Revision of Zymogen Granule Membrane Topology from Mon, 2010-05-10 14:45

Data Set Type: 

Proteomics
The University of Michigan, Ann Arbor, Michigan 48109
AttachmentSize
Office spreadsheet icon ZG Mol Topology dataset.xls215 KB

Molecular Topology of Zymogen Granule Proteins

This data set is from a study published in Molecular & Cellular Proteomics (7, 2323-2336, 2008) which used iTRAQ isobaric tags to label peptide from ZGs treated with proteinase K and separately peptides from untreated ZGs. Peptides from regions of the proteins outside the ZG membrane should be subject to proteinase K digestion. The data set posted here was obtained from this global topology analysis of ZG membrane proteins and included in the paper as Supplemental Table 3.

The results showed that tryptic peptides of ZGM proteins were separated into two distinct clusters according to their iTRAQ ratios for proteinase K treated vs control ZGs. The low iTRAQ ratio cluster included cytoplasm-orientated membrane and membrane-associated proteins and the second cluster having unchanged ratios included predominantly lumenal proteins. In order to more accurately assign the topology, we developed a statistical mixture model to provide probabilities for identified peptides to be cytoplasmic or luminal based on their iTRAQ ratios.

Using the statistical model, we obtained probabilities of being cytoplasmic or luminal for all 654 unique tryptic peptides detected in two independent studies (483 from experiment 1 and 484 from experiment 2) (Supplemental Table 3). All peptides were separated into two categories, cytoplasmic (color-coded as red) if cytoplasmic probability >0.50 or luminal (blue) if cytoplasmic probability <0.50 (Supplemental Tables 3). 313 peptides were common in both experiments among which 308 peptides (~98%) had consistent topology assignments and only 5 peptides (~2%) had inconsistent topology assignments in two experiments. These peptides led to the identification of 285 non-redundant proteins together with their topology information derived from the corresponding iTRAQ ratios. The protein names, accession numbers, peptide sequences, their iTRAQ ratios in two independent studies and corresponding cytoplasmic probabilities are summarized in Supplemental Table 3.

After a database and literature search, 73 highly likely contaminating mitochondrial and ribosomal proteins, indicated in the column "Likely contaminants" (Supplemental Table 3), were excluded from further analysis. By implementing this approach to global topology analysis of ZGM proteins, we obtained an experimentally constrained, comprehensive topology model of identified ZG membrane proteins.