Mass Spectrometry of Cardiac Calsequestrin Characterizes Microheterogeneity Unique to Heart and Indicative of Complex Intracellular Transit
| Autor: | Steven E. Cala, Arash Kiarash, Jeffrey J. O'Brian, Michal L. Ram |
|---|---|
| Rok vydání: | 2002 |
| Předmět: |
Spectrometry
Mass Electrospray Ionization Glycan Glycosylation Calsequestrin Biochemistry Cell Line symbols.namesake Dogs Animals Humans Protein Isoforms Myocyte Phosphorylation Molecular Biology Secretory pathway Molecular mass biology Chemistry Myocardium Tunicamycin Endoplasmic reticulum Cell Biology Golgi apparatus musculoskeletal system Recombinant Proteins Cell biology Protein Transport cardiovascular system symbols biology.protein tissues |
| Zdroj: | Journal of Biological Chemistry. 277:37154-37160 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m204370200 |
| Popis: | Cardiac calsequestrin concentrates in junctional sarcoplasmic reticulum in heart and skeletal muscle cells by an undefined mechanism. During transit through the secretory pathway, it undergoes an as yet uncharacterized glycosylation and acquires phosphate on CK2-sensitive sites. In this study, we have shown that active calsequestrin phosphorylation occurred in nonmuscle cells as well as muscle cells, reflecting a widespread cellular process. To characterize this post-translational modification and resolve individual molecular mass species, we subjected purified calsequestrin to mass spectrometry using electrospray ionization. Mass spectra showed that calsequestrin glycan structure in nonmuscle cells was that expected for an endoplasmic reticulum-localized glycoprotein and showed that each glycoform existed as four mass peaks representing molecules that also had 0-3 phosphorylation sites occupied. In heart, mass peaks indicated carbohydrate modifications characteristic of transit through Golgi compartments. Phosphorylation did not occur on every glycoform present, suggesting a far more complex movement of calsequestrin molecules in heart cells. Significant amounts of calsequestrin contained glycan with only a single mannose residue, indicative of a novel post-endoplasmic reticulum mannosidase activity. In conclusion, glyco- and phosphoforms of calsequestrin chart a complex cellular transport in heart, with calsequestrin following trafficking pathways not present or not accessible to the same molecules in nonmuscle. |
| Databáze: | OpenAIRE |
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