However, it is BYL719 Possible that at least some of them might be functionally membrane-associated through formation of protein complexes with membrane-anchored proteins. In a previous study we showed that several hydrophilic proteins are retained in the lipophilic membrane fraction due to interaction with hydrophobic proteins [21–23]. Relative abundance index To estimate the relative abundance of the

observed proteins, we used the emPAI algorithm, which is based on the calculation of identified peptides per protein and normalized by the theoretical number of peptides for the same protein (PAI). The outcome of the emPAI analysis is given for a selection of membrane proteins and lipoproteins with the highest values in Table 2 and 3, respectively. At the top of the membrane protein list is the possible proline rich antigen AZD5153 ic50 QNZ pra (Rv1078), with 5.66 mol %. This is a small protein with 25 kDa, and has 2 TMHs. When digested with trypsin, it constitutes 6 observable tryptic

peptides, where 5 of them were identified. This protein has also been observed in M. bovis [14, 24]. The membrane proteins Rv1078 and Rv1489 are the most abundant ones, but with no annotated biological functions. In the lipoprotein list only the first three proteins are assigned functions, while the 7 others have unknown biological functions. Table 2 List of the 14 most frequently observed membrane proteins. Sanger ID Gene name Protein identity No. of TMH a No. of observed peptides b emPAI (Mol %) c References Rv1078 pra Possible proline rich antigen 2 5 5.66 [14, 24] Rv1489 – Conserved hypothetical protein 2 5 1.30 [26] Rv1306 atpF Possible ATP synthase b chain 1 7 0.36 [14, 24–26] Rv2563 – Possible glutamine-transport transmembrane protein 4 13 0.35 [14, 25, 26, 32] Rv1234 – Possible transmembrane protein 2 7 0.26 [25, 26] Rv0072 – Possible glutamine-transport transmembrane protein 4 11

0.23 [25, 26] Rv0479c – Possible conserved membrane protein 1 11 0.23 [24–26] Rv2969c – Possible conserved membrane or secreted protein 1 11 0.19 [14, 24–26, 40] Rv2200c ctaC Possible transmembrane cytochrome C oxidase 3 13 0.17 [14, 24–26, 32] Rv2195 qcrA Possible rieske iron-sulfur protein 3 15 0.16 [14, 24–26, 40, 54] Rv1223 htrA Possible serine protease 1 19 0.15 [24, 26, 54] Rv1822 – Phosphatidylglycerophosphate Florfenicol synthase 4 5 0.14 [14] Rv2721c – Possible conserved transmembrane protein 2 12 0.13 [14, 24–26, 32] Rv3273 – Possible transmembrane carbonic anhydrase 10 11 0.11 [24–26, 54] a Number of TMH regions predicted by TMHMM version 2.0 publically available at http://​www.​cbs.​dtu.​dk/​services/​TMHMM/​. b Number of observed unique peptides from each protein. c Relative protein abundance provided in mol % concentration. Table 3 List of the 10 most frequently observed lipoproteins. Sanger ID Gene name Protein identity No. of observed peptides a emPAI (Mol %) b References Rv0432 sodC Possible periplasmic superoxide dismutase 6 2.36 [14, 24–26, 40] Rv3763 lpqH 19 kda lipoprotein antigen precursor 3 1.