ELECTROPHORESIS Supporting Information for Electrophoresis DOI 10.1002/elps.200406079 Michalis Aivaliotis, Carsten Corvey, Irene Tsirogianni, Michael Karas and Georgios Tsiotis Membrane proteome analysis of the green-sulfur bacterium Chlorobium tepidum ã 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Electrophoresis 2004, 25, 0001±0010 1 6 Addendum Table 1. List of identified proteins separated in 2-D gel with Triton X-100 No. Protein definition Accession Predicted Peptides Peptide TM GRAVYSignalP Aro- Predicted b) No./gene Mr/pI matched coverage a-heli- matic localization (%) cesa) anchor Transport and binding proteins 1 ArsA ATPase family protein Q8KDR5/CT0980 44541/5.0 17 48 1 20.117 ± ± Inner membrane 2 Outer membrane efflux Q8KED1/CT0758 52716/8.9 8 16 3 20.163 ± ± Outer membrane protein, putative 3 ABC transporter, periplasmic Q8KDW4/CT0930 33207/8.9 9 26 3 20.388 YES 1±25 ± Inner membrane substrate-binding protein Energy metabolism 4 ATP synthase F1, a-subunit Q8KAW8/ATPA 56869/6.3 31 50 2 20.051 ± ± Inner membrane 5 ATP synthase, b-chain Q8KAC9/ATPD 50125/5.0 37 77 1 20.130 ± ± Inner membrane 6 ATP synthase F0, B subunit Q8KGE9/ATPF 19437/7.7 8 33 1 20.225 YES 1±39 ± Inner membrane 7 Bacteriochlorophyl A protein Q46393/FMOA 40383/7.1 34 78 ± 20.288 ± ± Inner membrane 8 Chlorosome envelope O68991/GSMH 21779/4.9 7 44 2 20.125 ± ± Chlorosome protein H 9 Chlorosome envelope O68988/GSMI 25911/7.6 11 60 2 0.082 ± ± Chlorosome protein I 10 Chlorosome envelope Q8KEN5/CSMX 23970/5.7 9 40 ± 20.155 ± ± Chlorosome protein X 11 Cytochrome b6f complex Q9F722/PETC 18733/9.2 13 62 3 0.060 YES 1±24 ± Inner membrane Rieske, iron-sulfur subunit 12 Photosystem P840 RC Q8KEP5/PSCD 16607/9.7 8 45 ± 20.558 ± ± Inner membrane protein PscD 13 NADH dehydrogenase I, Q8KEC2/NDHH 20925/8.9 12 73 1 20.052 ± ± Cytoplasmic 20 kDa subunit Protein fate 14 ATP-dependent Cip protease, Q8KG79/CIPB-2 49654/5.3 28 62 ± 20.440 YES 1±4 ± Cytoplasmic ATP-binding subunit Cipb 15 Carboxyl-terminal protease Q8KDJ6/PRC 75483/9.7 13 20 ± 20.410 ± ± Unknown 16 Propyl oligopeptidase family Q8KCV9/CT1301 77944/6.5 27 54 2 20.428 ± ± Periplasmic protein 17 Chaperonin, 60 kDa Q8KF02/GROEL 58080/5.2 37 77 ± 20.196 ± ± Cytoplasmic 18 Heat shock protein HtpG Q8KE61/HTPG 71037/5.1 42 52 2 20.243 ± ± Cytoplasmic 19 Heat shock protein Q8KB28/CT1970 16008/4.9 15 83 1 20.562 ± ± Unknown Hsp20 family 20 Chaperone protein DnaK Q8KEP3/DNAK 68387/5.2 33 52 1 20.396 ± ± Unknown 21 Polypeptide deformylase Q8KCG7/DEF 20909/5.9 10 58 ± 20.133 ± ± Cytoplasmic 22 Type-II secretion system Q8KF93/CT0434 52852/6.1 10 34 ± 20.196 ± ± Cytoplasmic protein Protein synthesis 23 30S ribosomal protein S4 P59129 /RPSD 23319/10.2 20 74 ± 20.628 ± ± Cytoplasmic 24 Ribosomal protein L2 Q8KAH5/RPLB 30373/10.7 24 65 ± 20.613 ± ± Cytoplasmic 25 Ribosomal protein L5 Q8KAI4/RPLE 21038/10.3 15 67 ± 20.328 ± ± Cytoplasmic 26 Ribosomal protein L6 Q8KAI6/RPLF 19598/9.9 14 66 1 20.230 ± ± Cytoplasmic 27 50S ribosomal protein L9 Q8KAM4/RPLI 16489/7.9 8 58 ± 20.275 ± ± Cytoplasmic ã 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 2 M. Aivaliotis et al. Electrophoresis 2004, 25, 0001±0010 Table 1. Continued No. Protein definition Accession Predicted Peptides Peptide TM GRAVYSignalP Aro- Predicted b) No./gene Mr/pI matched coverage a-heli- matic localization (%) cesa) anchor 28 50S ribosomal protein L10 Q8KG17/RPLJ 19282/10.1 11 73 1 20.210 ± ± Cytoplasmic 29 Ribosomal protein L13 Q8KBK4/RPLM 16903/9.9 7 49 ± 20.558 ± ± Cytoplasmic 30 Ribosome recycling factor Q8KC49/FRR 21222/9.0 13 61 ± 20.638 ± ± Cytoplasmic Cell envelope 31 Outer surface protein, Q8KAL2/CT2144 20056/5.1 13 42 2 0.013 YES 1±19 F Outer membrane putative 32 Outer membrane protein Q8KFR8/OMPH 20430/9.5 22 67 1 20.448 YES 1±40 ± Outer membrane OmpH 33 Peptidoglycan-associated Q8KEP8/CT0638 17850/4.8 10 62 2 20.334 ± ± Outer membrane lipoprotein Unknown function 34 Pentapeptide repeat family Q8KAT8/CT2067 46535/6.6 16 36 3 20.149 ± ± Unknown protein 35 WD-repeat family protein Q8KB12/CT1986 35900/8.0 23 63 1 20.239 YES 1±8 ± Unknown 36 Hemaglutinin-related protein Q8KGA0/CT0068 22146/4.8 15 55 ± 20.208 ± F Outer membrane Central intermediary metabolism 37 Sulfide dehydrogenase, Q8KDM1/CT1025 45492/7.2 18 46 1 20.137 ± ± Periplasmic flavoprotein subunit, putative 38 Polysulfide reductase, Q8KAC2/CT2241 28963/8.1 12 56 1 20.345 YES 1±31 ± Cytoplasmic subunit B, putative 39 Sulfide-quinone reductase, Q8KG51/CT0117 53974/6.7 9 11 5 20.215 ± ± Inner membrane putative 40 Nitrogenase molybdenium- Q8KC89/NIFD 60680/6.1 14 32 3 20.336 ± ± Unknown iron protein, a-subunit Biosynthesis of cofactors, prosthetic groups and carriers 41 Zeta-carotene desaturase Q8KCK1/CT1414 50979/6.7 19 35 3 20.229 ± ± Unknown Cellular processes 42 Cell division protein FtsZ Q8KGD8/FTSZ 46167/5.9 20 41 1 20.220 ± ± Cytoplasmic/ inner membrane 43 Bacterioferritin comigratory Q8KEM5/BCP-1 17815/6.5 18 79 ± 20.551 ± ± Unknown protein, thiol peroxidase, putative 44 Bacterioferritin comigratory Q8KAZ7/BCP-2 16354/5.1 7 46 ± 0.022 ± ± Unknown protein, thiol peroxidase, putative 45 Peroxiredoxin, putative Q8KBN8/CT1747 24047/6.1 17 70 1 20.236 ± ± Cytoplasmic Purines, pyriminides, nucleosides, and nucleotides 46 Adeninephosphoribosyl- Q8KFM9/APT 19319/5.3 5 48 2 0.100 ± ± Cytoplasmic transferase DNA metabolism 47 Regulatory protein RecX Q8KBK8/CT1778 17890/9.5 7 42 ± 20.790 ± ± Cytoplasmic ã 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Electrophoresis 2004, 25, 0001±0010 Proteomic analysis of C. tepidum 3 Table 1. Continued No. Protein definition Accession Predicted Peptides Peptide TM GRAVYSignalP Aro- Predicted b) No./gene Mr/pI matched coverage a-heli- matic localization (%) cesa) anchor Regulatory functions 48 Transcriptional regulator, Q8KDZ1/CT0903 27147/7.6 8 30 ± 20.253 ± ± Cytoplasmic putative Conserved hypothetical proteins 49 Conserved hypothetical Q8KAK9/CT2147 42377/5.3 31 58 2 20.228 ± ± Unknown protein 50 Conserved hypothetical Q8KG37/CT0131 33752/5.9 21 52 1 0.043 ± ± Unknown protein 51 Conserved hypothetical Q8KBC2/CT1867 24297/8.8 16 69 1 20.247 YES 1±17 ± Unknown protein Hypothetical proteins 52 Hypothetical protein Q8KA97/CT2270 21451/8.9 11 44 1 0.115 YES 1±25 Y Outer membrane 53 Hypothetical protein Q8KCV6/CT1304 21332/6.0 11 59 1 20.153 ± ± Unknown 54 Hypothetical protein Q8KBI6/CT1801 32338/6.3 23 63 1 20.178 YES 1±20 ± Unknown 55 Hypothetical protein Q8KDH0/CT1080 28665/6.9 10 44 2 20.024 YES 1±23 ± Unknown 56 Hypothetical protein Q8KE01/CT0893 42440/8.6 17 54 3 20.409 YES 1±44 F Outer membrane 57 Hypothetical protein Q8KBI3/CT1804 42811/5.6 28 74 2 20.248 YES 1±36 F Outer membrane 58 Hypothetical protein Q8KAL8/CT2138 19418/5.4 6 27 ± 20.455 ± ± Unknown a) Predicted by TMtrep program b) Predicted by PSORT algorithm Table 2. List of identified proteins separated in 2-D gel with SDS and acetone treatment No. Protein definition Accession Predicted Peptides Peptides TM SignalP GRAVYAro- Predicted b) No./gene Mr/pI matched coverage a-heli- matic localization (%) cesa) anchor Transport and binding proteins 1 Toluene transport protein, Q8KAD3/CT2230 49260/6.0 19 49 3 YES 1±23 20.083 ± Inner membrane putative 2 ABC-type export system, Q8KEY0/CT0552 26711/5.5 21 55 1 ± 20.205 ± Inner membrane ATP-binding subunit 3 ABC-type export system, Q8KEX9/ CT0553 34024/9.9 7 25 2 YES 1±20 20.076 ± Inner membrane membrane fusion 4 Outer membrane efflux Q8KED1/CT0758 52716/8.9 12 31 3 YES 1±42 20.163 ± Ouuter membran protein, putative 5 ArsA ATPase family protein Q8KDR5/CT0980 44541/5.0 22 64 1 ± 20.117 ± Inner membrane 6 ArsA ATPase family protein Q8KAT5/CT2070 44671/5.1 11 41 ± ± 20.226 ± Inner membrane 7 Phosphate ABC transporter, Q8KDZ9/CT0895 28379/9.5 14 63 1 YES 1±24 0.020 ± Inner membrane periplasmic substrate- binding protein, putative Energy metabolism 30 NADH dehydrogenase I, Q8KEB8/NDHI 23569/7.4 13 44 1 ± 20.388 ± Cytoplasmic 23 kDa subunit or CT0771 ã 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 4 M. Aivaliotis et al. Electrophoresis 2004, 25, 0001±0010 Table 2. Continued No. Protein definition Accession Predicted Peptides Peptides TM SignalP GRAVYAro- Predicted b) No./gene Mr/pI matched coverage a-heli- matic localization (%) cesa) anchor 31 Thioredoxin reductase Q8KE48/TRXB 34049/5.2 19 71 1 ± 20.104 ± Cytoplasmic or CT0842 32 Cytochrome b6f complex Q9F722/PETC 18864/9.2 9 50 3 YES 1±19 0.060 ± Inner membrane Rieske, iron-sulfur subunit or CT0302 33 Transketolase, C-terminal Q8KE86/CT0804 35681/5.7 18 66 1 ± 20.023 ± Cytoplasmic subunit 34 Enolase Q8KB35/ENO-1 46383/5.8 25 55 ± ± 20.306 ± Cytoplasmic or CT1962 35 2,3-Bisphosphoglycerate- Q8KFC8/GPMA or 28434/6.1 12 45 ± ± 20.552 ± Cytoplasmic dependent phosphoglycerate GPM or CT0399 mutase 36 ATP synthase, b-chain Q8KAC9/ATPD 50125/5.0 29 81 1 ± 20.130 ± Inner membrane or CT2234 37 ATP synthase F1, a-subunit Q8KAW8/ATPA 56848/6.3 36 58 2 ± 20.051 ± Inner membrane or CT2033 38 Chlorosome envelope Q8KEN5/CSMX 23970/5.7 12 55 ± ± 20.155 ± Chlorosome protein X or CT0652 39 Chlorosome envelope O68988/CSMI 25929/7.6 11 45 2 ± 0.082 ± Chlorosome protein I or CT1382 40 Chlorosome envelope O68991/CSMH 21779/5.0 12 67 2 ± 20.125 ± Chlorosome protein H or CT1417 41 Bacteriochlorophyl A protein Q46393/FMOA 40383/7.1 34 86 ± ± 20.288 ± Inner membrane or CT1499 42 Carbohydrate kinase, Q8KDR9/CT0976 34608/4.5 22 66 2 ± 20.187 ± Unknown PfkB family 43 NAD-dependent epimerase/ Q8KFU2/CT0230 39061/5.6 9 43 2 ± 20.244 ± Unknown dehydratase family protein 44 Alcohol dehydrogenase, Q8KDA2/CT1152 40657/5.4 10 33 2 ± 0.015 ±