LC-MS/MS using the Q-Trap The hybrid linear ion trap mass - - PDF document

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Articles that describe this mass spectrometer:

• Hager JW, Le Blanc JC. J. Chromatrogr. A,

2003; 1020 (1): 3-9

• Le Blanc JC, Hager JW, Ilisiu AM, Hunter

C, Zhong F, Chu I. Proteomics ,2003; 3(6): 859-69

• Hager JW, Le Blanc JC. Rapid Commun.

Mass Spectrom. 2003; 17(10):1056- 64.

The hybrid linear ion trap mass spectrometer (Q TRAP MS) from MDS Sciex

LC-MS/MS using the Q-Trap

Willy Bienvenut Cours de 3eme cycle Romand 03/03/04

The Q-trap MS Q Trap (Quadrupole – linear trap)

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Triple quadrupole mass spectrometer

Q1 Q3 Q0 N2 CAD Gas Q2 Normal quadrupole or linear ion trap 3x10-5 Torr

Ion accumulation Fragmentation Exit lens Aux AC Skimmer Entrance plate Detector

Q0: ion accumulator (on/off) Q1: First quadrupole Q2: Fragmentation cell Q3: third quadrupole or linear ion trap Q0 Q1 Q2 Q3

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Q1 Q3 Q0 Q2

Q-Trap utilisation modes

Trap scan Fragment Resolving (fixed) Enhance produced ion isolate/Fragment/Scan Fragment Resolving (fixed) MS3 Trap scan RF-only RF-only Enhance resolution Trap scan RF-only RF-only Enhance multiply charged ions Trap scan RF-only RF-only Enhance MS Resolving (fixed) Fragment Resolving (fixed) Multiple reaction monitoring Fragment resolving (scanning) Fragment Resolving (scanning) Neutral loss scan Resolving (fixed) Fragment Resolving (scanning) Precursor ion Fragment resolving (scanning) Fragment Resolving (fixed) Product ion Fragment resolving (scanning) Fragment Resolving (scanning) Triple quadrupoles (Q1 & Q3) Resolving (scanning) RF-only RF-only Q3 scan RF-only RF-only Resolving (scanning) Q1 scan Q3 Q2 Q1 Q-Trap mode

Mode Triple Quad. MS/MS

• Only 1 ion pair is stable at any one time.
• Poor scanning efficiency.
• Very efficient for MRM.
• Very selective scans available.

Q0 Q1 Q2 Q3

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Trap scan Fragment Resolving (fixed) Enhance produced ion Trap scan RF-only RF-only Enhance resolution Trap scan RF-only RF-only Enhance MS Q3 Q2 Q1 Q-Trap mode

Standard ESI-MS/MS modes

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Alternative survey scan: the PIS scan

The precursor ion scan approach: Looking for well defined ions m/z values

216 Ac-K 216 175 60 70 159 72 86 m/z ions p-Y y1 (R) S P W V I/L Corresponding AA

Usual approach for small molecules detection and quantitation, e.g. drugs and metabolites. Adaptable to low MW peptide fragments such as immonium ions

6.66 3.13 1.1 8 5.58 6.98 4.88 4.06 2.36 5.96 9.56 5.88 2.26 6.90 6.54 3.92 1.60 5.27 4.28 5.24 7.72

% in SWISS- PROT V Y W T S P F M K L I H G E N C D Q R A AA

TIC

(Total Ion Current)

EMS ER EPI 1 EPI 2 EPI 2

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TIC PIS 86 ER EPI 1 EPI 2

Extraction of the data from the acquired files

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MW 200000 400000 600000 800000 1000000 1200000 0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 pI MW (Da)

pI= 7.0 +/- 2.0 / MW= 57.5 +/- 105.7 kDa

Aldose reductase (EC 1.1.1.21) (AR) (Aldehyde redu ALDR_HUMAN P15121 6.55 35722 102 102 102 100 100 56 Annexin A5 (Annexin V) (Lipocortin V) (Endonexin I ANX5_HUMAN P08758 4.94 35806 13 13 13 13 17 23 Glyceraldehyde 3-phosphate dehydrogenase, muscle ( G3P1_HUMAN P00354 6.60 35876 23 23 23 21 24 45 Glyceraldehyde 3-phosphate dehydrogenase, liver (E G3P2_HUMAN P04406 8.58 35922 4 4 4 4 4 8 Malate dehydrogenase, cytoplasmic (EC 1.1.1.37). MDHC_HUMAN P40925 6.89 36295 62 62 62 62 61 54 L-lactate dehydrogenase B chain (EC 1.1.1.27) (LDH LDHB_HUMAN P07195 5.72 36507 8 6 7 6 7 11 L-lactate dehydrogenase A chain (EC 1.1.1.27) (LDH LDHA_HUMAN P00338 8.46 36558 10 10 9 8 8 7 42 Heterogeneous nuclear ribonucleoproteins A2/B1 (hn ROA2_HUMAN P22626 8.97 37430 42 42 43 43 39 Transaldolase (EC 2.2.1.2). TAL1_HUMAN P37837 6.36 37540 123 Heterogeneous nuclear ribonucleoprotein A1 (Helix- ROA1_HUMAN P09651 9.26 38715 47 47 47 47 47 47 4 Glyceraldehyde 3-phosphate dehydrogenase, testis-s G3PT_HUMAN O14556 8.39 44501 88 88 88 Keratin, type I cytoskeletal 18 (Cytokeratin 18) ( K1CR_HUMAN P05783 5.34 47927 106 106 106 107 104 Phosphatidylinositol N-acetylglucosaminyltransfera PIGA_HUMAN P37287 8.63 54127 119 118 119 Annexin A11 (Annexin XI) (Calcyclin-associated ann ANXB_HUMAN P50995 7.53 54390 116 116 117 116 101 52 Keratin, type II cytoskeletal 1 (Cytokeratin 1) (K K2C1_HUMAN P04264 8.16 65886 93 93 93 91 90 19 Merlin (Moesin-ezrin-radixin-like protein) (Schwan MERL_HUMAN P35240 6.11 69690 96 Glycerol-3-phosphate dehydrogenase, mitochondrial GPDM_HUMAN P43304 6.17 76323 101 90 U4/U6 small nuclear ribonucleoprotein Prp3 (Pre-mR PRP3_HUMAN O43395 9.50 77529 104 Zinc finger CW-type coiled-coil domain protein 1. ZCW1_HUMAN Q9Y6X9 8.82 110724 96 96 96 96 96 Potassium voltage-gated channel subfamily H member KCH5_HUMAN Q8NCM2 7.51 111907 113 Tyrosine-protein kinase JAK3 (EC 2.7.1.112) (Janus JAK3_HUMAN P52333 6.64 125016 118 117 118 119 115 Ras GTPase-activating protein nGAP (RAS protein ac NGAP_HUMAN Q9UJF2 8.52 128558 124 119 97 Carbamoyl-phosphate synthase [ammonia], mitochondr CPSM_HUMAN P31327 5.92 160549 112 112 112 111 94 Eukaryotic translation initiation factor 4 gamma 1 I4G1_HUMAN Q04637 5.27 175535 123 122 123 Phosphatidylinositol-4-phosphate 3-kinase C2 domai PK3B_HUMAN O00750 6.87 184858 113 113 113 112 110 Tuberin (Tuberous sclerosis 2 protein). TSC2_HUMAN P49815 7.06 200749 124 123 124 125 Kinesin-like protein KIF13B (Kinesin-like protein K13B_HUMAN Q9NQT8 5.56 202666 94 45 CAD protein [Includes: Glutamine-dependent carbamo PYR1_HUMAN P27708 6.00 242917 115 115 115 116 112 99 Retinal-specific ATP-binding cassette transporter ABCR_HUMAN P78363 5.89 255944 89 Spectrin beta chain, brain 2 (Spectrin, non-erythr SPCP_HUMAN O15020 5.79 271295 86 Neurofibromin (Neurofibromatosis-related protein N NF1_HUMAN P21359 7.10 319372 117 116 117 118 114 Desmoplakin (DP) (250/210 kDa paraneoplastic pemph DESP_HUMAN P15924 6.44 331776 103 Low-density lipoprotein receptor-related protein 1 LRP1_HUMAN Q07954 5.16 502634 122 121 122 122 117 Nesprin 1 (Nuclear envelope spectrin repeat protei SNE1_HUMAN Q8NF91 5.38 1011042 100 protein description SP name SP entry pI MW 0.05% 0.10% 0.2% 0.5% 1% 2% 5% Dermcidin precursor (Preproteolysin) [Contains: Su DCD_HUMAN P81605 5.64 9259 114 114 114 115 113 102 Multisynthetase complex auxiliary component p43 [C MCA1_HUMAN Q12904 6.80 18218 83 83 83 82 82 70 51 Tumor protein D52 (N8 protein). TD52_HUMAN P55327 4.94 19863 52 52 52 52 51 41 Histone H1.2 (Histone H1d). H12_HUMAN P16403 10.94 21234 29 29 29 31 29 24 18 Thioredoxin-dependent peroxide reductase, mitochon PDX3_HUMAN P30048 5.77 21468 89 89 89 87 86 73 Peroxiredoxin 1 (EC 1.11.1.-) (Thioredoxin peroxid PDX1_HUMAN Q06830 8.27 22110 45 45 45 45 44 71 Tumor protein D54 (hD54) (D52-like 2). TD54_HUMAN O43399 5.26 22238 56 56 57 56 54 81 Histone H1.5 (Histone H1a). H15_HUMAN P16401 10.91 22449 41 41 41 46 46 30 Histone H1x. H1X_HUMAN Q92522 10.76 22487 97 97 97 86 85 75 Heat shock 27 kDa protein (HSP 27) (Stress-respons HS27_HUMAN P04792 5.98 22783 65 65 64 66 66 53 43 Proteasome subunit beta type 3 (EC 3.4.25.1) (Prot PSB3_HUMAN P49720 6.14 22949 82 82 82 80 81 68 48 60S ribosomal protein L14 (CAG-ISL 7). RL14_HUMAN P50914 10.94 23159 80 80 80 79 80 67 29 Glutathione S-transferase P (EC 2.5.1.18) (GST cla GTP_HUMAN P09211 5.44 23225 66 66 65 93 95 82 Ran-specific GTPase-activating protein (Ran bindin RANG_HUMAN P43487 5.19 23310 36 36 36 36 38 83 Nuclear protein Hcc-1 (HSPC316) (Proliferation ass HCC1_HUMAN P82979 6.10 23671 28 28 28 28 30 32 40S ribosomal protein S8. RS8_HUMAN P09058 10.32 24074 31 31 31 30 43 52 GTP-binding nuclear protein RAN (TC4) (Ran GTPase) RAN_HUMAN P17080 7.01 24423 73 73 73 73 111 98 59 Hypoxanthine-guanine phosphoribosyltransferase (EC HPRT_HUMAN P00492 6.24 24448 25 25 25 27 27 29 26 Protein-L-isoaspartate(D-aspartate) O-methyltransf PIMT_HUMAN P22061 6.78 24519 63 63 63 63 62 48 Small nuclear ribonucleoprotein associated protein RSMB_HUMAN P14678 11.20 24610 99 99 99 Elongation factor 1-beta (EF-1-beta). EF1B_HUMAN P24534 4.50 24633 64 64 69 64 63 66 26S proteasome non-ATPase regulatory subunit 9 (26 PSD9_HUMAN O00233 6.46 24654 86 86 86 84 74 60S ribosomal protein L10a (CSA-19). R10A_HUMAN P53025 9.94 24700 84 84 85 83 83 61 23 Ubiquitin carboxyl-terminal hydrolase isozyme L1 ( UBL1_HUMAN P09936 5.33 24824 79 79 79 94 92 92 Peroxiredoxin 6 (EC 1.11.1.-) (Antioxidant protein PDX6_HUMAN P30041 6.02 24904 19 19 19 18 18 18 L-3-phosphoserine phosphatase (EC 3.1.3.3) (PSP) ( SERB_HUMAN P78330 5.53 25022 54 54 54 54 53 42 54 FK506-binding protein 3 (EC 5.2.1.8) (Peptidyl-pro FKB3_HUMAN Q00688 9.29 25177 33 33 33 33 32 26 16 Growth factor receptor-bound protein 2 (GRB2 adapt GRB2_HUMAN P29354 5.89 25206 74 74 75 75 72 76 Proteasome subunit beta type 7 precursor (EC 3.4.2 PSB7_HUMAN Q99436 5.61 25295 94 94 94 92 91 84 12 U2 small nuclear ribonucleoprotein B"". RU2B_HUMAN P08579 9.72 25486 26 26 26 26 25 28 28 Platelet-activating factor acetylhydrolase IB beta PA1B_HUMAN Q29459 5.57 25569 111 111 111 110 Splicing factor, arginine/serine-rich 2 (Splicing SFR2_HUMAN Q01130 11.88 25575 75 75 74 74 76 30 Glutathione S-transferase Mu 1 (EC 2.5.1.18) (GSTM GTM1_HUMAN P09488 6.28 25581 91 91 91 89 88 Proteasome subunit alpha type 2 (EC 3.4.25.1) (Pro PSA2_HUMAN P25787 7.12 25767 70 70 70 71 70 56 58 HLA class II histocompatibility antigen, DR alpha 2DRA_HUMAN P01903 4.96 25987 24 24 24 24 23 16 Apoptosis regulator Bcl-X (Bcl-2-like 1 protein). BCLX_HUMAN Q07817 4.86 26049 108 108 108 106 106 93 40 Vacuolar ATP synthase subunit E (EC 3.6.3.14) (V-A VATE_HUMAN P36543 7.70 26145 87 87 87 85 84 74 Translin. TSN_HUMAN Q15631 6.01 26183 92 92 92 90 89 79 2 Isopentenyl-diphosphate delta-isomerase 1 (EC 5.3. IDI1_HUMAN Q13907 5.93 26319 100 100 100 97 97 87 50 Adenylate kinase isoenzyme 2, mitochondrial (EC 2. KAD2_HUMAN P54819 7.85 26347 15 15 15 16 15 17 Proteasome subunit alpha type 5 (EC 3.4.25.1) (Pro PSA5_HUMAN P28066 4.74 26411 35 35 35 35 37 31 46 Triosephosphate isomerase (EC 5.3.1.1) (TIM). TPIS_HUMAN P60174 6.51 26538 1 1 1 1 1 2 40S ribosomal protein S3. RS3_HUMAN P23396 9.68 26688 48 48 48 49 48 60 22 THO complex subunit 4 (Tho4) (Ally of AML-1 and LE THO4_HUMAN Q86V81 11.15 26888 98 21 Chloride intracellular channel protein 1 (Nuclear CLI1_HUMAN O00299 5.09 26923 21 21 22 22 21 27 7 3-hydroxyacyl-CoA dehydrogenase type II (EC 1.1.1. HCD2_HUMAN Q99714 7.65 26923 121 120 121 121 118 41 HLA class II histocompatibility antigen, DRB1-1 be 2B11_HUMAN P04229 7.16 27010 59 59 59 58 58 HLA class II histocompatibility antigen, DRB3-2 be 2B32_HUMAN P01913 6.83 27056 61 61 61 61 60 9 HLA class II histocompatibility antigen, DRB1-11 b 2B1B_HUMAN P20039 6.49 27230 44 44 44 44 41 34 Proteasome activator complex subunit 2 (Proteasome PSE2_HUMAN Q9UL46 5.44 27362 6 7 8 7 6 4 Splicing factor, arginine/serine-rich 7 (Splicing SFR7_HUMAN Q16629 11.83 27367 72 72 72 65 73 Glutathione transferase omega 1 (EC 2.5.1.18) (GST GTO1_HUMAN P78417 6.24 27566 49 49 49 48 57 51 Splicing factor, arginine/serine-rich 1 (pre-mRNA SFR1_HUMAN Q07955 10.37 27613 90 90 90 88 87 78 14-3-3 protein zeta/delta (Protein kinase C inhibi 143Z_HUMAN P29312 4.73 27745 2 2 2 2 2 1 10 14-3-3 protein tau (14-3-3 protein theta) (14-3-3 143T_HUMAN P27348 4.68 27764 3 3 3 3 3 3 Electron transfer flavoprotein beta-subunit (Beta- ETFB_HUMAN P38117 8.25 27844 58 58 58 59 56 Chloride intracellular channel protein 2 (XAP121). CLI2_HUMAN O15247 5.20 27885 95 95 95 95 93 85 Proteasome subunit alpha type 7 (EC 3.4.25.1) (Pro PSA7_HUMAN O14818 8.60 27887 81 81 81 78 79 62 1 14-3-3 protein beta/alpha (Protein kinase C inhibi 143B_HUMAN P31946 4.76 27951 5 5 5 5 5 5 5 14-3-3 protein gamma (Protein kinase C inhibitor p 143G_HUMAN P35214 4.80 28171 16 16 16 15 14 14 Proteasome subunit alpha type 3 (EC 3.4.25.1) (Pro PSA3_HUMAN P25788 5.19 28302 30 30 30 29 28 21 Calcium-dependent protease, small subunit (Calpain CANS_HUMAN P04632 5.05 28316 110 110 110 109 109 U2 small nuclear ribonucleoprotein A' (U2 snRNP-A' RU2A_HUMAN P09661 8.71 28416 37 37 37 38 33 36 32 Acidic leucine-rich nuclear phosphoprotein 32 fami A32A_HUMAN P39687 3.99 28585 22 22 21 23 22 19 31 Phosphoglycerate mutase 1 (EC 5.4.2.1) (EC 5.4.2.4 PMG1_HUMAN P18669 6.75 28673 46 46 46 42 40 37 13 Proteasome activator complex subunit 1 (Proteasome PSE1_HUMAN Q06323 5.78 28723 12 12 12 10 12 13 Proliferating cell nuclear antigen (PCNA) (Cyclin) PCNA_HUMAN P12004 4.57 28769 32 32 32 32 31 33 24 Acidic leucine-rich nuclear phosphoprotein 32 fami A32B_HUMAN Q92688 3.94 28788 14 14 14 14 13 15 44 Eukaryotic translation initiation factor 3 subunit IF31_HUMAN O75822 4.72 29062 53 53 53 53 45 38 20 14-3-3 protein epsilon (Mitochondrial import stimu 143E_HUMAN P42655 4.63 29174 7 8 6 9 9 6 6 60S ribosomal protein L7. RL7_HUMAN P18124 10.66 29226 38 38 38 37 34 39 39 40S ribosomal protein S4, X isoform (Single copy a RS4_HUMAN P12750 10.16 29467 69 69 68 69 67 55 34 Proteasome subunit alpha type 4 (EC 3.4.25.1) (Pro PSA4_HUMAN P25789 7.58 29484 55 55 56 55 55 44 17

pI= 7.1 +/- 2.2 MW= 28.4 +/- 5.3 kDa

10000 20000 30000 40000 50000 60000 0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 pI MW

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Influence of the threshold

20 40 60 80 100 120 140 0.0% 1.0% 2.0% 3.0% 4.0% 5.0% 6.0% 7.0% 8.0% 9.0% 10.0% threshold for data extraction

Taille des fichier (Mo) Protein identifiées par Mascot (absolute number) Faux positifs (absolute number) Litigieux (1 seul peptide ou 2 peptides dont 1<au threshold mascot) (abolute number) proteines clairement identifiée (absolute number)) Faux positifs (%) Litigieux (1 seul peptide ou 2 peptides dont 1<au threshold mascot) (%) proteines clairement identifiée (%)

Threshold 0.05% 0.10% 0.20% 0.50% 1.00% 2.00% 5.00% 10.00% Taille des fichier (Mo) 5.063 4.993 4.800 4.284 3.627 2.610 1.021 0.264 Protein identifiées par Mascot (absolute number) 125 124 125 126 120 103 59 14 Faux positifs (absolute number) 13 12 13 15 13 12 3 Litigieux (1 seul peptide ou 2 peptides dont 1<au threshold mascot) (abolute number) 32 30 29 29 24 19 26 10 proteines clairement identifiée (absolute number)) 80 82 83 82 83 72 30 4 Faux positifs (%) 10.4 9.7 10.4 11.9 10.8 11.7 5.1 0.0 Litigieux (1 seul peptide ou 2 peptides dont 1<au threshold mascot) (%) 25.6 24.2 23.2 23.0 20.0 18.4 44.1 71.4 proteines clairement identifiée (%) 64.0 66.1 66.4 65.1 69.2 69.9 50.8 28.6

Influence of the threshold

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Extraction of the data from the acquired files

Treatment of the data for protein identification: MASCOT

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Pappin, DJC, Hojrup, P and Bleasby, AJ, Rapid identification of proteins by peptide-mass fingerprinting. Curr. Biol., 3(6) 327-32 (1993).

MASCOT : Mowse scoring algorythm

The first stage of a Mowse search is to compare the calculated peptide masses for each entry in the sequence database with the set of experimental data. Each calculated value which falls within a given mass tolerance of an experimental value counts as a match. A molecular weight range for the intact protein can be used as a pre-filter. Simpliest scorring method: just counting the number of matching peptides. Technique used in PeptIdent, PepSea, PeptideSearch Mowse uses empirically determined factors to assign a statistical weight to each individual peptide match.

Definitions copy from http://www.matrixscience.com/

The matrix of weighting factors is calculated during the database build stage, as follows: A frequency factor matrix, F, is created, in which each row represents an interval of 100 Da in peptide mass, and each column an interval of 10 kDa in intact protein mass. As each sequence entry is processed, the appropriate matrix elements fi,j are incremented so as to accumulate statistics on the size distribution of peptide masses as a function of protein mass. The elements of F are then normalised by dividing the elements of each 10 kDa column by the largest value in that column to give the Mowse factor matrix M: After searching the experimental mass values against a calculated peptide mass database, the score for each entry is calculated according to:

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The total score is the absolute probability that the observed match is a random event. Reporting probabilities directly can be confusing. Partly because they encompass a very wide range of magnitudes, and also because a "high" score is a "low" probability, which can be ambiguous. For this reason, we report scores as -10*LOG10(P), where P is the absolute

• probability. A probability of 10-20 thus becomes a score of 200.

Probability Based Mowse

Mascot incorporates a probability based implementation of the Mowse algorithm. By casting the Mowse score into a probabilistic framework, we gain a number of additional benefits:

• 1. A simple rule can be used to judge whether a result is significant or not.
• 2. Different types of matching (peptide masses and fragment ions) can be

combined in a single search.

• 3. Scores from different searches and on different databases can be compared.
• 4. Search parameters can be optimised more readily by iteration.

Matches using mass values (either peptide masses or MS/MS fragment ion masses) are always handled on a probabilistic basis.

Significance Level

Given an absolute probability that a match is random, and knowing the size of the sequence database being searched, it becomes possible to provide an objective measure of the significance of a result. A commonly accepted threshold is that an event is significant if it would be expected to occur at random with a frequency of less than 5%. The master results page for typical peptide mass fingerprint search (open in new window) reports that "Scores greater than 69 are significant (p<0.05)". The histogram of the score distribution looks like this:

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Expectation Values

Each protein score in a peptide mass fingerprint, and each ions score in an MS/MS search, is accompanied by an expectation value. This is the number of matches with equal or better scores that are expected to occur by chance alone. It is directly equivalent to the E-value in a Blast search result. For a score that is exactly on the default significance threshold, (p<0.05), the expectation value is also 0.05. Increase the score by 10 and the expectation value drops to 0.005. The lower the expectation value, the more significant the score. Mass Tolerances The score in a peptide mass fingerprint is usually inversely related to the mass tolerance. This is not always the case for an MS/MS ions search, where increasing the peptide mass tolerance may have little effect on the score. This is because most of the discrimination comes from the MS/MS fragment ion matches. Opening up the peptide mass tolerance means that Mascot has to test many more peptides, (and so the search takes longer!), but the major contributions to the final score, the MS/MS fragment ion matches, are unchanged. In fact, if the peptide mass tolerance is set too tightly, in an effort to improve discrimination, one or more of the peptide matches may be lost, which will dramatically reduce the overall score.

Limitations

Like any statistical approach, the Probability Based Mowse algorithm depends on assumptions and models. One of these assumptions is that the entries in the sequence databases are random sequences. This is not always a good assumption. Some of the most glaring examples involve extended repeats, such as AAC62527, porcine submaxillary apomucin. Although the molecular weight of this protein is 1.2 MDa, over 80% of the sequence is composed of an identical 7 kDa repeat. It is difficult to know how to treat such cases.

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If a single experimental peptide mass is allowed to match to multiple calculated masses, then a single experimental mass which matches within a repeat will give a huge and meaningless score. But, if duplicate matches are not permitted, it will be virtually impossible to get a match to such a protein because the number of measurable mass values is too small to give a statistically significant score. Another assumption is that the experimental measurements are independent

• determinations. This will not be true if the data include multiple mass values for the

same peptide, even if these are from ions with different charge states in an electrospray LC-MS run. Good peak detection and thresholding (in both mass and time domains for LC-MS) are essential for any scoring algorithm to give meaningful results.

Sequence Query Scoring

Amino acid sequence or composition information, if present, is treated as a rigorous filter on the candidate sequences. Ambiguous sequence or composition data can be used (in a manner similar to a regular expression search in computing) but it still functions as a filter, not a probabilistic match of the type found in a Blast or Fasta search.

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Protein description SP name Uniprot entry pI MW Rank (first analysis) Score peptides doublons % recov Hypothetical protein. Q8WYX2 Q8WYX2 10.38 14140 8 156 3 1 6 Basigin precursor Leukocyte activation antigen M6 Collagenase stimulatory factor Extr BASI_HUMAN P35613 5.31 27222 13 98 2 7 Nuclease sensitive element binding protein 1 Y box binding protein-1 Y-box transcriptio YB1_HUMAN P16991 9.87 35924 6 243 4 7 L-lactate dehydrogenase B chain LDHB_HUMAN P07195 5.72 36507.3 Actin, cytoplasmic 1 Beta-actin (egalement ACTB_HUMAN) ACTB_HUMAN P02570 5.29 41606 14 91 2 7 Nucleosome assembly protein 1-like 4 Nucleosome a NPL4_HUMAN Q99733 4.60 42823 Eukaryotic initiation factor 4A-I eIF4A-I eIF-4 IF41_HUMAN P04765 5.32 46154 12 110 2 7 Protein disulfide isomerase A6 precursor EC 5.3.4 PDA6_HUMAN Q15084 4.95 46171 Lupus La protein Sjogren syndrome type B antigen LA_HUMAN P05455 6.68 46837 2 635 10 24 Alpha enolase EC 4.2.1.11 2-phospho-D-glycerate ENOA_HUMAN P06733 6.99 47038 11 133 5 13 26S protease regulatory subunit 4 P26s4. PRS4_HUMAN Q03527 5.87 49185 Heterogeneous nuclear ribonucleoprotein H hnRNP H ROH1_HUMAN P31943 5.89 49229 19 52 1 2 Tubulin alpha-4 chain Alpha-tubulin 4. TBA4_HUMAN P05215 4.95 49924 5 288 8 2 18 RuvB-like 1 EC 3.6.1.- 49-kDa TATA box-binding RUV1_HUMAN Q9Y265 6.02 50228 Tubulin beta-4 chain Tubulin beta-III TBB4_HUMAN Q13509 4.83 50433 17 70 3 5 U1 small nuclear ribonucleoprotein 70 kDa U1 snRN RU17_HUMAN P08621 9.94 51557 3 597 10 24 ATP synthase beta chain, mitochondrial precursor ATPB_HUMAN P06576 5.00 51769 15 73 3 6 Serine hydroxymethyltransferase, mitochondrial pre GLYM_HUMAN P34897 8.11 52560 9 153 3 6 Cytosol aminopeptidase EC 3.4.11.1 Leucine amin AMPL_HUMAN P28838 6.29 52640 XAGL protein Fibroblast growth factor 2-interacting factor Q9Y4J7 Q9Y4J7 5.72 56770 18 56 1 3 T-complex protein 1, delta subunit TCP-1-delta ou Q96C51 TCPD_HUMAN P50991 7.52 57839 7 183 5 11 Importin alpha-2 subunit Karyopherin alpha-2 subunit SRP1-alpha RAG cohort protein 1 IMA2_HUMAN P52292 5.25 57862 16 73 2 5 T-complex protein 1, zeta subunit TCP-1-zeta CC TCPZ_HUMAN P40227 6.24 58024 22 38 1 1 Tyrosyl-tRNA synthetase EC 6.1.1.1 Tyrosyl--tRNA ligase TyrRS SYY_HUMAN P54577 6.61 59143 24 36 1 2 Glucose-6-phosphate isomerase EC 5.3.1.9 GPI G6PI_HUMAN P06744 8.42 63147 4 412 12 36 Probable RNA-dependent helicase p68 DEAD-box prot DDX5_HUMAN P17844 9.06 69148 10 141 4 9

Histidyl-tRNA synthetase

SYH_HUMAN P12081 5.72 57410.51 1 1492 33 6 54

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Protein description SP name Uniprot entry pI MW Rank (first analysis) Score peptides doublons % recov Rank (first analysis) Score peptides doublons % recov Hypothetical protein. Q8WYX2 Q8WYX2 10.38 14140 8 156 3 1 6 6 161 4 1 8 Basigin precursor Leukocyte activation antigen M6 Collagenase stimulatory factor Extr BASI_HUMAN P35613 5.31 27222 13 98 2 7 13 95 2 6 Nuclease sensitive element binding protein 1 Y box binding protein-1 Y-box transcriptio YB1_HUMAN P16991 9.87 35924 6 243 4 7 11 135 2 4 L-lactate dehydrogenase B chain LDHB_HUMAN P07195 5.72 36507.3 24 37 1 1 Actin, cytoplasmic 1 Beta-actin (egalement ACTB_HUMAN) ACTB_HUMAN P02570 5.29 41606 14 91 2 7 9 148 4 13 Nucleosome assembly protein 1-like 4 Nucleosome a NPL4_HUMAN Q99733 4.60 42823 15 76 1 3 Eukaryotic initiation factor 4A-I eIF4A-I eIF-4 IF41_HUMAN P04765 5.32 46154 12 110 2 7 Protein disulfide isomerase A6 precursor EC 5.3.4 PDA6_HUMAN Q15084 4.95 46171 16 69 2 4 Lupus La protein Sjogren syndrome type B antigen LA_HUMAN P05455 6.68 46837 2 635 10 24 3 486 9 23 Alpha enolase EC 4.2.1.11 2-phospho-D-glycerate ENOA_HUMAN P06733 6.99 47038 11 133 5 13 7 155 5 12 26S protease regulatory subunit 4 P26s4. PRS4_HUMAN Q03527 5.87 49185 19 48 3 1 2 Heterogeneous nuclear ribonucleoprotein H hnRNP H ROH1_HUMAN P31943 5.89 49229 19 52 1 2 21 45 1 2 Tubulin alpha-4 chain Alpha-tubulin 4. TBA4_HUMAN P05215 4.95 49924 5 288 8 2 18 8 154 4 14 RuvB-like 1 EC 3.6.1.- 49-kDa TATA box-binding RUV1_HUMAN Q9Y265 6.02 50228 17 62 2 5 Tubulin beta-4 chain Tubulin beta-III TBB4_HUMAN Q13509 4.83 50433 17 70 3 5 20 456 2 4 U1 small nuclear ribonucleoprotein 70 kDa U1 snRN RU17_HUMAN P08621 9.94 51557 3 597 10 24 2 591 10 27 ATP synthase beta chain, mitochondrial precursor ATPB_HUMAN P06576 5.00 51769 15 73 3 6 10 145 4 12 Serine hydroxymethyltransferase, mitochondrial pre GLYM_HUMAN P34897 8.11 52560 9 153 3 6 12 116 2 3 Cytosol aminopeptidase EC 3.4.11.1 Leucine amin AMPL_HUMAN P28838 6.29 52640 5 166 4 6 XAGL protein Fibroblast growth factor 2-interacting factor Q9Y4J7 Q9Y4J7 5.72 56770 18 56 1 3 T-complex protein 1, delta subunit TCP-1-delta ou Q96C51 TCPD_HUMAN P50991 7.52 57839 7 183 5 11 Importin alpha-2 subunit Karyopherin alpha-2 subunit SRP1-alpha RAG cohort protein 1 IMA2_HUMAN P52292 5.25 57862 16 73 2 5 T-complex protein 1, zeta subunit TCP-1-zeta CC TCPZ_HUMAN P40227 6.24 58024 22 38 1 1 Tyrosyl-tRNA synthetase EC 6.1.1.1 Tyrosyl--tRNA ligase TyrRS SYY_HUMAN P54577 6.61 59143 24 36 1 2 Glucose-6-phosphate isomerase EC 5.3.1.9 GPI G6PI_HUMAN P06744 8.42 63147 4 412 12 36 4 278 9 25 Probable RNA-dependent helicase p68 DEAD-box prot DDX5_HUMAN P17844 9.06 69148 10 141 4 9 14 77 2 2

Histidyl-tRNA synthetase

SYH_HUMAN P12081 5.72 57410.51 1 1492 33 6 54 1 1066 22 4 42

Rank of identification or Mascot score

Protein description SP name Uniprot entry pI MW Rank (first analysis) Score peptides doublons % recov Rank (first analysis) Score peptides doublons % recov Hypothetical protein. Q8WYX2 Q8WYX2 10.38 14140 8 156 3 1 6 6 161 4 1 8 Basigin precursor Leukocyte activation antigen M6 Collagenase stimulatory factor Extr BASI_HUMAN P35613 5.31 27222 13 98 2 7 13 95 2 6 Nuclease sensitive element binding protein 1 Y box binding protein-1 Y-box transcriptio YB1_HUMAN P16991 9.87 35924 6 243 4 7 11 135 2 4 L-lactate dehydrogenase B chain LDHB_HUMAN P07195 5.72 36507.3 24 37 1 1 Actin, cytoplasmic 1 Beta-actin (egalement ACTB_HUMAN) ACTB_HUMAN P02570 5.29 41606 14 91 2 7 9 148 4 13 Nucleosome assembly protein 1-like 4 Nucleosome a NPL4_HUMAN Q99733 4.60 42823 15 76 1 3 Eukaryotic initiation factor 4A-I eIF4A-I eIF-4 IF41_HUMAN P04765 5.32 46154 12 110 2 7 Protein disulfide isomerase A6 precursor EC 5.3.4 PDA6_HUMAN Q15084 4.95 46171 16 69 2 4 Lupus La protein Sjogren syndrome type B antigen LA_HUMAN P05455 6.68 46837 2 635 10 24 3 486 9 23 Alpha enolase EC 4.2.1.11 2-phospho-D-glycerate ENOA_HUMAN P06733 6.99 47038 11 133 5 13 7 155 5 12 26S protease regulatory subunit 4 P26s4. PRS4_HUMAN Q03527 5.87 49185 19 48 3 1 2 Heterogeneous nuclear ribonucleoprotein H hnRNP H ROH1_HUMAN P31943 5.89 49229 19 52 1 2 21 45 1 2 Tubulin alpha-4 chain Alpha-tubulin 4. TBA4_HUMAN P05215 4.95 49924 5 288 8 2 18 8 154 4 14 RuvB-like 1 EC 3.6.1.- 49-kDa TATA box-binding RUV1_HUMAN Q9Y265 6.02 50228 17 62 2 5 Tubulin beta-4 chain Tubulin beta-III TBB4_HUMAN Q13509 4.83 50433 17 70 3 5 20 456 2 4 U1 small nuclear ribonucleoprotein 70 kDa U1 snRN RU17_HUMAN P08621 9.94 51557 3 597 10 24 2 591 10 27 ATP synthase beta chain, mitochondrial precursor ATPB_HUMAN P06576 5.00 51769 15 73 3 6 10 145 4 12 Serine hydroxymethyltransferase, mitochondrial pre GLYM_HUMAN P34897 8.11 52560 9 153 3 6 12 116 2 3 Cytosol aminopeptidase EC 3.4.11.1 Leucine amin AMPL_HUMAN P28838 6.29 52640 5 166 4 6 XAGL protein Fibroblast growth factor 2-interacting factor Q9Y4J7 Q9Y4J7 5.72 56770 18 56 1 3

Histidyl-tRNA synthetase

SYH_HUMAN P12081 5.72 57410.51 1 1492 33 6 54 1 1066 22 4 42 T-complex protein 1, delta subunit TCP-1-delta ou Q96C51 TCPD_HUMAN P50991 7.52 57839 7 183 5 11 Importin alpha-2 subunit Karyopherin alpha-2 subunit SRP1-alpha RAG cohort protein 1 IMA2_HUMAN P52292 5.25 57862 16 73 2 5 T-complex protein 1, zeta subunit TCP-1-zeta CC TCPZ_HUMAN P40227 6.24 58024 22 38 1 1 Tyrosyl-tRNA synthetase EC 6.1.1.1 Tyrosyl--tRNA ligase TyrRS SYY_HUMAN P54577 6.61 59143 24 36 1 2 Glucose-6-phosphate isomerase EC 5.3.1.9 GPI G6PI_HUMAN P06744 8.42 63147 4 412 12 36 4 278 9 25 Probable RNA-dependent helicase p68 DEAD-box prot DDX5_HUMAN P17844 9.06 69148 10 141 4 9 14 77 2 2

% of sequence recovery

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Protein description SP name Uniprot entry pI MW Rank (first analysis) Score peptides doublons % recov Rank (first analysis) Score peptides doublons % recov Hypothetical protein. Q8WYX2 Q8WYX2 10.38 14140 8 156 3 1 6 6 161 4 1 8 Basigin precursor Leukocyte activation antigen M6 Collagenase stimulatory factor Extr BASI_HUMAN P35613 5.31 27222 13 98 2 7 13 95 2 6 Nuclease sensitive element binding protein 1 Y box binding protein-1 Y-box transcriptio YB1_HUMAN P16991 9.87 35924 6 243 4 7 11 135 2 4 L-lactate dehydrogenase B chain LDHB_HUMAN P07195 5.72 36507.3 24 37 1 1 Actin, cytoplasmic 1 Beta-actin (egalement ACTB_HUMAN) ACTB_HUMAN P02570 5.29 41606 14 91 2 7 9 148 4 13 Nucleosome assembly protein 1-like 4 Nucleosome a NPL4_HUMAN Q99733 4.60 42823 15 76 1 3 Eukaryotic initiation factor 4A-I eIF4A-I eIF-4 IF41_HUMAN P04765 5.32 46154 12 110 2 7 Protein disulfide isomerase A6 precursor EC 5.3.4 PDA6_HUMAN Q15084 4.95 46171 16 69 2 4 Lupus La protein Sjogren syndrome type B antigen LA_HUMAN P05455 6.68 46837 2 635 10 24 3 486 9 23 Alpha enolase EC 4.2.1.11 2-phospho-D-glycerate ENOA_HUMAN P06733 6.99 47038 11 133 5 13 7 155 5 12 26S protease regulatory subunit 4 P26s4. PRS4_HUMAN Q03527 5.87 49185 19 48 3 1 2 Heterogeneous nuclear ribonucleoprotein H hnRNP H ROH1_HUMAN P31943 5.89 49229 19 52 1 2 21 45 1 2 Tubulin alpha-4 chain Alpha-tubulin 4. TBA4_HUMAN P05215 4.95 49924 5 288 8 2 18 8 154 4 14 RuvB-like 1 EC 3.6.1.- 49-kDa TATA box-binding RUV1_HUMAN Q9Y265 6.02 50228 17 62 2 5 Tubulin beta-4 chain Tubulin beta-III TBB4_HUMAN Q13509 4.83 50433 17 70 3 5 20 456 2 4 U1 small nuclear ribonucleoprotein 70 kDa U1 snRN RU17_HUMAN P08621 9.94 51557 3 597 10 24 2 591 10 27 ATP synthase beta chain, mitochondrial precursor ATPB_HUMAN P06576 5.00 51769 15 73 3 6 10 145 4 12 Serine hydroxymethyltransferase, mitochondrial pre GLYM_HUMAN P34897 8.11 52560 9 153 3 6 12 116 2 3 Cytosol aminopeptidase EC 3.4.11.1 Leucine amin AMPL_HUMAN P28838 6.29 52640 5 166 4 6 XAGL protein Fibroblast growth factor 2-interacting factor Q9Y4J7 Q9Y4J7 5.72 56770 18 56 1 3

Histidyl-tRNA synthetase

SYH_HUMAN P12081 5.72 57410.51 1 1492 33 6 54 1 1066 22 4 42 T-complex protein 1, delta subunit TCP-1-delta ou Q96C51 TCPD_HUMAN P50991 7.52 57839 7 183 5 11 Importin alpha-2 subunit Karyopherin alpha-2 subunit SRP1-alpha RAG cohort protein 1 IMA2_HUMAN P52292 5.25 57862 16 73 2 5 T-complex protein 1, zeta subunit TCP-1-zeta CC TCPZ_HUMAN P40227 6.24 58024 22 38 1 1 Tyrosyl-tRNA synthetase EC 6.1.1.1 Tyrosyl--tRNA ligase TyrRS SYY_HUMAN P54577 6.61 59143 24 36 1 2 Glucose-6-phosphate isomerase EC 5.3.1.9 GPI G6PI_HUMAN P06744 8.42 63147 4 412 12 36 4 278 9 25 Probable RNA-dependent helicase p68 DEAD-box prot DDX5_HUMAN P17844 9.06 69148 10 141 4 9 14 77 2 2

Identification variability

SWISS-PROT name Uniprot entry pI MW LR 90' 7020 Score peptide s doublo ns % recov LR 90' 7049 Score peptide s doublo ns % recov LR 90' 7059 Score peptide s doublo ns % recov SFR6_HUMAN Q13247 11.42 39587 26 49 1 2 ST13_HUMAN P50502 5.18 41332 15 77 1 3 24 72 2 ACTB_HUMAN P02570 5.29 41606 12 258 7 12 181 4 10 FEN1_HUMAN P39748 8.8 42593 18 53 2 4 20 77 3 7 NPL4_HUMAN Q99733 4.60 42823 23 77 2 Q8NC51 Q8NC51 8.66 44938 25 65 1 19 83 2 4 EFTU_HUMAN P49411 6.31 45045 30 50 1 ROF_HUMAN P52597 5.38 45672 21 41 1 3 17 177 4 22 69 2 5 IF41_HUMAN P04765 5.32 46154 27 62 2 27 49 2 6 PDA6_HUMAN Q15084 4.95 46171 10 142 3 9 10 342 7 1 8 292 6 1 15 LA_HUMAN P05455 6.68 46837 15 204 6 1 ENOA_HUMAN P06733 6.99 47038 4 362 9 29 7 400 11 1 6 358 10 1 22 Q9NR72 Q9NR72 4.98 48154 28 42 1 2 Q8WU90 Q8WU90 5.22 48602 25 53 1 3 Q8WU90 Q8WU90 5.22 48602 32 46 1 HE47_HUMAN Q13838 5.44 48991 22 38 2 3 ROH1_HUMAN P31943 5.89 49229 16 199 4 23 68 3 6 TBB1_HUMAN P07437 4.75 49759 2 505 11 1 27 2 605 13 1 3 482 14 2 33 TBB2_HUMAN P05217 4.79 49831 3 494 12 2 27 4 563 13 2 2 575 16 3 37 EF1G_HUMAN P26641 6.25 50119 16 62 1 2 28 62 2 EF11_HUMAN P04720 9.10 50141 9 166 3 10 11 284 7 1 14 153 4 1 6 TBA1_HUMAN P05209 4.94 50152 12 108 4 12 18 171 7 1 9 251 8 1 19 RUV1_HUMAN Q9Y265 6.02 50228 26 64 2 30 39 1 2 ROK_HUMAN Q07244 5.39 50976 11 116 5 13 8 361 9 10 191 5 9 ATPB_HUMAN P06576 5.00 51769 5 287 7 19 9 356 8 7 355 9 24 GLYM_HUMAN P34897 8.11 52560 13 217 5 16 133 5 7 PDA3_HUMAN P30101 5.61 54265 8 207 5 13 5 435 13 1 11 183 6 11 ATPA_HUMAN P25705 8.28 55209 7 221 5 10 6 412 10 4 420 10 21 SERA_HUMAN O43175 6.29 56651 14 93 2 4 22 103 3 15 152 4 7 Q9Y4J7 Q9Y4J7 5.72 56770 31 47 1 PTB_HUMAN P26599 9.22 57221 17 54 2 5 TCPB_HUMAN P78371 6.01 57488 13 93 2 3 19 147 3 21 69 2 4 KPY1_HUMAN P14618 7.95 57806 20 45 2 2 14 211 6 13 159 5 11 CH60_HUMAN P10809 5.24 57963 1 745 19 4 33 1 1441 38 10 1 1320 34 5 49 SYY_HUMAN P54577 6.61 59143 24 58 2 4 TCPQ_HUMAN P50990 5.42 59621 21 111 2 TCPG_HUMAN P49368 6.10 60403 19 45 2 2 18 92 3 5 IEFS_HUMAN P31948 6.40 62639 20 134 4 17 98 4 6 PLSL_HUMAN P13796 5.20 70289 6 271 6 11 3 568 14 1 5 416 12 1 17

Identification variability (3 analyses)

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• u ST13-like tumor suppressor.

• u Q9BVP6

• u CTCL tumor antigen Cervical cancer suppressor 3.Q8IUB0

Influence of the LC gradient length Sequential m/z range analyses: cumulative analysis

Protein description pI MW Uniprot entry SP name NR 300- 410 6909 peptides NR 400- 510 6910 peptides NR 500- 610 6912 peptides NR 600- 710 6913 peptides NR 700- 810 6914 peptides NR 800- 910 6915 peptides NR 900- 1010 6916 peptides NR 1200- 1310 6919 peptides cumul NRs peptides doublon s % recov 60 kDa heat shock protein, mitochondrial precursor 5.24 57963 P10809 CH60_HUMAN 285 7 290 7 151 5 182 4 80 1 162 3 101 1 1105 29 5 42 Alpha enolase EC 4.2.1.11 2-phospho-D-glycerate 6.99 47038 P06733 ENOA_HUMAN 79 3 107 3 90 2 109 3 352 11 1 31 Tubulin beta-2 chain. 4.79 49831 P05217 TBB2_HUMAN 211 4 56 2 317 8 1 16 L-plastin Lymphocyte cytosolic protein 1 LCP-1 5.20 70289 P13796 PLSL_HUMAN 69 3 124 3 47 1 42 1 44 1 316 11 2 16 Tubulin beta-1 chain et TBB5_HUMAN 4.75 49759 P07437 TBB1_HUMAN 51 2 178 4 249 7 17 Heterogeneous nuclear ribonucleoprotein K hnRNP K 5.39 50976 Q07244 ROK_HUMAN 45 1 96 3 67 1 231 6 13 Protein disulfide isomerase A3 precursor EC 5.3.4 5.61 54265 P30101 PDA3_HUMAN 113 3 74 1 211 5 14 Hypothetical protein FLJ90489. 8.66 44938 Q8NC51 Q8NC51 103 2 40 1 176 4 11 Actin, cytoplasmic 1 Beta-actin (egalement ACTB_HUMAN) 5.29 41606 P02570 ACTB_HUMAN 56 2 42 1 51 1 165 6 1 19 Lupus La protein Sjogren syndrome type B antigen 6.68 46837 P05455 LA_HUMAN 106 2 58 1 164 3 9 Pyruvate kinase, M1 isozyme EC 2.7.1.40 Pyruvat et KPY2_HUMAN 7.95 57806 P14618 KPY1_HUMAN 121 4 121 4 5 Elongation factor Tu, mitochondrial precursor EF-Tu P43 6.31 45045 P49411 EFTU_HUMAN 49 1 43 1 119 3 6 Tubulin alpha-1 chain Alpha-tubulin 1. et TBA6_HUMAN 4.94 50152 P05209 TBA1_HUMAN 74 2 53 1 118 5 1 132 T-complex protein 1, beta subunit TCP-1-beta CC 6.01 57488 P78371 TCPB_HUMAN 70 1 116 3 5 Elongation factor 1-alpha 1 EF-1-alpha-1 Elonga

• u CTCL tumor antigen Cervical cancer suppressor

3.Q8IUB0 9.10 50141 P04720 EF11_HUMAN 67 1 116 3 5 Protein disulfide isomerase A6 precursor EC 5.3.4 4.95 46171 Q15084 PDA6_HUMAN 40 1 45 1 107 3 6 ATP synthase beta chain, mitochondrial precursor 5.00 51769 P06576 ATPB_HUMAN 74 1 107 2 4 Serine hydroxymethyltransferase, mitochondrial pre 8.11 52560 P34897 GLYM_HUMAN 36 1 102 3 5 Nucleosome assembly protein 1-like 4 Nucleosome a 4.60 42823 Q99733 NPL4_HUMAN 40 1 40 1 80 2 6 T-complex protein 1, theta subunit TCP-1-theta 5.42 59621 P50990 TCPQ_HUMAN 66 1 80 2 4 Putative GTP-binding protein PTD004 PRO2455. 7.64 44744 Q9NTK5 PTD4_HUMAN 65 1 79 2 6 Nucleosome assembly protein 1-like 1 NAP-1 relate 4.36 45374 P55209 NPL1_HUMAN 73 2 7 Elongation factor 1-gamma EF-1-gamma eEF-1B gam 6.25 50119 P26641 EF1G_HUMAN 42 1 68 2 5 U1 small nuclear ribonucleoprotein 70 kDa U1 snRN 9.94 51557 P08621 RU17_HUMAN 68 1 66 1 2 Probable ATP-dependent RNA helicase p47 HLA-B ass

• u Q9BVP6

5.44 48991 Q13838 HE47_HUMAN 62 1 62 1 2 Heterogeneous nuclear ribonucleoprotein F hnRNP F 5.38 45672 P52597 ROF_HUMAN 48 2 6 T-complex protein 1, delta subunit TCP-1-delta ou Q96C51 7.52 57839 P50991 TCPD_HUMAN 40 1 40 1 1 T-complex protein 1, eta subunit TCP-1-eta CCT- 7.55 59367 Q99832 TCPH_HUMAN 40 1 40 1 2 60S ribosomal protein L4 L1. 11.07 47697 P36578 RL4_HUMAN 36 2 36 2 3 Cervical SH3P7 Mucin-associated protein. 4.98 48154 Q9NR72 Q9NR72 36 1 36 1 2

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http://www.u.arizona.edu/~yingying/massSpec/tutorial.htm