ELECTROPHORESIS Supporting Information for Electrophoresis DOI 10.1002/elps.200600696 Jinfeng Liu, Yun Cai, Jinglan Wang, Qi Zhou, Bing Yang, Zhuang Lu, Liyan Jiao, Dongyang Zhang, Shaohui Sui, Ying Jiang, Wantao Ying and Xiaohong Qian Phosphoproteome profile of human liver Chang’s cell based on 2-DE with fluorescence staining and MALDI-TOF/TOF MS ª 2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.electrophoresis-journal.com Figures S1:The distribution of the protein spots selected for MALDI-TOF/TOF-MS identification. Table S1: Cellular location and functional classification of phosphoproteins in Chang’s cells Protein name Protein ID Subcellular Protein function location HS9B_ Heat shock P08238 cytoplasm positive regulation of nitric oxide protein HSP90-beta biosynthesis；nitric-oxide synthase (HSP 84) regulator activity；binding G3BP_ Ras-GTPase- Q13283 nucleus transport； Ras protein signal activating protein cytoplasm transduction；catalytic activity binding protein 1 VIME_ Vimentin P08670 cytoplasm structural constituent of cytoskeleton cytoskeleton protein binding cell motility RLA0_ 60S acidic P05388 ribosome binding；protein ribosomal proteinP0 biosynthesis；translational elongation；structural constituent of ribosome HS27_Heat shock 27 P04792 cytoplasm protein binding；regulation of kDa protein(HSP 27) cytoskeleton translational initiation；protein cell surface folding；anti-apoptosis；cell motility；response to unfolded protein NDKA_Nucleoside P15531 nucleus binding; biosynthesis; catalytic diphosphatekinase A cytoplasm activity; cell cycle; regulation of apoptosis EF1B_Elongation P24534 eukaryotic translation elongation factor factor 1-beta translation activity；protein binding；protein elongation factor biosynthesis；translational 1 complex elongation PSA5_ Proteasome P28066 proteasome core threonine endopeptidase activity subunit alpha type 5 complex ubiquitin-dependent protein catabolism TCTP_Translationally P13693 cytoplasm;multiv calcium ion controlletumor protein esicular body; transport、homeostasis、binding； tubulin anti-apoptosis；regulation of apoptosis RLA2_ 60S acidic P05387 ribosome binding；protein ribosomal proteinP2 biosynthesis；translational elongation；structural constituent of ribosome Z255_ Zinc finger Q9UID9 nucleus binding；transcription protein 255 Table S2: The list of proteins identified by MALDI-TOF/TOF-MS in Chang’s cells Spot Protein Name Accession Position Number 1 NCL protein IPI00183526.4 CDNA FLJ45706 fis, clone FEBRA2028457, highly similar to Nucleolin IPI00444262.1 Hypothetical protein FLJ34423 IPI00604517.1 Hypothetical protein NCL IPI00604620.1 2 Nucleolin IPI00217506.4 3 Endoplasmin precursor IPI00027230.3 4 GARS protein IPI00465260.1 5 Splice Isoform A of Ras-GTPase-activating protein binding p IPI00009057.1 Ras-GTPase-activating prot IPI00012442.1 Splice Isoform B of Ras-GTPase-activating protein binding protein 2 IPI00179890.2 6 CDNA FLJ26493 fis, clone KDN06317, highly similar to Ras-GTPase- activating protein bin IPI00442863.1 7 Cytoskeleton associated protein 2 IPI00071824.2 Clusterin precursor IPI00291262.3 Clusterin isoform 1 IPI00400826.1 8 Similar to Lamin A/C IPI00514320.2 Lamin A\/C IPI00514817.3 Chaperonin Containing TCP1, subunit 3 isoform C IPI00552715.1 T-complex protein 1, gamma subunit IPI00553185.2 9 Hypothetical protein DKFZp761K0511 IPI00334775.3 10 Splice Isoform 1 of Mitochondrial inner membrane protein IPI00009960.6 Splice Isoform 3 of Mitochondrial inner membrane protein IPI00470829.2 Splice Isoform 2 of Mitochondrial inner membrane protein IPI00554469.1 11 Heat shock protein 90Ad IPI00555957.1 12 Heat shock protein HSP 90-alpha 2 IPI00382470.2 Tax_Id IPI00414676.5 Heat shock protein 90Bc IPI00555614.1 Heat shock protein HSP 90-alp IPI00604607.1 13 Splice Isoform A of Lami IPI00021405.3 Splice Isoform C of Lamin A/C IPI00216952.1 Splice Isoform ADelta10 of Lamin A/C IPI00216953.1 Bifunctional purine biosynthesis protein PURH IPI00289499.3 14 Nucleosome assembly protein 1-like IPI00023860.1 Synapse associated protein 1 IPI00059242.3 15 Splice Isoform 1 of Calumenin precursor IPI00014537.1 Splice Isoform 2 of Calumenin precursor IPI00045396.1 16 Splice Isoform 1 of Heterogeneous nuclear ribonucleoprotein K IPI00216049.1 Splice Isoform 2 of Heterogeneous nucl IPI00216746.1 PREDICTED: similar to heterogeneous nuclear ribonucleoprotein K IPI00398052.2 Heterogeneous nuclear ribonucleoprotein K IPI00514561.1 17 Vimentin IPI00418471.5 Ribonuclease/angiogenin inhibitor IPI00465081.2 Hypothetical protein FLJ36605 IPI00477807.3 Placental ribonuclease inhibitor IPI00550069.2 Ribonuclease/angiogenin inhibitor IPI00550654.2 Placental ribonuclease inhibitor IPI00550874.1 Vimentin IPI00552689.1 18 Keratin, type I cytoskeletal 17 IPI00450768.4 19 78 kDa glucose-regulated protein precursor IPI00003362.1 20 T-complex protein 1, epsilon subunit IPI00010720.1 21 Protein disulfide-isomerase A3 precursor IPI00025252.1 22 Keratin, type II cytoskeletal 1 IPI00220327.2 Keratin, type II cytoskele IPI00290078.2 Keratin, type II cytoskeletal 7 IPI00306959.7 48 kDa protein IPI00332128.1 Hypothetical protein DKFZp566O224 IPI00383815.3 Keratin 8 variant IPI00418411.4 Keratin b20 IPI00431749.1 Hypothetical protein FLJ45472 IPI00443478.1 Keratin, type II cytoskeletal 8 IPI00554648.1 Keratin 1 IPI00556624.1 23 Pyruvate kinase 3 isoform 2 IPI00220644.6 Pyruvate kinase M2 IPI00383237.3 Pyruvate kinase 3 isoform 1 variant IPI00479186.2 Hypothetical protein IPI00604528.1 24 Tryptophanyl-tRNA synthetase IPI00295400.1 TrypTophanyl-TRNA synTheTase isoform b IPI00412737.1 25 Succinyl-CoA:3-ketoacid-coenzyme A transferase 1, mitochondrial precursor IPI00026516.1 26 Heat shock 70kDa protein 1A IPI00514377.3 27 Tubulin beta-? chain IPI00007752.1 Tubulin beta-2 chain IPI00011654.2 Tubulin beta-3 chain IPI00013683.2 Tubulin beta-4 chain IPI00023598.1 Tubulin, beta polypeptide paralog IPI00031370.3 Similar to Tubulin beta-4 chain IPI00034283.2 Tubulin, beta, 4 IPI00152453.1 Tubulin, beta 4 IPI00160897.3 Class IVb beta tubulin IPI00549172.2 TUBB protein IPI00549853.2 28 Tubulin alpha-1 chain IPI00007750.1 TUBA6 protein IPI00166768.1 Tubulin alpha-3 chain IPI00180675.4 Tubulin alpha-8 chain IPI00216005.5 Tubulin alpha-6 chain IPI00218343.4 K-ALPHA-1 protein IPI00328163.1 Tubulin alpha-ubiquitous chain IPI00387144.4 Similar to alpha tubulin IPI00410402.1 46 kDa protein IPI00414274.2 29 50 kDa protein IPI00174849.4 30 Nucleoporin 54kDa variant IPI00172580.4 Hypothetical protein DKFZp434L1613 IPI00385666.1 Splice Isoform 2 of Nucleoporin p54 IPI00386702.1 31 26S protease regulatory subunit 7 IPI00021435.2 PREDICTED: similar to proteasome (prosome, macropain) 26S subunit, ATPase 2 IPI00454870.1 32 Glutathione synthetase IPI00010706.1 33 Actin-like protein 3 IPI00028091.1 34 Actin, alpha 1, skeletal muscle IPI00514530.3 35 Sorting nexin 6, isoform a IPI00258833.1 Sorting nexin 6 IPI00298111.5 45 kDa protein IPI00513753.1 36 D-3-phosphoglycerate dehydrogenase IPI00011200.4 37 Rab GDP dissociation inhibitor alpha IPI00010154.3 Rab GDP dissociation inhibitor beta IPI00031461.1 GDP dissociation inhibitor 2 IPI00513829.1 GDP dissociation inhibitor 2 IPI00513929.1 GDP dissociation inhibitor 2 IPI00514395.1 38 Alpha enolase, lung specific IPI00013769.1 Gamma enolase IPI00216171.2 Beta enolase IPI00218474.4 Alpha enolase IPI00465248.4 39 Splice Isoform Long of Glucose-6-phosphate 1-dehydrogenase IPI00216008.2 Glucose-6-phosphate dehydroGenase IPI00289800.6 40 Splice Isoform 1 of SET protein IPI00072377.1 Splice Isoform 2 of SET protein IPI00301311.1 PREDICTED: similar to SET protein (Phosphatase 2A inhibitor I2PP2A) (I-2PP2A) (Template IPI00397984.2 41 Septin-2 IPI00014177.1 42 Splice Isoform C2 of Heterogeneous nuclear ribonucleoproteins C1/C2 IPI00013944.2 Heterogeneous nuclear ribonucleoprotein C-like dJ845O24.4 IPI00027569.1 Splice Isoform C1 of Heterogeneous nuclear IPI00216592.1 ATPase family, AAA domain containing 3A IPI00295992.2 Hypothetical protein FLJ10709 IPI00396219.4 Full-length cDNA clone CS0DA009YK08 of Neuroblastoma of Homo sapiens IPI00477313.1 HNRPC protein IPI00552125.1 24 kDa protein IPI00556280.2 43 Splice Isoform 2 of Nucleophosmin IPI00220740.1 PREDICTED: similar to nucleophosmin 1 IPI00455423.1 06 PREDICTED: similar to nucleophosmin 1 IPI00457100.1 Nucleophosmin IPI00549248.3 Nucleophosmin IPI00554482.1 44 Splice Isoform 1 of RNA-binding protein 4 IPI00003704.2 45 COP9 sig IPI00171844.3 COP9 signalosome subunit 4 variant IPI00604797.1 46 Macrophage capping protein IPI00027341.1 47 Splice Isoform 1 of 3'(2'),5'-bisphosphate nucleotidase 1 IPI00410214.1 Splice Isoform 2 of 3'(2'),5'-bisphosphate nucleotidase 1 IPI00410215.4 3'(2'), 5'-bisphosphate nucleotidase 1 IPI00479394.2 48 Maspin precursor IPI00472082.1 49 DnaJ homolog subfamily B member 11 precursor IPI00008454.1 50 HypotHetical protein LOC345651 IPI00003269.1 Actin, aortic smooth muscle IPI00008603.1 Actin, alpha skeletal muscle IPI00021428.1 Actin, cytoplasmic 1 IPI00021439.1 Actin, cytoplasmic 2 IPI00021440.1 Actin, alpha cardiac IPI00023006.1 Actin, gamma-enteric smooth muscle IPI00025416.1 Actin, alpha 1, skeletal muscle IPI00515047.1 Actin, cytoplasmic 1 IPI00550724.3 FKSG30 IPI00555900.1 51 Galactokinase IPI00019383.1 Hypothetical protein FLJ26554 IPI00442827.1 52 Proliferating cell nuclear antigen IPI00021700.1 53 F-actin capping protein alpha-1 subunit IPI00005969.1 F-actin capping protein alpha-2 subunit IPI00026182.4 MGC14151 protein IPI00304922.1 PREDICTED: similar to SERTA domain containing 4 IPI00402160.2 54 60S acidic ribosomal protein