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ADPr-Peptide Synthesis

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ADP-ribosylation and NAD+ Utilizing Enzymes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1813))

Abstract

Synthetic mono-ADPr-peptides are useful for structural, biochemical, and proteomics studies. We describe here a protocol for the preparation of mono-ADPr-peptides based on a fairly standard Fmoc-based solid-phase synthesis. Phosphoribosylated precursor building blocks are introduced into the peptide chain on solid-phase and subsequently converted to ADPr-sites by chemical phosphorylation with adenosine phosphoramidite. Suitably protected phosphoribosylated glutamine, asparagine, and citrulline building blocks described in this protocol allow introduction of ADP-Gln, ADPr-Asn, and ADPr-Cit into peptide chains as demonstrated for three peptides. Trifunctional amino acids, for which base-sensitive side-chain protection is available, can be accommodated in the sequences flanking the ADPr-cites.

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Abbreviations

ACN:

Acetonitrile

AcOH:

Acetic acid

CSO:

(1S)-(+)-(10-camphorsulfonyl)-oxaziridine

DBU:

1,8-diazabicycloundec-7-ene

DCM:

Dichloromethane

DiPEA:

Diisopropylethylamine

Dmab:

4-(N-[1-(4,4-dimethyl-2,6-dioxocyclohexylidene)-3-methylbutyl]amino) benzyl ester

DMF:

Dimethylformamide

Et2O:

Diethyl ether

EtOH:

Ethanol

HCTU:

O-(1H-6-Chlorobenzotriazole-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate

HFIP:

1,1,1,3,3,3-hexafluoro-2-propanol

NMP:

N-methylpyrrolidone

TBDPS:

tert-butyldiphenylsilyl

TBSOTf:

tert-butyldimethylsilyl triflate

TEA:

Triethylamine

Tfa:

Trifluroacetamide (protective group)

TFA:

Trifluoroacetic acid

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Acknowledgments

This work was supported by the Netherlands Organization for Scientific Research (NWO).

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Authors and Affiliations

  1. Gorlaeus Laboratories, Leiden Institute of Chemistry, Universiteit Leiden, Leiden, The Netherlands

    Hans A. V. Kistemaker, Jim Voorneveld & Dmitri V. Filippov

Authors
  1. Hans A. V. Kistemaker
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  2. Jim Voorneveld
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  3. Dmitri V. Filippov
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Corresponding author

Correspondence to Dmitri V. Filippov .

Editor information

Editors and Affiliations

  1. Department of Biology, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA

    Paul Chang

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Kistemaker, H.A.V., Voorneveld, J., Filippov, D.V. (2018). ADPr-Peptide Synthesis. In: Chang, P. (eds) ADP-ribosylation and NAD+ Utilizing Enzymes. Methods in Molecular Biology, vol 1813. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-8588-3_24

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  • DOI: https://doi.org/10.1007/978-1-4939-8588-3_24

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-8587-6

  • Online ISBN: 978-1-4939-8588-3

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