HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: T500010-MCP200v1 4/9/1391    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Glossary Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Che, F.-Y. Articles by Fricker, L. D. Search for Related Content PubMed PubMed Citation Articles by Che, F.-Y. Articles by Fricker, L. D. Social Bookmarking                      What's this?

Molecular & Cellular Proteomics 4:1391-1405, 2005.
© 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Technology

Quantitative Neuropeptidomics of Microwave-irradiated Mouse Brain and Pituitary^*

Fa-Yun Che, Jihyeon Lim, Hui Pan, Reeta Biswas and Lloyd D. Fricker

From the Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461

In neuropeptidomics, the degradation of a small fraction of abundant proteins overwhelms the low signals from neuropeptides, and many neuropeptides cannot be detected by mass spectrometry without extensive purification. Protein degradation was prevented when mice were sacrificed with focused microwave irradiation, permitting the detection of hypothalamic neuropeptides by mass spectrometry. Here we report an alternative and very simple method utilizing an ordinary microwave oven to inhibit enzymatic degradation. We used this technique to identify brain and pituitary neuropeptides. Quantitative analysis using mass spectrometry in combination with stable isotopic labeling was performed to determine the effect of microwave irradiation on relative levels of neuropeptides and protein degradation fragments. Microwave irradiation greatly reduced the levels of degradation fragments of proteins. In contrast, neuropeptide levels were increased about 2–3 times in hypothalamus by the microwave irradiation but not increased in pituitary. In a second experiment, three brain regions (hypothalamus, hippocampus, and striatum) from microwave-irradiated mice were analyzed. Altogether 41 neuropeptides or fragments of secretory pathway proteins were identified after microwave treatment; some of these are novel. These peptides were derived from 15 proteins: proopiomelanocortin, proSAAS, proenkephalin, preprotachykinins A and B, provasopressin, prooxytocin, melanin-concentrating hormone, proneurotensin, chromogranins A and B, secretogranin II, prohormone convertases 1 and 2, and peptidyl amidating monooxygenase. Although some protein degradation fragments were still found after microwave irradiation, these appear to result from protein breakdown during the extraction and not to an enzymatic reaction during the postmortem period. Two of the protein fragments corresponded to novel protein forms: VAP-33 with a 7-residue N-terminal extension and ß tubulin with a glutathione on the Cys near the N terminus. In conclusion, microwave irradiation with an ordinary microwave oven effectively inhibits enzymatic postmortem protein degradation, increases the recovery of neuropeptides, and makes it possible to conduct neuropeptidomic studies with mouse brain tissues.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				A. S. Heimann, I. Gomes, C. S. Dale, R. L. Pagano, A. Gupta, L. L. de Souza, A. D. Luchessi, L. M. Castro, R. Giorgi, V. Rioli, et al. Hemopressin is an inverse agonist of CB1 cannabinoid receptors PNAS, December 18, 2007; 104(51): 20588 - 20593. [Abstract] [Full Text] [PDF]

				L. D. Fricker Neuropeptidomics to Study Peptide Processing in Animal Models of Obesity Endocrinology, September 1, 2007; 148(9): 4185 - 4190. [Abstract] [Full Text] [PDF]

				M. Falth, K. Skold, M. Svensson, A. Nilsson, D. Fenyo, and P. E. Andren Neuropeptidomics Strategies for Specific and Sensitive Identification of Endogenous Peptides Mol. Cell. Proteomics, July 1, 2007; 6(7): 1188 - 1197. [Abstract] [Full Text] [PDF]