Abstract

RNA-based protein synthesis produces l-amino acid-containing proteins and peptides. d-amino acid-containing peptides (DAACPs) can be generated from l-amino acid peptides via post-translational modification. In the nervous system, the conformational change of a single l-amino acid in a peptide to its d-form results in altered bioactivity, with some DAACPs having orders-of-magnitude enhanced efficacy. However, this modification is often overlooked when characterizing endogenous peptides. Here, with the use of matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF)/TOF mass spectrometry, neuropeptides that have the second residue isomerized to the d-isoform are distinguished from their l-epimers via differences in the relative amounts of specific fragment ions during tandem MS. With the appropriate fragment ions chosen, and in some cases with the use of metal adducts, epimer discrimination is optimized. Specifically, the cardioexcitatory peptide Asn-DTrp-Phe-amide (NdWFa) was assayed directly from neurons isolated from the sea slug Aplysia californica; the fraction of the peptide with the second residue (W) in the d- versus l-form was 90 ± 10%. We demonstrate that this approach is well suited for confirming DAACPs directly from cells and tissue, advancing our understanding of the l to d modification and the role it plays in cell-to-cell signaling.

Original languageEnglish (US)
Pages (from-to)2794-2800
Number of pages7
JournalAnalytical chemistry
Volume83
Issue number7
DOIs
StatePublished - Apr 1 2011

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Neuropeptides
Neurons
Mass spectrometry
Amino Acids
Peptides
Ions
Cell signaling
Neurology
Bioactivity
Amides
Ionization
Desorption
Protein Isoforms
Proteins
Metals
RNA
Tissue
Lasers

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

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title = "Distinguishing endogenous d-amino acid-containing neuropeptides in individual neurons using tandem mass spectrometry",
abstract = "RNA-based protein synthesis produces l-amino acid-containing proteins and peptides. d-amino acid-containing peptides (DAACPs) can be generated from l-amino acid peptides via post-translational modification. In the nervous system, the conformational change of a single l-amino acid in a peptide to its d-form results in altered bioactivity, with some DAACPs having orders-of-magnitude enhanced efficacy. However, this modification is often overlooked when characterizing endogenous peptides. Here, with the use of matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF)/TOF mass spectrometry, neuropeptides that have the second residue isomerized to the d-isoform are distinguished from their l-epimers via differences in the relative amounts of specific fragment ions during tandem MS. With the appropriate fragment ions chosen, and in some cases with the use of metal adducts, epimer discrimination is optimized. Specifically, the cardioexcitatory peptide Asn-DTrp-Phe-amide (NdWFa) was assayed directly from neurons isolated from the sea slug Aplysia californica; the fraction of the peptide with the second residue (W) in the d- versus l-form was 90 ± 10{\%}. We demonstrate that this approach is well suited for confirming DAACPs directly from cells and tissue, advancing our understanding of the l to d modification and the role it plays in cell-to-cell signaling.",
author = "Lu Bai and Romanova, {Elena V.} and Sweedler, {Jonathan V.}",
year = "2011",
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language = "English (US)",
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TY - JOUR

T1 - Distinguishing endogenous d-amino acid-containing neuropeptides in individual neurons using tandem mass spectrometry

AU - Bai, Lu

AU - Romanova, Elena V.

AU - Sweedler, Jonathan V.

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Y1 - 2011/4/1

N2 - RNA-based protein synthesis produces l-amino acid-containing proteins and peptides. d-amino acid-containing peptides (DAACPs) can be generated from l-amino acid peptides via post-translational modification. In the nervous system, the conformational change of a single l-amino acid in a peptide to its d-form results in altered bioactivity, with some DAACPs having orders-of-magnitude enhanced efficacy. However, this modification is often overlooked when characterizing endogenous peptides. Here, with the use of matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF)/TOF mass spectrometry, neuropeptides that have the second residue isomerized to the d-isoform are distinguished from their l-epimers via differences in the relative amounts of specific fragment ions during tandem MS. With the appropriate fragment ions chosen, and in some cases with the use of metal adducts, epimer discrimination is optimized. Specifically, the cardioexcitatory peptide Asn-DTrp-Phe-amide (NdWFa) was assayed directly from neurons isolated from the sea slug Aplysia californica; the fraction of the peptide with the second residue (W) in the d- versus l-form was 90 ± 10%. We demonstrate that this approach is well suited for confirming DAACPs directly from cells and tissue, advancing our understanding of the l to d modification and the role it plays in cell-to-cell signaling.

AB - RNA-based protein synthesis produces l-amino acid-containing proteins and peptides. d-amino acid-containing peptides (DAACPs) can be generated from l-amino acid peptides via post-translational modification. In the nervous system, the conformational change of a single l-amino acid in a peptide to its d-form results in altered bioactivity, with some DAACPs having orders-of-magnitude enhanced efficacy. However, this modification is often overlooked when characterizing endogenous peptides. Here, with the use of matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF)/TOF mass spectrometry, neuropeptides that have the second residue isomerized to the d-isoform are distinguished from their l-epimers via differences in the relative amounts of specific fragment ions during tandem MS. With the appropriate fragment ions chosen, and in some cases with the use of metal adducts, epimer discrimination is optimized. Specifically, the cardioexcitatory peptide Asn-DTrp-Phe-amide (NdWFa) was assayed directly from neurons isolated from the sea slug Aplysia californica; the fraction of the peptide with the second residue (W) in the d- versus l-form was 90 ± 10%. We demonstrate that this approach is well suited for confirming DAACPs directly from cells and tissue, advancing our understanding of the l to d modification and the role it plays in cell-to-cell signaling.

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