Organellar proteomics: analysis of pancreatic zymogen granule membranes.
The zymogen granule (ZG) is the specialized organelle in pancreatic acinar cells for digestive enzyme storage and regulated secretion and has been a model for studying secretory granule functions. In an initial effort to comprehensively understand the functions of this organelle, we conducted a proteomic study to identify proteins from highly purified ZG membranes. By combining two-dimensional gel electrophoresis and two-dimensional LC with tandem mass spectrometry, 101 proteins were identified from purified ZG membranes including 28 known ZG proteins and 73 previously unknown proteins, including SNAP29, Rab27B, Rab11A, Rab6, Rap1, and myosin Vc. Moreover several hypothetical proteins were identified that represent potential novel proteins. The ZG localization of nine of these proteins was further confirmed by immunocytochemistry. To distinguish intrinsic membrane proteins from soluble and peripheral membrane proteins, a quantitative proteomic strategy was used to measure the enrichment of intrinsic membrane proteins through the purification process. The iTRAQ ratios correlated well with known or Transmembrane Hidden Markov Model-predicted soluble or membrane proteins. By combining subcellular fractionation with high resolution separation and comprehensive identification of proteins, we have begun to elucidate zymogen granule functions through proteomic and subsequent functional analysis of its membrane components.
Chen
Xuequn
X
Department of Biological Chemistry, The University of Michigan, Ann Arbor, Michigan 48109, USA. xuequnc@umich.edu
Walker
Angela K
AK
Strahler
John R
JR
Simon
Eric S
ES
Tomanicek-Volk
Sarah L
SL
Nelson
Bradley B
BB
Hurley
Mary C
MC
Ernst
Stephen A
SA
Williams
John A
JA
Andrews
Philip C
PC
D000818:Animals; D004792:Enzyme Precursors; D007425:Intracellular Membranes / Q000737:chemistry* / Q000378:metabolism*; D008565:Membrane Proteins / Q000032:analysis* / Q000302:isolation & purification; D010179:Pancreas / Q000166:cytology*; D021381:Protein Transport; D040901:Proteomics; D051381:Rats; D022142:Secretory Vesicles / Q000737:chemistry / Q000378:metabolism*
D004792:Enzyme Precursors; D008565:Membrane Proteins