Combining Bottom-Up and Top-Down Mass Spectrometric Strategies for De Novo Sequencing of the Crustacean Hyperglycemic Hormone from Cancer borealis
peer-reviewed · Analytical Chemistry · 2009
| Date | 2009-01-01 |
| Type | peer-reviewed |
| Venue | Analytical Chemistry |
| Publisher | American Chemical Society (ACS) |
| Contribution | downstream-application |
| DOI | 10.1021/ac801910g |
| Citations (OpenAlex) | 32 |
| Venue 2-year citedness | 7.10 |
Abstract
The crustacean hyperglycemic hormone (CHH) is a 72-amino acid residue polypeptide with multiple physiological effects. The X-organ/sinus gland is the primary source for CHH and its family members. However, the amino acid sequence of CHH in Cancer borealis , a premier model system for neuromodulation, has not been characterized. In this study, a novel hybrid strategy combining “bottom-up” and “top-down” methodologies enabled direct sequencing of CHH peptide in the sinus gland of C. borealis . Multiple mass spectrometry (MS)-based techniques were employed to characterize the CHH peptide, including direct tissue analysis by MALDI-FT-ICR-MS, de novo sequencing of tryptic digested CHH by nano-LC/ESI-Q-TOF MS and intact CHH analysis by LC/FT-ICR-MS. In-trap cleaning removed the extensive matrix adducts of CHH in the direct tissue analysis by MALDI-FT-ICR-MS. Fragmentation efficiency of the intact CHH was drastically improved after the reduction-alkylation of the disulfide bonds. The sequence coverage was further enhanced by employing multiple complementary fragmentation techniques. Overall, this example is the largest neuropeptide de novo sequenced in C. borealis by mass spectrometric methods.
Methods and tools
- Crustacean hyperglycemic hormone sequencing: Combines bottom-up and top-down mass spectrometry to sequence the 72-residue crustacean hyperglycemic hormone of Cancer borealis de novo, the sequence having been unavailable for this species.
Cited by (4)
- High-definition De Novo Sequencing of Crustacean Hyperglycemic Hormone (CHH)-family Neuropeptides (2012) both
- Algorithms for the de novo sequencing of peptides from tandem mass spectra (2011) crossref
- Extensive <i>De Novo</i> Sequencing of New Parvalbumin Isoforms Using a Novel Combination of Bottom-Up Proteomics, Accurate Molecular Mass Measurement by FTICR−MS, and Selected MS/MS Ion Monitoring (2010) crossref
- Top-Down <i>de Novo</i> Protein Sequencing of a 13.6 kDa Camelid Single Heavy Chain Antibody by Matrix-Assisted Laser Desorption Ionization-Time-of-Flight/Time-of-Flight Mass Spectrometry (2010) crossref