Organizing MS/MS Proteomic Data for Publication with Scaffold

1. Organizing MS/MS Proteomic Data for Publication with Scaffold Brian C. Searle Proteome Software Inc. Portland, Oregon ABRF2009, Memphis TN February 10th, 2009 Creative Commons Attribution

2. Publication Standards • In 2006 MCP published guidelines for reporting peptide and protein identifications • Other proteomics journals have adopted similar standards

3. What do the Guidelines Do? The guidelines ensure enough information to: • Understand and critically assess the results • Enforce a low level of reliability • Provide enough data concerning potentially questionable results to allow for reassessment

4. What do the Guidelines Do? The guidelines ensure enough information to: • Understand and critically assess the results • Enforce a low level of reliability • Provide enough data concerning potentially questionable results to allow for reassessment Publication guidelines have played a critical role in the acceptance of proteomic analysis

5. Publication Standards are Hard!

6. Publication Standards are Hard! They're hard for: • the authors who have to comply

7. Publication Standards are Hard! They're hard for: • the authors who have to comply • the reviewers who must police compliance

8. Publication Standards are Hard! They're hard for: • the authors who have to comply • the reviewers who must police compliance • the journals because there’s a huge amount of supplemental data in non-standard formats • PowerPoint guarantee readability, but difficult to use • RAW file formats can expire, but are much more useful

9. Publication Standards are Hard! They're hard for: • the authors who have to comply • the reviewers who must police compliance • the journals because there’s a huge amount of supplemental data in non-standard formats But they don’t have to be!

10. Scaffold Goals • Make it easier to organize data • Make tables useful for publication • Make figures that could be dropped directly into manuscripts • Clear fit when the guidelines were announced

11. Scaffold Makes theGuidelines Easier for Authors • Collate relevant data from search engine files • Most search engines supported: Mascot, SEQUEST, X! Tandem, Phenyx, SpectrumMill, OMSSA, IdentityE

12. Scaffold Makes theGuidelines Easier for Authors • Collate relevant data from search engine files • Most search engines supported: Mascot, SEQUEST, X! Tandem, Phenyx, SpectrumMill, OMSSA, IdentityE • Collapse related parameters across all files • Clearly point out missing metadata • Produce comparable search engine and instrument independent probabilities

13. Collate Relevant Data • Peak picking software, version, altered parameters • Database Selection • Database name and version • Species restriction • Number of proteins searched • Database search parameters • Search engine name and version • Enzyme specificity • # missed cleavages • Fixed/variable modifications • Mass tolerances • Peptide selection criteria

14. Collate Relevant Data • Peak picking software, version, altered parameters • Database Selection • Database name and version • Species restriction • Number of proteins searched • Database search parameters • Search engine name and version • Enzyme specificity • # missed cleavages • Fixed/variable modifications • Mass tolerances • Peptide selection criteria

15. Collate Relevant Data

16. Collate Relevant Data • This Button Gets: • Protein accession numbers • Number of unique peptides • % sequence coverage

17. Collate Relevant Data • While This Button Gets: • Peptide sequences identified • Precursor m/z and charge • Score and peptide probability

18. Collate Relevant Data • One hit wonders and modifications require validation • Distribute the Scaffold file to reviewers

19. Collate Relevant Data • Similar table and reports for iTRAQ and TMT quantitative data in Scaffold Q+ That’s it!

20. Making the Guidelines Easier for Reviewers to Police • All metadata is guaranteed to be present

21. Making the Guidelines Easier for Reviewers to Police All metadata is guaranteed to be present Quick to review key proteins and spectra

22. Making the Guidelines Easier for Reviewers to Police All metadata is guaranteed to be present Quick to review key proteins and spectra Easy to validate statistical assumptions

23. Probability Assumptions Usually Work

24. But Not Always!

25. Distributing Guideline Compliant Data • Journals can distribute Scaffold files containing the entire data set

26. Distributing Guideline Compliant Data • Journals can distribute Scaffold files containing the entire data set • Authors can share their raw data • Peak list exports for a variety of formats (MGF, DTA, PKL, etc)

27. Distributing Guideline Compliant Data • Journals can distribute Scaffold files containing the entire data set • Authors can share their raw data • Peak list exports for a variety of formats (MGF, DTA, PKL, etc) • Journals are allowed to distribute the original version of the free viewer so the files can ALWAYS be opened!

28. Conclusions Scaffold makes it: • Easier to pass publishing data guidelines • Easier to review data and to police standards • Easier to safely distribute supplemental material in a useful format