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Christina I. Schroeder - One of the best experts on this subject based on the ideXlab platform.

  • Efficient enzymAtic ligAtion of inhibitor cystine knot spider venom peptides: using SortAse A to form double-knottins thAt probe voltAge-gAted sodium chAnnel NAV1.7
    Bioconjugate chemistry, 2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provid...

  • Efficient EnzymAtic LigAtion of Inhibitor Cystine Knot Spider Venom Peptides: Using SortAse A To Form Double-Knottins ThAt Probe VoltAge-GAted Sodium ChAnnel NAV1.7
    2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provide novel tools for enhAncing our understAnding of, And design strAtegies for, therApeuticAlly relevAnt voltAge-gAted ion chAnnels

  • semienzymAtic cyclizAtion of disulfide rich peptides using SortAse A
    Journal of Biological Chemistry, 2014
    Co-Authors: Xinying Jia, Soohyun Kwon, Chingi Anderson Wang, Yenhua Huang, Lai Yue Chan, Chia Chia Tan, Johan K Rosengren, Jason Mulvenna, Christina I. Schroeder
    Abstract:

    Disulfide-rich cyclic peptides hAve generAted greAt interest in the development of peptide-bAsed therApeutics due to their exceptionAl stAbility towArd chemicAl, enzymAtic, or thermAl AttAck. In pArticulAr, they hAve been used As scAffolds onto which bioActive epitopes cAn be grAfted to tAke AdvAntAge of the fAvorAble biophysicAl properties of disulfide-rich cyclic peptides. To dAte, the most commonly used method for the heAd-to-tAil cyclizAtion of peptides hAs been nAtive chemicAl ligAtion. In recent yeArs, however, enzyme-mediAted cyclizAtion hAs become A promising new technology due to its efficiency, sAfety, And cost-effectiveness. SortAse A (SrtA) is A bActeriAl enzyme with trAnspeptidAse Activity. It recognizes A C-terminAl pentA-Amino Acid motif, LPXTG, And cleAves the Amide bond between Thr And Gly to form A thioAcyl-linked intermediAte. This intermediAte undergoes nucleophilic AttAck by An N-terminAl poly-Gly sequence to form An Amide bond between the Thr And N-terminAl Gly. Here, we demonstrAte thAt SortAse A cAn successfully be used to cyclize A vAriety of smAll disulfide-rich peptides, including the cyclotide kAlAtA B1, α-conotoxin Vc1.1, And sunflower trypsin inhibitor 1. These peptides rAnge in size from 14 to 29 Amino Acids And contAin three, two, or one disulfide bond, respectively, within their heAd-to-tAil cyclic bAckbones. Our findings provide proof of concept for the potentiAl broAd ApplicAbility of enzymAtic cyclizAtion of disulfide-rich peptides with therApeutic potentiAl.

Akello J. Agwa - One of the best experts on this subject based on the ideXlab platform.

  • Efficient enzymAtic ligAtion of inhibitor cystine knot spider venom peptides: using SortAse A to form double-knottins thAt probe voltAge-gAted sodium chAnnel NAV1.7
    Bioconjugate chemistry, 2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provid...

  • Efficient EnzymAtic LigAtion of Inhibitor Cystine Knot Spider Venom Peptides: Using SortAse A To Form Double-Knottins ThAt Probe VoltAge-GAted Sodium ChAnnel NAV1.7
    2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provide novel tools for enhAncing our understAnding of, And design strAtegies for, therApeuticAlly relevAnt voltAge-gAted ion chAnnels

David J. Craik - One of the best experts on this subject based on the ideXlab platform.

  • Efficient enzymAtic ligAtion of inhibitor cystine knot spider venom peptides: using SortAse A to form double-knottins thAt probe voltAge-gAted sodium chAnnel NAV1.7
    Bioconjugate chemistry, 2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provid...

  • Efficient EnzymAtic LigAtion of Inhibitor Cystine Knot Spider Venom Peptides: Using SortAse A To Form Double-Knottins ThAt Probe VoltAge-GAted Sodium ChAnnel NAV1.7
    2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provide novel tools for enhAncing our understAnding of, And design strAtegies for, therApeuticAlly relevAnt voltAge-gAted ion chAnnels

Linda V. Blomster - One of the best experts on this subject based on the ideXlab platform.

  • Efficient enzymAtic ligAtion of inhibitor cystine knot spider venom peptides: using SortAse A to form double-knottins thAt probe voltAge-gAted sodium chAnnel NAV1.7
    Bioconjugate chemistry, 2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provid...

  • Efficient EnzymAtic LigAtion of Inhibitor Cystine Knot Spider Venom Peptides: Using SortAse A To Form Double-Knottins ThAt Probe VoltAge-GAted Sodium ChAnnel NAV1.7
    2018
    Co-Authors: Akello J. Agwa, Linda V. Blomster, David J. Craik, Christina I. Schroeder
    Abstract:

    GAting modifier toxins from spider venom Are disulfide-rich peptides thAt typicAlly comprise A stAbilizing inhibitor cystine knot (ICK). These knottin peptides Are being pursued As therApeutic leAds for A rAnge of conditions linked to trAnsmembrAne proteins. Recently, double-knottin peptides discovered in spider venom And produced by recombinAnt expression hAve provided insights into the phArmAcology of trAnsmembrAne chAnnels. Here, we use chemoenzymAtic ligAtion to produce double-knottins to probe the effect of bivAlent modulAtion on the voltAge-gAted sodium chAnnel subtype 1.7 (NAV1.7), which is implicAted in pAin signAling. MonovAlent knottins were oxidAtively folded And then biochemicAlly conjugAted using SortAse A, to form double-knottins. The structurAl integrity of the peptides wAs confirmed using NMR, And fluorescence-bAsed Activity AssAys provided evidence suggesting thAt coincubAted monovAlent And bivAlent knottins cAn cooperAtively modulAte NAV1.7. We AnticipAte thAt double-knottins will provide novel tools for enhAncing our understAnding of, And design strAtegies for, therApeuticAlly relevAnt voltAge-gAted ion chAnnels

Ykelien L. Boersma - One of the best experts on this subject based on the ideXlab platform.

  • Engineering the specificity of Streptococcus pyogenes SortAse A by loop grAfting
    Proteins, 2020
    Co-Authors: Magdalena Wójcik, Wim J Quax, Kamil Szala, Ronald Van Merkerk, Ykelien L. Boersma
    Abstract:

    SortAses Are A group of enzymes displAyed on the cell-wAll of GrAm-positive bActeriA. They Are responsible for the AttAchment of virulence fActors onto the peptidoglycAn in A trAnspeptidAtion reAction through recognition of A pentApeptide substrAte. Most housekeeping SortAses recognize one specific pentApeptide motif; however, Streptococcus pyogenes SortAse A (SpSrtA WT) recognizes LPETG, LPETA And LPKLG motifs. Here, we exAmined SpSrtA's flexible substrAte specificity by investigAting the role of the β7/β8 loop in determining substrAte specificity. We exchAnged the β7/β8 loop in SpSrtA with corresponding β7/β8 loops from StAphylococcus Aureus (SASrtA WT) And BAcillus AnthrAcis (BASrtA WT). While the BASrtA-derived vAriAnt showed no enzymAtic Activity towArd either LPETG or LPETA substrAtes, the Activity of the SASrtA-derived mutAnt towArd the LPETA substrAte wAs completely Abolished. InsteAd, the mutAnt hAd An improved Activity towArd LPETG, the preferred substrAte of SASrtA WT.

  • IdentificAtion of potentiAl Antivirulence Agents by substitution-oriented screening for inhibitors of Streptococcus pyogenes SortAse A
    European Journal of Medicinal Chemistry, 2018
    Co-Authors: Magdalena Wójcik, Nikolaos Eleftheriadis, Alexander Dömling, Martijn R H Zwinderman, Frank J Dekker, Ykelien L. Boersma
    Abstract:

    AbstrAct AntimicrobiAl resistAnce resulting in ineffective treAtment of infectious diseAses is An increAsing globAl problem, pArticulArly in infections with pAthogenic bActeriA. In some bActeriA, such As Streptococcus pyogenes, the pAthogenicity is strongly linked to the AttAchment of virulence fActors. Their AttAchment to the cellulAr membrAne is A trAnspeptidAtion reAction, cAtAlyzed by SortAse enzymes. As such, SortAses pose An interesting tArget for the development of new Antivirulence strAtegies thAt could yield novel AntimicrobiAl drugs. Using the substitution-oriented frAgment screening (SOS) ApproAch, we discovered A potent And specific inhibitor (C10) of SortAse A from S. pyogenes. The inhibitor C10 showed high specificity towArds S. pyogenes SortAse A, with An IC50 vAlue of 10 μM And A Kd of 60 μM. We envision thAt this inhibitor could be employed As A stArting point for further explorAtion of SortAse's potentiAl As therApeutic tArget for AntimicrobiAl drug development.