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Measuring D-amino acid-containing neuropeptides with capillary electrophoresis

Department of Chemistry & Beckman Institute, University of Illinois, 600 South Mathews Ave., 63-5, Urbana, IL 61801, USA.
The Analyst (Impact Factor: 4.11). 09/2005; 130(8):1198-203. DOI: 10.1039/b504717j
Source: PubMed

ABSTRACT

Neuropeptides are heavily posttranslationally modified (PTM) gene products that are often characterized by a variety of mass spectrometric approaches. Recently, the occurrence of amino acids in the D-form has been documented in several neuropeptides. As this modification has no associated mass shift, this particular PTM is difficult to evaluate using mass spectrometry (MS) alone. Here we demonstrate several approaches using capillary electrophoresis (CE) with absorbance and laser-induced fluorescence (LIF) for the separation of native and derivatized molluscan peptides containing D-amino acids. The combination of peptide derivatization followed by CE/LIF is well suited for single cell measurements because of its ability to characterize the peptides in such small samples. In order to verify this approach, the D-Trp-containing peptide NdWFa (NH2-Asn-D-Trp-Phe-CONH2), present in individual neurons from the marine mollusk Aplysia californica, has been characterized. The mass spectra show that NdWFa and/or NWFa are present in specific neurons; CE/LIF analysis of these cells demonstrates that NdWFa is the dominant form of the peptide.

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    • "excited fluorescence and single laser sources (Gostkowski et al., 1998; Sheeley et al., 2005). Other, straightforward strategies have also been used, such as those dependent on D-amino acid oxidase (Nagata et al., 1985) or more specifically on D-aspartate oxidase (Fisher et al., 1992; Nagata et al., 1985). "
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