SPECTRA LIBRARY ASSISTED DE NOVO PEPTIDE SEQUENCING FOR HCD AND ETD SPECTRA PAIRS

Transcription

1 SPECTRA LIBRARY ASSISTED DE NOVO PEPTIDE SEQUENCING FOR HCD AND ETD SPECTRA PAIRS 1 Yan Yan Department of Computer Science University of Western Ontario, Canada

2 OUTLINE Background Tandem mass spectrometry Peptide sequencing methods Proposed method Use of spectra libraries Spectra merging Peptide tags De novo sequencing model Experiments and results Data Experiments and comparison Conclusions 2

3 BACKGROUND Mass spectrometry (MS) An analytical technique measuring mass-to-charge ratio (m/z) of individual compounds. Tandem mass spectrometry (MS/MS) It contains two or more mass analyzers. It breaks the compounds into smaller fragment ions. Fragmentation techniques Collision-induced dissociation (CID) High-energy collisional dissociation (HCD) Electron transfer dissociation (ETD) MS/MS experiments Input: protein samples Output: tandem mass (MS/MS)spectra 3

4 MS/MS PROCESS Protein sample Sample preparation Protein digestion enzyme Peptide sample Peptide separation MS Ion source Mass analyzer 1 MS/MS Precursor ions of interest Detector Mass analyzer 2 Fragment ions Fragmentation CID/HCD/ETD/ 4

5 BACKGROUND Different ion types of MS/MS There are commonly 6 different ions,and they form 3 complimentary ion pairs. B-ionsand y-ionsare themost common ionsin CID/HCD spectra. C-ionsand z-ions are common in ETD spectra. Losing small molecules, such as ammonia (NH 3 ) and water (H 2 O). Peptide fragmentation notation. 5

6 TWO WAYS OF PEPTIDE SEQUENCING 6 Frank et al. JPR

7 BACKGROUND Spectra library Set of annotated experimental MS/MS spectra Publicly availablelibraries chemdata.nist.gov Additional informationhelp with de novo sequencing 7

8 PROPOSED METHOD Spectra merging Peak selection (within each spectrum) Find all length 2 paths ;select middle peak output i j k m aa1 m aa2 Find complimentary ion pairs and select them Output: all selected peaks form one spectrum S 8

9 PROPOSED METHOD Improved spectra merging -- use spectra libraries as training sets Calculate significant scores Middle ion of the 2-tags Complementary ion pairs 9

10 PROPOSED METHOD Improved spectra merging -- use spectra libraries as training sets Calculate significant scores Middle ion of the 2-tags Complementary ion pairs Assign significant scores on peaks during selection Peak I (m/z, charge, score) 10

11 PROPOSED METHOD De novo sequencing Generate peptide tags from both spectra Graph with multiple edge types* (GMET) Tag1 Tag2 N-terminal C-terminal 11 *:Y. Yan, A. Kusalik, and F.-X. Wu, NovoHCD: De novo peptide sequencing from HCD spectra, NanoBioscience, IEEE Transactions on, vol. 13, no. 2, pp , June 2014.

12 PROPOSED METHOD Peptide tag improvement Generate length-3 tags Merge tags with two successive amino acids overlap (and ions overlap) E.g. t i = TAG and t j = AGT t ij = TAGT Assign significant scores to tags Rank candidate peptides with significant scores 12

13 EXPERIMENTS AND RESULTS Experiment MS/MS spectra dataset Dataset # of spectra Spectrum charge Selected pairs SCX_HCD_decon 1952 SCX_ETD_decon 612 SCX_HCD_no_decon 2557 SCX_ETD_no_decon to to Spectra libraries human peptide spectral library (from chemdata.nist.gov) of 183,140 HCD spectra 100,000 ETD spectra of synthetic, unmodified peptides 13

14 EXPERIMENTS AND RESULTS Results Significant score calculated using spectra libraries 14

15 EXPERIMENTS AND RESULTS Results Full length accuracy top three candidates output 15

16 EXPERIMENTS AND RESULTS Results Accuracy comparison with different output SCX_HCD_decon and SCX_ETD_decon dataset pair SCX_HCD_no_decon and SCX_ETD_no_decon dataset pair 16

17 EXPERIMENTS AND RESULTS Results Computational time 17

18 CONCLUSIONS Spectra libraries adds additional information for better peptide sequencing performance. Full length accuracy increases up to 11% when only the highest ranked candidates output. Computation time saves up to 40% when merged long peptide tags were used. 18

19 19 THANK YOU

20 FUTURE WORK PLAN Experiments Compare with more methods Use more datasets Algorithm improvement Other ways of selecting signal peaks to merge spectra Spectrum specific features Method development Multiple spectra sequencing 20

21 BACKGROUND Peptides Peptides are organic compounds consisting of 2 or more amino acids. There are 20 standard amino acids in nature. Two amino acids connect to each other by forming a peptide bondand losinga molecule ofwater. Amino acid structure. Dec Formation of a peptide from two amino acids. Dec

22 BACKGROUND ECD/ETD spectra New data that has been available in recent years. C-Ions and z-ions are dominant in ETD/ECD spectra. Ions lose small molecules, such as ammonia (NH3) and water (H2O). 22

23 BACKGROUND CID/HCD spectra Common ion types and mass calculation 23

24 BACKGROUND ECD/ETD spectra Common ion types and mass calculation 24

25 BACKGROUND Complimentary ion pairs In CID/HCD b i -ion + y n-i -ion = MH + +H = M parent + 2H In ECD/ETD c i -ion + z n-i -ion = M parent + 3H (c i -1 )-ion + (z n-i +1)-ion = M parent + 3H 25

26 PROPOSED METHOD Spectra merging Peak selection (within each spectrum) 1. Find all length 2 paths ;output middle peak Relationship : amino acid difference Ions: regular or with loss of small molecules output i j k m aa1 m aa2 2. Find complimentary ion pairs and output Ions: consider different charges Output peaks to form spectra S 26

27 PROPOSED METHOD (HAVE PROBLEMS!!) De novo sequencing Tags Find 3-tags from each of the original spectrum (denoted as S " and S # ) Find peaks associated with a tag t in S Calculate AACs separated by t (use P %&'' ) Threshold control if more tags needed Extend paths to form Graphs with multiple edge type (GMETs) with AAC restriction Ion types including b/y- and c/z-ions Ion type determined by previous step Assembling tags and partial peptides from GMETs to be candidate peptides 27

28 EXPERIMENTS AND RESULTS Experiment Data Select spectra pairs having the same peptide sequences 28 *:Results inferred by previous published two methods for HCD and ExD data, respectively.

29 EXPERIMENTS AND RESULTS Experiment Data Select spectra pairs having the same peptide sequences 29 *:Results inferred by previous published two methods for HCD and ExD data, respectively.