Proteomic analysis of platelet alpha-granules using mass spectrometry.
BACKGROUND
Platelets have three major types of secretory organelles: lysosomes, dense granules, and alpha-granules. alpha-Granules contain several adhesive proteins involved in hemostasis, as well as glycoproteins involved in inflammation, wound healing, and cell-matrix interactions. This article represents the first effort to define the platelet alpha-granule proteome using mass spectrometry (MS).


METHODS
We prepared a subcellular fraction enriched in intact alpha-granules from human platelets using sucrose gradient ultracentrifugation. alpha-Granule proteins were separated and identified using sodium dodecylsulfate polyacrylamide gel electrophoresis and liquid chromatography-tandem MS.


RESULTS
In the sucrose fraction enriched in alpha-granules, we identified 284 non-redundant proteins, 44 of which appear to be new alpha-granule proteins, on the basis of a literature review. Immunoelectron microscopy confirmed the presence of Scamp2, APLP2, ESAM and LAMA5 in platelet alpha-granules for the first time. We identified 65% of the same proteins that were detected in the platelet releasate (J. A. Coppinger et al. [Blood 2004;103: 2096-104]) as well as additional soluble and membrane proteins. Our method provides a suitable tool for analyzing the granule proteome of patients with storage pool deficiencies.
Maynard
D M
DM
Section on Human Biochemical Genetics, Medical Genetics Branch, National Human Genome Research Institute, NIH, Bethesda, MD 20892-1851, USA. maynardd@mail.nih.gov
Heijnen
H F G
HF
Horne
M K
MK
White
J G
JG
Gahl
W A
WA
D001792:Blood Platelets / Q000648:ultrastructure*; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D013058:Mass Spectrometry; D046529:Microscopy, Electron, Transmission; D015388:Organelles / Q000378:metabolism* / Q000648:ultrastructure; D040901:Proteomics