Neuropeptide profiling of the bovine hypothalamus: Thermal stabilization is an effective tool in inhibiting post-mortem degradation

• Published in: Proteomics (Weinheim) 2011-04, Vol.11 (7), p.1264-1276
• Main Authors: Colgrave, Michelle L., Xi, Li, Lehnert, Sigrid A., Flatscher-Bader, Traute, Wadensten, Henrik, Nilsson, Anna, Andren, Per E., Wijffels, Gene
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Analytical, structural and metabolic biochemistry; Animal proteomics; Animals; Autopsy; Biological and medical sciences; Bovine; Cattle; Chromatography, Liquid; Electrophoresis, Gel, Two-Dimensional; FARMACI; Female; Fundamental and applied biological sciences. Psychology; Hormones; Hot Temperature; Hypothalamus; Hypothalamus - chemistry; Hypothalamus - metabolism; Mass Spectrometry; Mice; Miscellaneous; Molecular Sequence Data; Neuropeptide; Neuropeptides; Neuropeptides - analysis; Neuropeptides - metabolism; Peptide Fragments - analysis; PHARMACY; Pituitary gland; Post-mortem degradation; Postmortem Changes; Proprotein Convertases - analysis; Proprotein Convertases - metabolism; Protein Array Analysis; Protein Denaturation; Protein Processing, Post-Translational; Protein Stability; Proteins; Reproduction; Species Specificity; Stabilization
• ISSN: 1615-9853; 1615-9861; 1615-9861
• DOI: 10.1002/pmic.201000423

The hypothalamus is the central regulatory region of the brain that links the nervous system to the endocrine system via the pituitary gland. It synthesizes and secretes neuropeptide hormones, which in turn act to stimulate or inhibit the secretion of pituitary hormones. We have undertaken a detailed MS investigation of the peptides present in the bovine hypothalamus by adapting a novel heat stabilization methodology, which improved peptide discovery to direct our studies into the molecular mechanisms involved in bovine reproduction. The untreated samples contained large numbers of protein degradation products that interfered with the analysis of the neuropeptides. In the thermally stabilized samples, we were able to identify many more neuropeptides that are known to be expressed in the bovine hypothalamus. Furthermore, we have characterized a range of post‐translational modifications that indicate the presence of processed intact mature neuropeptides in the stabilized tissue samples, whereas we detected many trimmed or truncated peptides resulting from post‐mortem degradation in the untreated tissue samples. Altogether, using an optimized workflow, we were able to identify 140 candidate neuropeptides. We also nominate six new candidate neuropeptides derived from proSAAS, secretogranin‐2 and proTRH.