors: Heidi Simonis, Claudia Yaghootfam, Volkmar Gieselmann, Marc Sylvester, Ulrich Matzner
    Abstract:

    Protein engineering is A meAns to optimize protein therApeutics developed for the treAtment of so fAr incurAble diseAses including cAncers And genetic disorders. Here we report on An engineering ApproAch in which we successfully increAsed the cAtAlytic rAte constAnt of An enzyme thAt is presently evAluAted in enzyme replAcement therApies (ERT) of A lysosomAl storAge diseAse (LSD). Although ERT is A treAtment option for mAny LSDs, outcomes Are lAgging fAr behind expectAtions for most of them. This hAs been Ascribed to insufficient enzyme Activities AccumulAting in tissues difficult to tArget such As brAin And peripherAl nerves. We show for humAn ArylsulfAtAse A (hARSA) thAt the Activity of A therApeutic enzyme cAn be substAntiAlly increAsed by reversing Activity-diminishing And by inserting Activity-promoting Amino Acid substitutions thAt hAd occurred in the evolution of hominids And non-humAn mAmmAls, respectively. The potentiAl of this ApproAch, here designAted As evolutionAry redesign, wAs highlighted by the observAtion thAt murinizAtion of only 1 or 3 Amino Acid positions increAsed the hARSA Activity 3- And 5-fold, with little impAct on stAbility, respectively. The two kineticAlly optimized hARSA vAriAnts showed no immunogenic potentiAl in ERT of A humAnized ARSA knockout mouse model of metAchromAtic leukodystrophy (MLD) And reduced lysosomAl storAge of kidney, peripherAl And centrAl nervous system up to 3-fold more efficiently thAn wild-type hARSA. Due to their sAfety profile And higher therApeutic potentiAl the engineered hARSA vAriAnts might represent mAjor AdvAnces for future enzyme-bAsed therApies of MLD And stimulAte AnAlogous ApproAches for other enzyme therApeutics.

  • potentiAl of surfActAnt coAted nAnopArticles to improve brAin delivery of ArylsulfAtAse A
    Journal of Controlled Release, 2017
    Co-Authors: Tilman Schuster, Volkmar Gieselmann, Astrid Muhlstein, Claudia Yaghootfam, Olga Maksimenko, E V Shipulo, Svetlana Gelperina, Jorg Kreuter, Ulrich Matzner
    Abstract:

    AbstrAct The lysosomAl storAge disorder (LSD) metAchromAtic leukodystrophy (MLD) is cAused by A deficiency of the soluble, lysosomAl hydrolAse ArylsulfAtAse A (ASA). The diseAse is chArActerized by AccumulAtion of 3-O-sulfogAlActosylcerAmide (sulfAtide), progressive demyelinAtion of the nervous system And premAture deAth. Enzyme replAcement therApy (ERT), bAsed on regulAr intrAvenous injections of recombinAnt functionAl enzyme, is in clinicAl use for severAl LSDs. For MLD And other LSDs with centrAl nervous system (CNS) involvement, however, ERT is limited by the blood-brAin bArrier (BBB) restricting trAnsport of therApeutic enzymes from the blood to the brAin. In the present study, the potentiAl of different types of surfActAnt-coAted biodegrAdAble nAnopArticles to increAse brAin delivery of ASA wAs evAluAted. Three different strAtegies to bind ASA to nAnopArticle surfAces were compAred: (1) Adsorption, (2) high-Affinity binding viA the streptAvidin-biotin system, And (3) covAlent binding. Adsorption Allowed binding of high Amounts of Active ASA. However, in presence of phosphAte-buffered sAline or serum rApid And complete desorption occurred, rendering this strAtegy ineffective for in vivo ApplicAtions. In contrAst, stAble immobilizAtion with negligible dissociAtion wAs Achieved by high-Affinity And covAlent binding. Consequently, we AnAlyzed the brAin tArgeting of two stAbly nAnopArticle-bound ASA formulAtions in ASA−/− mice, An AnimAl model of MLD. CompAred to free ASA, injected As A control, the biodistribution of nAnopArticle-bound ASA wAs Altered in peripherAl orgAns, but no increAse of brAin levels wAs detectAble. The fAilure to improve brAin delivery suggests thAt the ASA glycoprotein interferes with processes required to tArget surfActAnt-coAted nAnopArticles to brAin cApillAry endotheliAl cells.

  • ArylsulfAtAse A overexpressing humAn ipsc derived neurAl cells reduce cns sulfAtide storAge in A mouse model of metAchromAtic leukodystrophy
    Molecular Therapy, 2015
    Co-Authors: Jonas Doerr, Ulrich Matzner, Volkmar Gieselmann, Annika Bockenhoff, Benjamin Ewald, Julia Ladewig, Matthias Eckhardt, Oliver Brustle, Philipp Koch
    Abstract:

    MetAchromAtic leukodystrophy (MLD) is An inherited lysosomAl storAge disorder resulting from A functionAl deficiency of ArylsulfAtAse A (ARSA), An enzyme thAt cAtAlyzes desulfAtion of 3-O-sulfogAlActosylcerAmide (sulfAtide). LAck of Active ARSA leAds to the AccumulAtion of sulfAtide in oligodendrocytes, SchwAnn cells And some neurons And triggers progressive demyelinAtion, the neuropAthologicAl hAllmArk of MLD. SeverAl therApeutic ApproAches hAve been explored, including enzyme replAcement, Autologous hemAtopoietic stem cell-bAsed gene therApy, intrAcerebrAl gene therApy or cell-bAsed gene delivery into the centrAl nervous system (CNS). However, long-term treAtment of the blood-brAin-bArrier protected CNS remAins chAllenging. Here we used MLD pAtient-derived induced pluripotent stem cells (iPSCs) to generAte long-term self-renewing neuroepitheliAl stem cells And AstrogliAl progenitors for cell-bAsed ARSA replAcement. Following trAnsplAntAtion of ARSA-overexpressing precursors into ARSA-deficient mice we observed A significAnt reduction of sulfAtide storAge up to A distAnce of 300 µm from grAfted cells. Our dAtA indicAte thAt neurAl precursors generAted viA reprogrAmming from MLD pAtients cAn be engineered to AmeliorAte sulfAtide AccumulAtion And mAy thus serve As Autologous cell-bAsed vehicle for continuous ARSA supply in MLD-Affected brAin tissue.

  • compArison of five peptide vectors for improved brAin delivery of the lysosomAl enzyme ArylsulfAtAse A
    The Journal of Neuroscience, 2014
    Co-Authors: Annika Bockenhoff, Volkmar Gieselmann, Philipp Wolte, Hansjoachim Galla, Sandra Cramer, Simeon Knieling, Claudia Wohlenberg, Ulrich Matzner
    Abstract:

    Enzyme replAcement therApy (ERT) is A treAtment option for lysosomAl storAge disorders (LSDs) cAused by deficiencies of soluble lysosomAl enzymes. ERT depends on receptor-mediAted trAnsport of intrAvenously injected recombinAnt enzyme to lysosomes of pAtient cells. The blood–brAin bArrier (BBB) prevents efficient trAnsfer of therApeutic polypeptides from the blood to the brAin pArenchymA And thus hinders effective treAtment of LSDs with CNS involvement. We compAred the potentiAl of five brAin-tArgeting peptides to promote brAin delivery of the lysosomAl enzyme ArylsulfAtAse A (ASA). Fusion proteins between ASA And the protein trAnsduction domAin of the humAn immunodeficiency virus TAT protein (TAt), An Angiopep peptide (Ang-2), And the receptor-binding domAins of humAn Apolipoprotein B (ApoB) And ApoE (two versions, ApoE-I And ApoE-II) were generAted. All ASA fusion proteins were enzymAticAlly Active And tArgeted to lysosomes when Added to cultured cells. In contrAst to wild-type ASA, which is tAken up by mAnnose-6-phosphAte receptors, All chimeric proteins were AdditionAlly endocytosed viA mAnnose-6-phosphAte-independent routes. For ASA-Ang-2, ASA-ApoE-I, And ASA-ApoE-II, uptAke wAs pArtiAlly due to the low-density lipoprotein receptor-relAted protein 1. TrAnsendotheliAl trAnsfer in A BBB cell culture model wAs elevAted for ASA-ApoB, ASA-ApoE-I, And ASA-ApoE-II. BrAin delivery wAs, however, increAsed only for ASA-ApoE-II. ApoE-II wAs Also superior to wild-type ASA in reducing lysosomAl storAge in the CNS of ASA-knock-out mice treAted by ERT. Therefore, the ApoE-derived peptide AppeArs useful to treAt metAchromAtic leukodystrophy And possibly other neurologicAl disorders more efficiently.

Euridice Carmona - One of the best experts on this subject based on the ideXlab platform.

  • interAction of ArylsulfAtAse A AsA with its nAturAl sulfoglycolipid substrAtes A computAtionAl And site directed mutAgenesis study
    Glycoconjugate Journal, 2009
    Co-Authors: Matthias Schenk, Daniela Costa Santos, Nongnuj Tanphaichitr, Euridice Carmona, Chaitanya A K Koppisetty, Smita Bhatia, Pergeorg Nyholm
    Abstract:

    ArylsulfAtAse A (ASA) hydrolyzes sulfAte esters with A pH optimum of 5. InterActions between p-nitrocAtechol sulfAte (NCS, ArtificiAl substrAte) And Active site residues of ASA Are reveAled from their co-crystAl structure. Since equivAlent ASA interActions with its nAturAl substrAtes, sulfogAlActosylcerAmide (SGC) And sulfogAlActosylglycerolipid (SGG), Are not known, we computAtionAlly docked SGC/SGG to the ASA crystAl structure. Our dockings suggested thAt Cys69 wAs the Active site residue, And Lys302 & Lys123 As residues Anchoring the sulfAte group of SGC/SGG to the Active site, As observed for NCS. We further confirmed these results using 2 recombinAnt ASA mutAnts: C69A And CKK (Cys69, Lys302 And Lys123-All mutAted to AlA). Both ASA mutAnts fAiled to desulfAte SGC/SGG, And CKK showed minimAl binding to [14C]SGC, Although C69A still hAd Affinity for this sulfoglycolipid. However, our dockings suggested AdditionAl intermoleculAr hydrogen bonding And hydrophobic interActions between ASA And SGC/SGG, thus contributing to the specificity of SGC/SGG As nAturAl substrAtes.

  • sperm surfAce ArylsulfAtAse A cAn disperse the cumulus mAtrix of cumulus oocyte complexes
    Journal of Cellular Physiology, 2007
    Co-Authors: Araya Anupriwan, Daniela Costa Santos, Tony Rupar, Euridice Carmona, Sitthichai Iamsaard, Krittalak Chakrabandhu, Benjamin K Tsang, Nongnuj Tanphaichitr
    Abstract:

    Cumulus cell lAyers of expAnded cumulus oocyte complexes (COCs) Are interlinked with networks of hyAluronic Acid, chondroitin sulfAte B proteoglycAns And link proteins, And they cAn be dispersed by sperm surfAce hyAluronidAses. In this report, we showed thAt ArylsulfAtAse A (AS-A), existing on the sperm heAd surfAce, Also hAd this dispersion Action. Purified AS-A free of proteAse, hyAluronidAse And chondroitinAse Activities could disperse the cumulus mAtrix of expAnded COCs. However, this COC dispersion Action wAs not AssociAted with AS-A desulfAtion Activity, AssAyed by using p-nitrocAtecholsulfAte (ArtificiAl substrAte). COCs incubAted for 1 h with sperm pretreAted with Anti-AS-A IgG in the presence of Apigenin (A hyAluronidAse inhibitor) did not exhibit mAtrix dispersion, whereAs severAl cumulus lAyers were AlreAdy dispersed in COCs incubAted with sperm pretreAted with preimmune IgG. Furthermore, sperm from AS-A null mice showed A significAnt delAy in COC dispersion, compAred with wild-type sperm. Within 1 h of sperm-COC co-incubAtion, the size of COCs incubAted with AS-A null sperm wAs 65% of the originAl dimension, whereAs thAt of COCs inseminAted with wild-type sperm wAs only 17%. A further delAy in COC dispersion by AS-A(-/-) mouse sperm wAs observed when Apigenin wAs present in the co-incubAtion. We Also showed for the first time thAt AS-A hAd A specific Affinity for chondroitin sulfAte B, A component of cumulus mAtrix proteoglycAn networks; this might provide A mechAnism of cumulus mAtrix destAbilizAtion induced by sperm surfAce AS-A.

  • Acquisition of ArylsulfAtAse A onto the mouse sperm surfAce during epididymAl trAnsit
    Biology of Reproduction, 2003
    Co-Authors: Wattana Weerachatyanukul, Araya Anupriwan, Louis Hermo, Euridice Carmona, Michael G Wade, Solange Maria Da Silva, Peter Rippstein, Prasert Sobhon, Prapee Sretarugsa, Nongnuj Tanphaichitr
    Abstract:

    ArylsulfAtAse A (AS-A) is locAlized to the sperm surfAce And pArticipAtes in sperm-zonA pellucidA binding. We investigAted how AS-A, usuAlly known As An AcrosomAl enzyme, trAfficked to the sperm surfAce. Immunocytochemistry of the mouse testis confirmed the existence of AS-A in the AcrosomAl region of round And elongAting spermAtids. However, immunofluorescence And flow cytometry indicAted the Absence of AS-A on the surfAce of live testiculAr sperm. In contrAst, positive AS-A stAining wAs observed in the heAds of live cAudAl epididymAl And vAs deferens sperm. The results suggested thAt Acquisition of AS-A on the sperm surfAce occurred during epididymAl trAnsit. Immunocytochemistry of the epididymis reveAled AS-A in nArrow And ApicAl cells in the initiAl segment And in cleAr cells in All epididymAl regions. However, these epitheliAl cells Are in the minority And Are not involved in secretory Activity. In the cAudAl epididymis And vAs deferens, AS-A wAs Also locAlized to principAl cells, the mAjor epitheliAl cells. BecAuse principAl cells hAve secretory Activity, they mAy secrete AS-A into the epididymAl fluid. This hypothesis wAs supported by our results reveAling the presence of AS-A in the epididymAl And vAs deferens fluid (determined by immunoblotting And ELISA) And An AS-A trAnscript in the epididymis (by reverse trAnscription polymerAse chAin reAction). AlexA-430 AS-A bound to epididymAl sperm with high Affinity (Kd = 46 nM). This binding wAs inhibited by treAtment of sperm with An Antibody AgAinst sperm surfAce sulfogAlActosylglycerolipid. This finding suggests thAt AS-A in the epididymAl fluid mAy deposit onto sperm viA its Affinity to sulfogAlActosylglycerolipid.

  • role of sperm surfAce ArylsulfAtAse A in mouse sperm zonA pellucidA binding
    Biology of Reproduction, 2002
    Co-Authors: Julierut Tantibhedhyangkul, Araya Anupriwan, Wattana Weerachatyanukul, Euridice Carmona, Hongbin Xu, Dominick Michaud, Nongnuj Tanphaichitr
    Abstract:

    We hAve previously described the zonAe pellucidAe (ZP) binding Ability of A pig sperm surfAce protein, P68. Our recent results on peptide sequencing of 3 P68 tryptic peptides And moleculAr cloning of pig testis ArylsulfAtAse A (AS-A) reveAled the identity of P68 As AS-A. In this report, we demonstrAte the presence of AS-A on the mouse sperm surfAce And its role in ZP binding. Using Anti-AS-A Antibody, we hAve shown by immunoblotting thAt AS-A wAs present in A Triton X-100 extrAct of mouse sperm. The presence of AS-A on the sperm plAsmA membrAne wAs conclusively demonstrAted by indirect immunofluorescence, immunogold electron microscopy, And AS-A’s desulfAtion Activity on live mouse sperm. The AS-A remAined on the heAd surfAce of in vivo cApAcitAted sperm, As reveAled by positive immunofluorescent stAining of oviductAl/uterine sperm. SignificAntly, the role of mouse sperm surfAce AS-A on ZP binding wAs demonstrAted by dose-dependent decreAses of sperm-ZP binding on sperm pretreAtment with Anti-AS-A IgG/FAb. Furthermore, AlexA-430 conjugAted AS-A bound to mouse ZP of unfertilized eggs but not to fertilized ones, And this level of binding increAsed And ApproAched sAturAtion with increAsing AlexA430 AS-A concentrAtions. Moreover, in vivo fertilizAtion wAs mArkedly decreAsed when mouse sperm pretreAted with AntiAS-A IgG were ArtificiAlly inseminAted into femAles. All of these results designAted A new function for AS-A in mouse gAmete interAction. fertilizAtion, gAmete biology, sperm

  • role of sperm surfAce ArylsulfAtAse A in mouse sperm zonA pellucidA binding
    Biology of Reproduction, 2002
    Co-Authors: Julierut Tantibhedhyangkul, Araya Anupriwan, Wattana Weerachatyanukul, Euridice Carmona, Dominick Michaud, Nongnuj Tanphaichitr
    Abstract:

    We hAve previously described the zonAe pellucidAe (ZP) binding Ability of A pig sperm surfAce protein, P68. Our recent results on peptide sequencing of 3 P68 tryptic peptides And moleculAr cloning of pig testis ArylsulfAtAse A (AS-A) reveAled the identity of P68 As AS-A. In this report, we demonstrAte the presence of AS-A on the mouse sperm surfAce And its role in ZP binding. Using Anti-AS-A Antibody, we hAve shown by immunoblotting thAt AS-A wAs present in A Triton X-100 extrAct of mouse sperm. The presence of AS-A on the sperm plAsmA membrAne wAs conclusively demonstrAted by indirect immunofluorescence, immunogold electron microscopy, And AS-A's desulfAtion Activity on live mouse sperm. The AS-A remAined on the heAd surfAce of in vivo cApAcitAted sperm, As reveAled by positive immunofluorescent stAining of oviductAl/uterine sperm. SignificAntly, the role of mouse sperm surfAce AS-A on ZP binding wAs demonstrAted by dose-dependent decreAses of sperm-ZP binding on sperm pretreAtment with Anti-AS-A IgG/FAb. Furthermore, AlexA-430 conjugAted AS-A bound to mouse ZP of unfertilized eggs but not to fertilized ones, And this level of binding increAsed And ApproAched sAturAtion with increAsing AlexA-430 AS-A concentrAtions. Moreover, in vivo fertilizAtion wAs mArkedly decreAsed when mouse sperm pretreAted with Anti-AS-A IgG were ArtificiAlly inseminAted into femAles. All of these results designAted A new function for AS-A in mouse gAmete interAction.

Araya Anupriwan - One of the best experts on this subject based on the ideXlab platform.

  • presence of ArylsulfAtAse A And sulfogAlActosylglycerolipid in mouse ovAries locAlizAtion to the corpus luteum
    Endocrinology, 2008
    Co-Authors: Araya Anupriwan, Matthias Schenk, Rapeepun Vanichviriyakit, Daniela Costa Santos, Arman Yaghoubian, Alexander T H Wu, Kessiri Kongmanas, Trish Berger, Fang Liu, Kym F Faull
    Abstract:

    ArylsulfAtAse A (AS-A) is A lysosomAl enzyme, which cAtAlyzes the desulfAtion of certAin sulfogAlActolipids, including sulfogAlActosylglycerolipid (SGG), A molecule implicAted in cell Adhesion. In this report, immunocytochemistry reveAled the selective presence of AS-A in the corpus luteum of mouse ovAries. Immunoblotting indicAted thAt mouse corpus luteum AS-A hAd A moleculAr mAss of 66 kDA, similAr to AS-A of other tissues. Corpus luteum AS-A wAs Active, cApAble of desulfAting the ArtificiAl substrAte, p-nitrocAtechol sulfAte, At the optimum pH of five. To understAnd further the role of AS-A in femAle reproduction, levels of AS-A were determined during corpus luteum development in pseudopregnAnt mice And during luteolysis After cessAtion of pseudopregnAncy. Immunocytochemistry, immunoblotting And desulfAtion Activity showed thAt AS-A expression wAs evident At the onset of pseudopregnAncy in the newly formed corporA luteA, And its level increAsed steAdily during glAnd development. The increAse in the expression And Activity of AS-A continued throughout luteolysis After the decreAse in serum progesterone levels. We Also observed the selective presence of SGG on the luteAl cell surfAce in developed corporA luteA, As shown by immunofluorescence of mouse ovAry sections As well As high-performAnce thin-lAyer chromAtogrAphy of lipids isolAted from mouse And pig corporA luteA. The identity of the “SGG” bAnd on the thin lAyer silicA plAte wAs further vAlidAted by electrosprAy ionizAtion mAss spectrometry. SignificAntly, SGG disAppeAred in regressing corporA luteA. Therefore, lysosomAl AS-A mAy be involved in cell-surfAce remodeling during luteolysis by desulfAting SGG After its endocytosis And tArgeting to the lysosome.

  • sperm surfAce ArylsulfAtAse A cAn disperse the cumulus mAtrix of cumulus oocyte complexes
    Journal of Cellular Physiology, 2007
    Co-Authors: Araya Anupriwan, Daniela Costa Santos, Tony Rupar, Euridice Carmona, Sitthichai Iamsaard, Krittalak Chakrabandhu, Benjamin K Tsang, Nongnuj Tanphaichitr
    Abstract:

    Cumulus cell lAyers of expAnded cumulus oocyte complexes (COCs) Are interlinked with networks of hyAluronic Acid, chondroitin sulfAte B proteoglycAns And link proteins, And they cAn be dispersed by sperm surfAce hyAluronidAses. In this report, we showed thAt ArylsulfAtAse A (AS-A), existing on the sperm heAd surfAce, Also hAd this dispersion Action. Purified AS-A free of proteAse, hyAluronidAse And chondroitinAse Activities could disperse the cumulus mAtrix of expAnded COCs. However, this COC dispersion Action wAs not AssociAted with AS-A desulfAtion Activity, AssAyed by using p-nitrocAtecholsulfAte (ArtificiAl substrAte). COCs incubAted for 1 h with sperm pretreAted with Anti-AS-A IgG in the presence of Apigenin (A hyAluronidAse inhibitor) did not exhibit mAtrix dispersion, whereAs severAl cumulus lAyers were AlreAdy dispersed in COCs incubAted with sperm pretreAted with preimmune IgG. Furthermore, sperm from AS-A null mice showed A significAnt delAy in COC dispersion, compAred with wild-type sperm. Within 1 h of sperm-COC co-incubAtion, the size of COCs incubAted with AS-A null sperm wAs 65% of the originAl dimension, whereAs thAt of COCs inseminAted with wild-type sperm wAs only 17%. A further delAy in COC dispersion by AS-A(-/-) mouse sperm wAs observed when Apigenin wAs present in the co-incubAtion. We Also showed for the first time thAt AS-A hAd A specific Affinity for chondroitin sulfAte B, A component of cumulus mAtrix proteoglycAn networks; this might provide A mechAnism of cumulus mAtrix destAbilizAtion induced by sperm surfAce AS-A.

  • Acquisition of ArylsulfAtAse A onto the mouse sperm surfAce during epididymAl trAnsit
    Biology of Reproduction, 2003
    Co-Authors: Wattana Weerachatyanukul, Araya Anupriwan, Louis Hermo, Euridice Carmona, Michael G Wade, Solange Maria Da Silva, Peter Rippstein, Prasert Sobhon, Prapee Sretarugsa, Nongnuj Tanphaichitr
    Abstract:

    ArylsulfAtAse A (AS-A) is locAlized to the sperm surfAce And pArticipAtes in sperm-zonA pellucidA binding. We investigAted how AS-A, usuAlly known As An AcrosomAl enzyme, trAfficked to the sperm surfAce. Immunocytochemistry of the mouse testis confirmed the existence of AS-A in the AcrosomAl region of round And elongAting spermAtids. However, immunofluorescence And flow cytometry indicAted the Absence of AS-A on the surfAce of live testiculAr sperm. In contrAst, positive AS-A stAining wAs observed in the heAds of live cAudAl epididymAl And vAs deferens sperm. The results suggested thAt Acquisition of AS-A on the sperm surfAce occurred during epididymAl trAnsit. Immunocytochemistry of the epididymis reveAled AS-A in nArrow And ApicAl cells in the initiAl segment And in cleAr cells in All epididymAl regions. However, these epitheliAl cells Are in the minority And Are not involved in secretory Activity. In the cAudAl epididymis And vAs deferens, AS-A wAs Also locAlized to principAl cells, the mAjor epitheliAl cells. BecAuse principAl cells hAve secretory Activity, they mAy secrete AS-A into the epididymAl fluid. This hypothesis wAs supported by our results reveAling the presence of AS-A in the epididymAl And vAs deferens fluid (determined by immunoblotting And ELISA) And An AS-A trAnscript in the epididymis (by reverse trAnscription polymerAse chAin reAction). AlexA-430 AS-A bound to epididymAl sperm with high Affinity (Kd = 46 nM). This binding wAs inhibited by treAtment of sperm with An Antibody AgAinst sperm surfAce sulfogAlActosylglycerolipid. This finding suggests thAt AS-A in the epididymAl fluid mAy deposit onto sperm viA its Affinity to sulfogAlActosylglycerolipid.

  • role of sperm surfAce ArylsulfAtAse A in mouse sperm zonA pellucidA binding
    Biology of Reproduction, 2002
    Co-Authors: Julierut Tantibhedhyangkul, Araya Anupriwan, Wattana Weerachatyanukul, Euridice Carmona, Hongbin Xu, Dominick Michaud, Nongnuj Tanphaichitr
    Abstract:

    We hAve previously described the zonAe pellucidAe (ZP) binding Ability of A pig sperm surfAce protein, P68. Our recent results on peptide sequencing of 3 P68 tryptic peptides And moleculAr cloning of pig testis ArylsulfAtAse A (AS-A) reveAled the identity of P68 As AS-A. In this report, we demonstrAte the presence of AS-A on the mouse sperm surfAce And its role in ZP binding. Using Anti-AS-A Antibody, we hAve shown by immunoblotting thAt AS-A wAs present in A Triton X-100 extrAct of mouse sperm. The presence of AS-A on the sperm plAsmA membrAne wAs conclusively demonstrAted by indirect immunofluorescence, immunogold electron microscopy, And AS-A’s desulfAtion Activity on live mouse sperm. The AS-A remAined on the heAd surfAce of in vivo cApAcitAted sperm, As reveAled by positive immunofluorescent stAining of oviductAl/uterine sperm. SignificAntly, the role of mouse sperm surfAce AS-A on ZP binding wAs demonstrAted by dose-dependent decreAses of sperm-ZP binding on sperm pretreAtment with Anti-AS-A IgG/FAb. Furthermore, AlexA-430 conjugAted AS-A bound to mouse ZP of unfertilized eggs but not to fertilized ones, And this level of binding increAsed And ApproAched sAturAtion with increAsing AlexA430 AS-A concentrAtions. Moreover, in vivo fertilizAtion wAs mArkedly decreAsed when mouse sperm pretreAted with AntiAS-A IgG were ArtificiAlly inseminAted into femAles. All of these results designAted A new function for AS-A in mouse gAmete interAction. fertilizAtion, gAmete biology, sperm

  • role of sperm surfAce ArylsulfAtAse A in mouse sperm zonA pellucidA binding
    Biology of Reproduction, 2002
    Co-Authors: Julierut Tantibhedhyangkul, Araya Anupriwan, Wattana Weerachatyanukul, Euridice Carmona, Dominick Michaud, Nongnuj Tanphaichitr
    Abstract:

    We hAve previously described the zonAe pellucidAe (ZP) binding Ability of A pig sperm surfAce protein, P68. Our recent results on peptide sequencing of 3 P68 tryptic peptides And moleculAr cloning of pig testis ArylsulfAtAse A (AS-A) reveAled the identity of P68 As AS-A. In this report, we demonstrAte the presence of AS-A on the mouse sperm surfAce And its role in ZP binding. Using Anti-AS-A Antibody, we hAve shown by immunoblotting thAt AS-A wAs present in A Triton X-100 extrAct of mouse sperm. The presence of AS-A on the sperm plAsmA membrAne wAs conclusively demonstrAted by indirect immunofluorescence, immunogold electron microscopy, And AS-A's desulfAtion Activity on live mouse sperm. The AS-A remAined on the heAd surfAce of in vivo cApAcitAted sperm, As reveAled by positive immunofluorescent stAining of oviductAl/uterine sperm. SignificAntly, the role of mouse sperm surfAce AS-A on ZP binding wAs demonstrAted by dose-dependent decreAses of sperm-ZP binding on sperm pretreAtment with Anti-AS-A IgG/FAb. Furthermore, AlexA-430 conjugAted AS-A bound to mouse ZP of unfertilized eggs but not to fertilized ones, And this level of binding increAsed And ApproAched sAturAtion with increAsing AlexA-430 AS-A concentrAtions. Moreover, in vivo fertilizAtion wAs mArkedly decreAsed when mouse sperm pretreAted with Anti-AS-A IgG were ArtificiAlly inseminAted into femAles. All of these results designAted A new function for AS-A in mouse gAmete interAction.

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