Microsomes are derived mostly from endoplasmic reticulum and are an ideal target to investigate compound metabolism, membrane-bound enzyme functions, lipid-protein interactions, and drug-drug interactions. To better understand the molecular mechanisms of the liver and its diseases, mouse liver microsomes were isolated and enriched with differential centrifugation and sucrose gradient centrifugation, and microsome membrane proteins were further extracted from isolated microsomal fractions by the carbonate method. The enriched microsome proteins were arrayed with two-dimensional gel electrophoresis (2DE) and carbonate-extracted microsome membrane proteins with one-dimensional gel electrophoresis (1DE). A total of 183 2DE-arrayed proteins and 99 1DE-separated proteins were identified with tandem mass spectrometry. A total of 259 nonredundant microsomal proteins were obtained and represent the proteomic profile of mouse liver microsomes, including 62 definite microsome membrane proteins. The comprehensive bioinformatics analyses revealed the functional categories of those microsome proteins and provided clues into biological functions of the liver. The systematic analyses of the proteomic profile of mouse liver microsomes not only reveal essential, valuable information about the biological function of the liver, but they also provide important reference data to analyze liver disease-related microsome proteins for biomarker discovery and mechanism clarification of liver disease.
The liver, a vital organ, has a wide range of physiological functions and plays a major role in metabolism, biosynthesis, and chemical neutralizing. Liver diseases, such as viral hepatitis and liver cancer, pose a worldwide public health challenge. The Human Liver Proteome Project (HLPP) was launched in 2002 to better understand molecular liver functions and diseases, and liver proteome expression profile is one of the major parts of HLPP [
Microsomes are composed primarily of closed sacs of membrane called vesicles that are derived mostly from endoplasmic reticulum (ER). As for liver, in addition to components of the protein secretary pathway, microsomes contain a multitude of proteins that are involved in lipid/lipoprotein biosynthesis and drug metabolism. The liver microsome is an ideal way to study the metabolism of compounds, the functional properties of membrane-bound enzymes, lipid-protein interactions, and drug-drug interactions [
Two-dimensional gel electrophoresis (2DE) is one of the most widespread techniques for the proteomic profiling of soluble proteins and visualizes isoforms and posttranslational modifications in a proteome [
Although many of microsome proteins have been studied, many more remain to be isolated and characterized. With the improvement of current methodologies and experimental techniques, more proteomic data will be obtained. Also, biological interpretation of proteomic data and extracting biological knowledge are essential to further understanding liver function.
In our study, 2DE was first used to array the isolated microsome proteins of the liver. Because of the low performance of 2DE in separating membrane proteins [
Male C57 mice (9 weeks old) were purchased from the Experimental Animal Center of Central South University (Changsha, China). The mice were starved overnight for liver subcellular fractionation. All experiments were performed with the approval of the institutional ethics committee on animal research.
Microsome apparatus-rich fractions were prepared from mice livers with differential centrifugation and sucrose gradient centrifugation as described [
Electron microscopy and Western blotting were used to detect and validate the purity of prepared microsomes. For electron microscope analysis, the prepared microsomes were fixed with 2.5% glutaraldehyde for 24 h and 2% OsO4 for 2 h, dehydrated with alcohol (50%, 70%, 90%, and 100% in turn), and processed into epoxy resin. Thin sections (500 Å
2DE was performed as described by the manufacturer (Amersham Biosciences). Protein samples (400
Microsome membrane proteins were further extracted by the carbonate method from isolated microsomal fractions [
The proteins contained in the 2D gel spots and 1D gel bands were subjected to in-gel digestion with trypsin. Gel spots or bands were excised and destained with 100 mM NH4HCO3 in 50% acetonitrile (ACN) at room temperature. The proteins were reduced with 10 mM dithiothreitol (DDT) (56°C; 30 min) and alkylated with 50 mM iodoacetamide in 100 mM NH4HCO3 (dark, room temperature, 30 min). The gel pieces that contained proteins were dried and then incubated in the digestion solution (40 mM NH4HCO3, 9% ACN, and 20
The tryptic peptide mixture was fractionated with reverse-phase (RP) high-performance liquid chromatography (HPLC) by using an Ultimate nano-HPLC system (Dionex). Peptide samples were purified and concentrated with a C18-PepMap precolumn and then separated on an analytical C18-PepMap column (75
Protein annotations were obtained primarily from UniProt 7.0 including accession, entry name, comments such as function, catalytic activity, subcellular location, and similarity. The Cytoscape plugin, Biological Networks Gene Ontology (BinGO), was used to find statistically overrepresented GO categories of the protein dataset. An online tool, WebGestalt (
It was essential to obtain a highly pure fraction to conduct proteomic characterization of microsomes. The purity of prepared microsomes was monitored with electron microscope and Western blotting analysis. A large number of nearly spherical membrane vesicles were visualized with electron microscope without other contaminated organelle compositions (see Supplemental Figure 1(a) in Supplementary Material available online at doi:10.1155/2012/832569). Western blotting analyses showed that, with the standard immunoblotting protocol, the ER marker endoplasmin was enriched in the isolated microsome fractions without the contamination marker (mitochondrial marker OxPhos Complex IV subunit I, peroxisomal marker catalase, and cytoplasmic marker cadherin) being detected (Supplemental Figure 1(b)). The results demonstrated an optimized preparation of microsomes.
The 2DE reference maps display
Proteins identified from mouse liver microsomal preparations with 2DE-based strategy.
Spot no. | IPIa | Protein name | Predicted TMD | Location |
---|---|---|---|---|
90,91 | IPI00108939 | glyceraldehyde-3-phosphate dehydrogenase, spermatogenic | 0 | ER |
6 | IPI00111936 | UDP-glucuronosyltransferase 1-2 precursor, microsomal | 1 | ER |
145 | IPI00121833 | Acetyl-coenzyme A acyltransferase 1 | 0 | ER |
102 | IPI00622235 | Transitional endoplasmic reticulum ATPase | 0 | ER |
6,61,194 | IPI00122815 | Prolyl 4-hydroxylase, beta polypeptide | 0 | ER |
17 | IPI00123176 | Similar to glyceraldehyde-3-phosphate dehydrogenase, 37 kDa protein | 0 | ER |
134,135 | IPI00123342 | Hypoxia upregulated 1 | 1 | ER |
2 | IPI00129526 | Endoplasmin | 0 | ER |
139 | IPI00131459 | Nucleoside diphosphate kinase A | 0 | ER |
179 | IPI00132874 | Splice isoform 1 of monoglyceride lipase | 0 | ER |
163 | IPI00133522 | Protein disulfide-isomerase precursor | 0 | ER |
49 | IPI00134058 | Thioredoxin domain containing protein 4 precursor | 0 | ER |
108,145,65 | IPI00135284 | Similar to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) ISOFORM 1 | 0 | ER |
147,148,149 | IPI00135686 | Mus musculus adult male kidney cDNA, RIKEN full-length enriched library, clone: 0610008 | 1 | ER |
174,178,179,183 | IPI00135726 | Similar to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) | 0 | ER |
49,50 | IPI00163011 | Thioredoxin domain containing protein 5 precursor | 0 | ER |
137 | IPI00226993 | Thioredoxin | 0 | ER |
148 | IPI00229551 | ADAM4 | 1 | ER |
62,157,158,162 | IPI00230108 | Glucose-regulated protein, full insert sequence | 0 | ER |
148 | IPI00271869 | Similar to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) | 0 | ER |
146,147,149,150,153 | IPI00273646 | Glyceraldehyde-3-phosphate dehydrogenase | 0 | ER |
187 | IPI00555023 | Glutathione S-transferase P 1 | 0 | ER |
144 | IPI00319652 | Glutathione peroxidase | 0 | ER |
84 | IPI00319992 | 78 kDa glucose-regulated protein precursor | 0 | ER |
153 | IPI00320208 | Elongation factor 1-beta | 0 | ER |
118 | IPI00323357 | Heat shock cognate 71 kDa protein | 0 | ER |
173 | IPI00323661 | Similar to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) | 0 | ER |
145,201 | IPI00462605 | Similar to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) | 0 | ER |
127 | IPI00469307 | Alpha-2-macroglobulin receptor-associated protein precursor | 0 | ER |
143,152,203 | IPI00480343 | 2700050F09Rik protein | 0 | ER |
162 | IPI00831714 | Leucine-rich repeat-containing protein 7 | 0 | ER (integral to membrane) |
149,150 | IPI00352124 | Flavin containing monooxygenase 5 | 1 | ER (integral to membrane) |
131 | IPI00132397 | GTP-binding protein SAR1b | 0 | ER (peripheral membrane protein) |
107 | IPI00227657 | Stromal cell-derived factor 2-like protein 1 precursor | 0 | ER lumen |
145 | IPI00123281 | Expressed sequence AA959742 | 1 | ER membrane |
7 | IPI00222496 | Protein disulfide isomerase-associated 6 | 1 | ER, membrane proteinb |
7 | IPI00112322 | UDP-glucuronosyltransferase 2B5 precursor | 1 | ER, membrane proteinsb |
156 | IPI00331322 | Microsomal glutathione S-transferase 1 | 3 | ER, outer membrane |
151 | IPI00319973 | Membrane-associated progesterone receptor component 1 | 1 | ER, membrane-bound |
152 | IPI00170316 | Multiple coagulation factor deficiency protein 2 homolog precursor | 0 | ER-Golgi intermediate compartment |
170 | IPI00408892 | RAS-related protein RAB-7 | 0 | Golgi, endosomes, lysosomes |
153 | IPI00123316 | Splice isoform 1 of tropomyosin 1 alpha chain | 0 | Cytoplasm |
116 | IPI00133456 | Senescence marker protein-30 | 0 | Cytoplasm |
89,129,130,151,152 | IPI00135085 | Heme-binding protein | 0 | Cytoplasm |
165 | IPI00109061 | Tubulin beta-4 chain homolog | 0 | Cytoplasmic |
88 | IPI00109073 | Tubulin beta-4 chain | 0 | Cytoplasmic |
105,138 | IPI00110753 | Tubulin alpha-1 chain | 0 | Cytoplasmic |
113,166,167,197,204 | IPI00110827 | Actin, alpha skeletal muscle | 0 | Cytoplasmic |
9,129,130,151.153,166, | IPI00110850 | Actin, cytoplasmic 1 | 0 | Cytoplasmic |
90 | IPI00114162 | Fatty acid-binding protein, epidermal | 0 | Cytoplasmic |
145 | IPI00116277 | T-complex protein 1, delta subunit | 0 | Cytoplasmic |
144 | IPI00117264 | DJ-1 protein | 0 | Cytoplasmic |
191,164,165 | IPI00117348 | Tubulin alpha-2 chain | 0 | Cytoplasmic |
137,138,164 | IPI00117350 | Tubulin alpha-4 chain | 0 | Cytoplasmic |
141,153,165,132 | IPI00117352 | Tubulin beta-5 chain | 0 | Cytoplasmic |
126 | IPI00117914 | Arginase 1 | 0 | Cytoplasmic |
152 | IPI00120532 | 21 kDa protein | 0 | Cytoplasmic |
107,108,139,143,182 | IPI00125489 | 44 KD protein (Argininosuccinate synthase) | 0 | Cytoplasmic |
191 | IPI00626790 | Glutamine synthetase | 0 | Cytoplasmic |
176,182,194 | IPI00130950 | Betaine-homocysteine S-methyltransferase | 0 | Cytoplasmic |
99 | IPI00131204 | UDP-glucose pyrophosphorylase 2 | 0 | Cytoplasmic |
204 | IPI00136929 | Gamma actin-like protein | 0 | Cytoplasmic |
101,132 | IPI00169463 | Tubulin beta-2C Chain | 0 | Cytoplasmic |
202,133 | IPI00221400 | Alcohol dehydrogenase A chain | 0 | Cytoplasmic |
89 | IPI00221528 | Actin, cytoplasmic type 5 homolog | 0 | Cytoplasmic |
168 | IPI00221890 | Carbonic anhydrase III | 0 | Cytoplasmic |
202,133 | IPI00317740 | Guanine nucleotide-binding protein beta subunit 2-like 1 | 0 | Cytoplasmic |
159 | IPI00331174 | T-complex protein 1, eta subunit | 0 | Cytoplasmic |
154 | IPI00338039 | Tubulin, beta 2 | 0 | Cytoplasmic |
141 | IPI00348094 | Predicted: similar to tubulin M beta 1 | 0 | Cytoplasmic |
136 | IPI00404011 | Microtubule-associated protein | 0 | Cytoplasmic |
153 | IPI00421223 | Tropomyosin alpha 4 chain | 0 | Cytoplasmic |
194,195 | IPI00457825 | Similar to argininosuccinate synthase (Citrulline-aspartate ligase) | 0 | Cytoplasmic |
60 | IPI00462072 | Similar to alpha enolase (2-phospho-D-glycerate hydro-lyase) | 0 | Cytoplasmic |
178 | IPI00467066 | Glycine N-methyltransferase | 0 | Cytoplasmic |
63,109 | IPI00467833 | Triosephosphate isomerase | 0 | Cytoplasmic |
153 | IPI00605380 | Similar to tubulin alpha-2 chain (Alpha-tubulin 2) | 0 | Cytoplasmic |
162 | IPI00123313 | Ubiquitin-activating enzyme E1 1 | 0 | Cytoplasmic and nuclear |
64 | IPI00420745 | Proteasome subunit, alpha type 2, full insert sequence | 0 | Cytoplasmic and nuclear |
145 | IPI00320165 | Oxidoreductase HTATIP2 | 0 | Cytoplasmic and nuclear |
153 | IPI00117978 | Cytochrome c oxidase subunit IV isoform 1, mitochondrial precursor | 1 | Mitochondrial inner membrane |
19 | IPI00109167 | NADH-ubiquinone oxidoreductase 24 kDa subunit | 0 | Mitochondrial inner membrane |
158 | IPI00111885 | Ubiquinol-cytochrome-c reductase complex core protein I, mitochondrial precursor | 0 | Mitchondrial inner membrane |
175 | IPI00121322 | Electron transfer flavoprotein-ubiquinone oxidoreductase, mitochondrial precursor | 0 | Mitchondrial inner membrane |
196 | IPI00128023 | NADH-ubiquinone oxidoreductase 49 kDa subunit, mitochondrial precursor | 0 | Mitchondrial inner membrane |
134 | IPI00111908 | Predicted: carbamoyl-phosphate synthetase 1 | 0 | Mitochondrial |
145 | IPI00114840 | Endonuclease G, mitochondrial precursor | 0 | Mitochondrial |
70 | IPI00331555 | 2-oxoisovalerate dehydrogenase alpha subunit, mitochondrial precursor | 0 | Mitochondrial |
94,95 | IPI00115607 | Trifunctional enzyme beta subunit, mitochondrial precursor | 0 | Mitochondrial |
145 | IPI00115824 | NipSnap1 protein | 0 | Mitochondrial |
22 | IPI00116154 | Cytochrome c oxidase, subunit vb, full insert sequence | 0 | Mitochondrial |
15,146,147,148,149,100 | IPI00118986 | ATP synthase oligomycin sensitivity conferral protein, mitochondrial precursor | 0 | Mitochondrial |
127 | IPI00119138 | Ubiquinol-cytochrome-c reductase complex core protein 2, mitochondrial precursor | 0 | Mitochondrial |
147,148 | IPI00120984 | NADH-ubiquinone oxidoreductase 19 kDa subunit | 0 | Mitochondrial |
137 | IPI00129516 | Ubiquinol-cytochrome c reductase complex 11 kDa protein, mitochondrial precursor | 0 | Mitochondrial |
93,99,100,192,203 | IPI00130280 | ATP synthase alpha chain, mitochondrial precursor | 0 | Mitochondrial |
149,150 | IPI00132217 | Tetratricopeptide repeat protein 11 | 1 | Mitochondrial |
150,151 | IPI00132390 | NADH-ubiquinone oxidoreductase B15 subunit | 1 | Mitochondrial |
101,132,137,141,153 | IPI00170093 | NADH-ubiquinone oxidoreductase 23 kDa subunit, mitochondrial precursor | 0 | Mitochondrial |
92,93,94,95,96 | IPI00223092 | Hydroxyacyl-coenzyme A dehydrogenase/3-ketoacyl-coenzyme A | 0 | Mitochondrial |
142,143,152 | IPI00230507 | ATP synthase D chain, mitochondrial | 0 | Mitochondrial |
162 | IPI00308882 | NADH-ubiquinone oxidoreductase 75 kDa subunit, mitochondrial precursor | 0 | Mitochondrial |
149 | IPI00344004 | 13 KDa differentiation-associated protein | 0 | Mitochondrial |
145,146 | IPI00420718 | Hydroxymethylglutaryl-CoA synthase, mitochondrial precursor | 0 | Mitochondrial |
51 | IPI00308885 | 60 kDa heat shock protein, mitochondrial | 0 | Mitochondrial |
153 | IPI00462250 | Similar to adenine nucleotide translocase | 3 | Mitochondrial |
85,165,167,203 | IPI00468481 | ATP synthase beta chain, mitochondrial precursor | 0 | Mitochondrial |
147 | IPI00117281 | Phospholipid hydroperoxide glutathione peroxidase, mitochondrial precursor | 0 | Mitochondrial and cytoplasmic |
169 | IPI00133240 | Ubiquinol-cytochrome c reductase iron-sulfur subunit, mitochondrial precursor | 0 | Mitochondrial inner membrane |
200,201 | IPI00230351 | Succinate dehydrogenase [ubiquinone] flavoprotein subunit, mitochondrial precursor | 0 | Mitochondrial inner membrane |
174 | IPI00132042 | Pyruvate dehydrogenase E1 component beta subunit, mitochondrial precursor | 0 | Mitochondrial matrix |
156 | IPI00315794 | Cytochrome b5 outer mitochondrial membrane isoform precursor | 1 | Mitochondrial outer membrane |
65,109,144,170,176,100 | IPI00134746 | Argininosuccinate synthase | 0 | mitochondrion |
145,146 | IPI00338536 | Succinate dehydrogenase [ubiquinone] iron-sulfur protein, mitochondrial precursor | 0 | Mitochondrion |
112 | IPI00122547 | Voltage-dependent anion-selective channel protein 2 | 0 | Mitochondrion outer membrane |
147 | IPI00131186 | Splice isoform 2 of transcription factor BTF3 | 0 | Nuclear |
149 | IPI00317794 | Nucleolin | 0 | Nuclear |
53 | IPI00331146 | UMP-CMP kinase | 0 | Nuclear |
150 | IPI00458856 | Similar to ZNF91L isoform 1 | 0 | Nuclear |
6 | IPI00461822 | E1A binding protein p300 | 0 | Nuclear |
55 | IPI00126172 | RIKEN cDNA 4931406C07, PTD012 homolog | 0 | Nuclear |
150,151 | IPI00113241 | 40S ribosomal protein S19 | 0 | Ribosome |
104 | IPI00116908 | Similar to 40 s ribosomal protein S12 | 0 | Ribosome |
147 | IPI00849793 | 60S ribosomal protein L12 | 0 | Ribosome |
81,127,145 | IPI00125971 | 26S protease regulatory subunit S10B | 0 | Ribosome |
199 | IPI00123604 | 40S ribosomal protein SA | 0 | Ribosome |
125 | IPI00135640 | 26S protease regulatory subunit 8 | 0 | Ribosome |
149,150,151 | IPI00139780 | 60S ribosomal protein L23 | 0 | Ribosome |
149,150 | IPI00222546 | 60S ribosomal protein L22 | 0 | Ribosome |
145 | IPI00314950 | 60S acidic ribosomal protein P0 | 0 | Ribosome |
149 | IPI00322562 | 40S ribosomal protein S14 | 0 | Ribosome |
146 | IPI00331092 | 40S ribosomal protein S4, | 0 | Ribosome |
149,150 | IPI00331461 | 60S ribosomal protein L11 | 0 | Ribosome |
188 | IPI00351894 | Similar to ribosomal protein | 0 | Ribosome |
148 | IPI00849793 | 60S ribosomal protein L12 | 0 | Ribosome |
149 | IPI00465880 | 40S ribosomal protein S17 | 0 | Ribosome |
199 | IPI00123604 | 40S ribosomal protein SA | 0 | Ribosome |
4,11 | IPI00121209 | Apolipoprotein A-I precursor | 0 | Secreted |
149 | IPI00121837 | Ribonuclease 4 precursor | 1 | Secreted |
12 | IPI00122429 | Plasma retinol-binding protein precursor | 0 | Secreted |
163 | IPI00123920 | Alpha-1-antitrypsin 1–3 precursor | 0 | Secreted |
163 | IPI00123924 | Alpha-1-antitrypsin 1–4 precursor | 0 | Secreted |
163 | IPI00123927 | Alpha-1-antitrypsin 1–5 precursor | 0 | Secreted |
162 | IPI00128484 | Hemopexin precursor | 1 | Secreted |
3,117,118 | IPI00131695 | Serum albumin precursor | 0 | Secreted |
98 | IPI00139788 | Serotransferrin precursor | 0 | Secreted |
199 | IPI00323571 | Apolipoprotein E precursor | 0 | Secreted |
135,136 | IPI00377351 | Apolipoprotein A-IV precursor | 0 | Secreted |
163 | IPI00406302 | Alpha-1-antitrypsin 1-1 precursor | 0 | Secreted |
100,155,156 | IPI00466399 | 21 kDa protein | 0 | Secreted |
156 | IPI00480401 | Major urinary protein 1 precursor | 0 | Secreted |
122 | IPI00130661 | Tripeptidyl-peptidase I precursor | 0 | Secreted (lysosomal) |
101 | IPI00115302 | Branched chain ketoacid dehydrogenase E1, beta polypeptide | 0 | Membrane |
197 | IPI00120716 | Guanine nucleotide-binding protein G(I)/G(S)/G(T) beta subunit 1 | 0 | Membrane |
21,137 | IPI00120719 | Cytochrome c oxidase, subunit va, full insert sequence | 0 | Membrane |
125 | IPI00124790 | Polyposis locus protein 1-like 1 | 3 | Membrane |
129 | IPI00132076 | Catechol O-methyltransferase | 1 | Membrane |
130,142 | IPI00138406 | Ras-related protein Rap-1A | 0 | Membrane |
174 | IPI00162780 | Guanine nucleotide-binding protein G(I)/G(S)/G(T) beta subunit 2 | 0 | Membrane |
88,154 | IPI00230113 | Cytochrome b5 | 1 | Membrane |
199,200 | IPI00353727 | Annexin A4 | 0 | Membrane |
110 | IPI00117416 | Neighbor of COX4 | 0 | Unknown |
143 | IPI00121271 | Hypothetical S-adenosyl-L-methionine-dependent methyltransferases structure containing protein | 0 | Unknown |
144,108 | IPI00267667 | RIKEN cDNA 6330409N04, CLLL6 protein homolog | 0 | Unknown |
101 | IPI00269613 | Eukaryotic translation initiation factor 3 subunit 2 | 0 | Unknown |
149,150 | IPI00307837 | 51 kDa protein | 0 | Unknown |
203 | IPI00318204 | Sid6061p | 0 | Unknown |
105 | IPI00273646 | Similar to glyceraldehyde-3-phosphate dehydrogenase | 0 | Unknown |
189,190,194 | IPI00626790 | Glutamine synthetase | 0 | Unknown |
50 | IPI00345842 | 86 KDa PROTEIN | 0 | Unknown |
51 | IPI00350780 | 45 kDa protein | 0 | Unknown |
133 | IPI00381231 | 77 KDa protein | 0 | Unknown |
144 | IPI00923085 | Probable ubiquitin-conjugating enzyme E2 FLJ25076 homolog | 0 | Unknown |
146,147,150,173 | IPI00460295 | 44 KDa protein | 0 | Unknown |
156 | IPI00330913 | Major urinary protein 26 | 0 | Unknown |
59 | IPI00467988 | 169 kDa protein | 0 | Unknown |
100,155,156 | IPI00469517 | 21 kDa protein | 0 | Unknown |
149 | IPI00130554 | Splice isoform 1 of SNARE-associated protein Snapin | 0 | Vesicular membrane |
101,127,134 | IPI00131366 | Keratin, type II cytoskeletal 6B | 0 | Sarcolemma |
83,106,107 | IPI00121788 | Peroxiredoxin 1 | 0 | Microbody |
101,139 | IPI00348328 | Keratin Kb40 | 0 | Intermediate filament |
156 | IPI00137414 | Left-right dynein | 0 | Cilium |
aESI-Q-TOF identification, subcellular location are given for each ID number.
bThis protein is nonmembrane associated according to the annotation in the Swiss-Prot database but has one predicted TMD.
2DE pattern of mouse liver microsome. Microsomal proteins (400
The Na2CO3-treated microsome membrane proteins were separated on SDS-PAGE gels and visualized with Coomassie brilliant blue staining (Figure
Proteins identified from Na2CO3-extracted mouse liver microsomal membrane preparations with 1DE-based strategy.
Bands no. | Accession no. | Protein name | Predicted TMD | GRAVY score | Subcellular location | |
---|---|---|---|---|---|---|
9 | IPI00112322 | UDP-glucuronosyltransferase 2B5 precursor | 1 | −0.031 | 7.94 | ER |
9 | IPI00127223 | UDP glucuronosyltransferase 2 family, polypeptide B36 | 1 | −0.036 | 8.47 | ER |
9 | IPI00222496 | Protein disulfide-isomerase A6 | 1 | −0.292 | 5.05 | ER |
8 | IPI00417182 | UDP-glycosyltransferase 1 family polypeptide A5 | 1 | 0.044 | 8.33 | ER |
9 | IPI00116572 | Cytochrome P450, family 2, subfamily d, polypeptide 9 | 2 | −0.043 | 6.37 | ER |
15 | IPI00113655 | 40S ribosomal protein S6 | 0 | −0.918 | 10.68 | ER |
5 | IPI00129526 | Endoplasmin precursor (ER protein 99,94 kDa glucose-regulated protein) | 0 | −0.72 | 4.74 | ER |
13 | IPI00130985 | Short-chain dehydrogenase CRAD2 | 0 | 0.026 | 8.35 | ER |
6 | IPI00222809 | Similar to GDH/6PGL endoplasmic bifunctional protein | 0 | −0.18 | 6.61 | ER |
8 | IPI00230108 | Glucose-regulated protein, full insert sequence | 0 | −0.479 | 5.78 | ER |
10,11 | IPI00317356 | Paraoxonase 1 | 0 | −0.01 | 5.02 | ER |
7 | IPI00319992 | 78 kDa glucose-regulated protein precursor | 0 | −0.481 | 5.07 | ER |
13 | IPI00121079 | NADH-cytochrome b5 reductase 3 | 0 | −0.203 | 8.56 | ER, membrane bound |
9 | IPI00123964 | Cytochrome P450 2A5 | 1 | −0.203 | 9.23 | ER, membrane bound |
9 | IPI00114779 | Cytochrome P450 2C38 | 0 | −0.147 | 8.69 | ER, membrane bound |
17 | IPI00331322 | Microsomal glutathione S-transferase 1 | 3 | 0.14 | 9.67 | ER and mitochondrial outer membrane |
17 | IPI00119766 | Cis-retinol androgen dehydrogenase 1 | 0 | 0.005 | 9.25 | ER lumen |
8 | IPI00134691 | UDP-glucuronosyltransferase 1-1 precursor, microsomal | 2 | 0.087 | 8.87 | ER, integral to plasma membrane |
8 | IPI00128287 | Cytochrome P450 1A2 | 1 | −0.203 | 8.92 | ER, membrane bound |
10 | IPI00136910 | Cytochrome P450 2D11 | 2 | −0.009 | 6.15 | ER, membrane bound |
9 | IPI00308328 | Cytochrome P450 2F2 | 1 | −0.135 | 7.74 | ER, membrane bound |
9,10 | IPI00323908 | Cytochrome P450 2D10 | 2 | −0.073 | 6.16 | ER, membrane bound |
7 | IPI00112549 | Long-chain-fatty-acid-CoA ligase 1 | 1 | −0.045 | 6.81 | ER, type III membrane protein |
8 | IPI00133522 | Protein disulfide-isomerase precursor | 0 | −0.386 | 4.79 | ER |
9 | IPI00116572 | Cytochrome P450 2D9 | 0 | −0.063 | 5.93 | ER, membrane bound |
5,6 | IPI00119618 | Calnexin precursor | 1 | −0.875 | 4.5 | ER, type I membrane protein |
1,10,12,14,15 | IPI00319973 | Membrane-associated progesterone receptor component 1 | 1 | −0.616 | 4.57 | ER, membrane bound |
8 | IPI00132475 | Protein ERGIC-53 | 1 | −0.545 | 5.92 | ER-Golgi intermediate compartment (ERGIC), type I membrane protein |
8,17 | IPI00109061 | Tubulin beta-4 chain homolog | 0 | −0.406 | 4.78 | Cytoplasmic |
10 | IPI00110827 | Actin, alpha skeletal muscle | 0 | −0.232 | 5.23 | Cytoplasmic |
10,12,14 | IPI00110850 | Actin, cytoplasmic 1 | 0 | −0.2 | 5.29 | Cytoplasmic |
1,2,3 | IPI00111908 | Carbamoyl-phosphate synthase | 0 | −0.12 | 6.42 | Cytoplasmic |
1,8,9,13 | IPI00117348 | Tubulin alpha-2 chain | 0 | −0.23 | 4.94 | Cytoplasmic |
9,10,11,12,13 | IPI00117914 | Arginase 1 | 0 | −0.187 | 6.52 | Cytoplasmic |
17 | IPI00120451 | Fatty acid-binding protein, liver | 0 | −0.409 | 8.59 | Cytoplasmic |
9 | IPI00129028 | Similar to tubulin, alpha 3C isoform 1 | 0 | −0.204 | 4.98 | Cytoplasmic |
1–11,13,17 | IPI00130950 | Betaine-homocysteine S-methyltransferase | 0 | −0.36 | 8.01 | Cytoplasmic |
1,4,6,10,11,14,15 | IPI00134746 | Argininosuccinate synthase | 0 | −0.361 | 8.36 | Cytoplasmic |
3,4 | IPI00114710 | Pyruvate carboxylase, mitochondrial precursor | 0 | −0.173 | 6.25 | Mitochondrial |
17 | IPI00553333 | Hemoglobin subunit beta-1 | 0 | 0.092 | 7.13 | Mitochondrial |
9 | IPI00134809 | Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex | 0 | −0.171 | 9.1 | Mitochondrial |
17 | IPI00117978 | Cytochrome c oxidase subunit IV isoform 1, mitochondrial precursor | 1 | −0.412 | 9.25 | Mitochondrial inner membrane |
15,16 | IPI00315794 | Cytochrome b5 outer mitochondrial membrane isoform precursor | 1 | −0.602 | 4.79 | Mitochondrial outer membrane |
13 | IPI00321718 | Prohibitin-2 | 0 | −2.58 | 9.83 | Mitochondrial, cytoplasmic, nuclear |
13 | IPI00122547 | Voltage-dependent anion-selective channel protein 2 | 0 | −0.223 | 7.44 | Outer mitochondrial Membrane |
17 | IPI00114559 | Histone H2A type 1 | 0 | −0.572 | 11.22 | Nuclear |
16,17 | IPI00114642 | Histone H2B F | 0 | −0.762 | 10.32 | Nuclear |
8 | IPI00387318 | Cell cycle control protein 50A | 2 | −0.331 | 8.58 | Membrane |
15 | IPI00113849 | Splice isoform 2 of cell division control protein 42 homolog | 0 | −0.157 | 6.16 | Membrane |
13 | IPI00122549 | Splice isoform Pl-VDAC1 of voltage-dependent anion-selective channel protein 1 | 0 | −0.334 | 8.55 | Membrane |
15 | IPI00127408 | Ras-related C3 botulinum substrate 1 | 0 | −0.101 | 8.77 | Membrane |
15 | IPI00138406 | Ras-related protein Rap-1A | 0 | −0.375 | 6.39 | Membrane |
6 | IPI00116921 | Scavenger receptor class B member 1 | 2 | 0.073 | 8.29 | Integral membrane protein |
1 | IPI00121985 | Splice Isoform 1 of solute carrier organic anion transporter family, member 1B2 | 12 | 0.172 | 8.95 | Integral membrane protein |
9 | IPI00124830 | Integrin-associated protein precursor | 5 | 0.563 | 8.58 | Integral membrane protein |
14,15 | IPI00131176 | Cytochrome c oxidase subunit 2 | 2 | 0.27 | 4.6 | Integral membrane protein |
1 | IPI00132604 | Secretedretory carrier-associated membrane protein 3 | 4 | 0.028 | 7.55 | Integral membrane protein |
1 | IPI00135701 | Solute carrier organic anion transporter family, member 1A1 | 11 | 0.244 | 8.58 | Integral membrane protein |
1 | IPI00311682 | Sodium/potassium-transporting ATPase alpha-1 chain precursor | 10 | 0.002 | 5.3 | Integral membrane protein |
6 | IPI00331214 | Platelet glycoprotein IV | 2 | −0.053 | 8.61 | Integral membrane protein |
2 | IPI00119063 | Prolow-density lipoprotein receptor-related protein 1 | 1 | −0.502 | 5.17 | Integral to membrane |
1,12,16,17 | IPI00124790 | Polyposis locus protein 1-like 1 | 3 | 0.375 | 6.82 | Integral to membrane |
10 | IPI00129677 | Asialoglycoprotein receptor major subunit | 1 | −0.66 | 5.99 | Integral to membrane |
17 | IPI00467119 | Camello-like protein 1 | 1 | 0.302 | 9.61 | Integral to membrane |
5,8 | IPI00316329 | Keratin, type II cytoskeletal 1 | 0 | −0.588 | 8.2 | Intermediate filament |
10,11,12 | IPI00108844 | Cation-dependent mannose-6-phosphate receptor precursor | 1 | −0.168 | 5.24 | Type I membrane protein |
9 | IPI00109998 | H-2 class I histocompatibility antigen, D-B alpha chain precursor | 1 | −0.508 | 6.28 | Type I membrane protein |
4 | IPI00120245 | Integrin alpha-V precursor | 1 | −0.246 | 5.46 | Type I membrane protein |
3,4 | IPI00121190 | Epidermal growth factor receptor precursor | 2 | −0.316 | 6.46 | Type I membrane protein |
2 | IPI00126186 | Macrophage mannose receptor 1 precursor | 1 | −0.5 | 6.47 | Type I membrane protein |
5 | IPI00134549 | Splice isoform LAMP-2A of lysosome-associated membrane glycoprotein 2 precursor | 1 | −0.036 | 7.05 | Type I membrane protein |
13 | IPI00312018 | Malectin | 1 | −0.203 | 5.73 | Type I membrane protein |
3 | IPI00312063 | Low-density lipoprotein receptor precursor | 1 | −0.391 | 4.88 | Type I membrane protein |
16 | IPI00466570 | Transmembrane emp24 domain-containing protein 10 | 2 | −0.169 | 6.25 | Type I membrane protein |
4 | IPI00108535 | Carcinoembryonic antigen-related cell adhesion molecule 1 | 1 | −0.302 | 5.35 | Type I membrane protein |
5 | IPI00310059 | Polymeric-immunoglobulin receptor precursor | 1 | −0.499 | 5.26 | Type I membrane protein also secreted |
9 | IPI00121550 | Sodium/potassium-transporting ATPase beta-1 chain | 1 | −0.55 | 8.83 | Type II membrane protein |
2,3 | IPI00134585 | Glutamyl aminopeptidase | 1 | −0.344 | 5.28 | Type II membrane protein |
10 | IPI00307966 | ADP-ribosyl cyclase 1 | 1 | −0.106 | 8.64 | Type II membrane protein |
3 | IPI00319509 | Aminopeptidase N | 1 | −0.277 | 5.62 | Type II membrane protein |
3 | IPI00458003 | Ectonucleotide pyrophosphatase/phosphodiesterase 3 | 1 | −0.346 | 6.13 | Unknown |
9 | IPI00409409 | CD1D1 protein | 1 | −0.178 | 9.22 | Unknown |
7 | IPI00621548 | NADPH-cytochrome P450 reductase | 1 | −0.463 | 5.37 | Unknown |
9 | IPI00321644 | Cytochrome P450 2D26 | 2 | −0.105 | 6.16 | Unknown |
1 | IPI00127016 | Hydroxysteroid 17-beta dehydrogenase 6 | 0 | −0.075 | 8.63 | Unknown |
16 | IPI00221721 | Hypothetical krab box containing protein, full insert sequence | 0 | −0.142 | 9.84 | Unknown |
8 | IPI00224073 | Hypothetical peptidase family M20/M25/M40 containing protein, full insert sequence | 0 | −0.01 | 5.99 | Unknown |
16 | IPI00228379 | Ferritin light chain 2 | 0 | −0.479 | 6.37 | Unknown |
17 | IPI00266842 | 17 kDa protein | 0 | −0.668 | 10.48 | Unknown |
15 | IPI00379258 | Similar to ferritin light chain 2 | 0 | −0.454 | 8.51 | Unknown |
3 | IPI00405742 | Plexin B2 | 0 | −0.3 | 5.67 | Unknown |
2 | IPI00408258 | Structure-specific endonuclease subunit SLX4 | 0 | −0.714 | 5.33 | Unknown |
11 | IPI00462251 | Hypothetical protein LOC72792 isoform 1 | 0 | −0.429 | 5.82 | Unknown |
15 | IPI00605814 | Similar to Ferritin light chain 1 | 0 | −0.358 | 6.42 | Unknown |
10 | IPI00131366 | Keratin, type II cytoskeletal 6B | 0 | −0.488 | 8.32 | Sarcolemma |
10 | IPI00322209 | Keratin, type II cytoskeletal 8 | 0 | −0.602 | 5.7 | Sarcolemma |
8 | IPI00853991 | Similar to VH coding region | 0 | −0.102 | 5.31 | Secreted |
10 | IPI00126458 | MRNA | 1 | −0.55 | 5.66 | MHC class I protein complex |
15 | IPI00121788 | Peroxiredoxin 1 | 0 | −0.221 | 8.26 | Microbody |
1DE pattern and membrane-associated characteristic classification of Na2CO3-extracted microsomal membrane proteins. (a) 1DE pattern. Molecular weight markers are shown on the left and bands excised for MS analysis are indicated on the right. Lanes S1 and S2 were loaded with the same protein samples (50
Hydrophobicity is an important characteristic of a membrane protein. The grand average of hydropathy (GRAVY) scores (>−0.4) (
Among the 2DE dataset (
Proteins that are consistently present in both 2DE dataset of microsomal proteins (Table
Accession number | Protein name | Predicted TMD | GRAVY scores | Location | |
---|---|---|---|---|---|
IPI00108454 | Similar to 40S ribosomal protein S6 | 0 | −0.918 | 10.68 | ER |
IPI00112322a | UDP-glucuronosyltransferase 2B5 precursor | 1 | −0.031 | 7.94 | ER |
IPI00129526 | Endoplasmin precursor (ER protein 99, 94 kDa glucose-regulated protein) | 0 | −0.72 | 4.74 | ER |
IPI00133522 | Protein disulfide-isomerase precursor | 0 | −0.386 | 4.79 | ER |
IPI00222496a | Protein disulfide isomerase-associated 6 | 1 | −0.292 | 5.05 | ER |
IPI00230108 | Glucose-regulated protein, full insert sequence | 0 | −0.479 | 5.78 | ER |
IPI00319992 | 78 kDa glucose-regulated protein precursor | 0 | −0.481 | 5.07 | ER |
IPI00331322a | Microsomal glutathione S-transferase 1 | 3 | 0.14 | 9.67 | ER and mitochondrial outer membrane |
IPI00319973a | Membrane-associated progesterone receptor component 1 | 1 | −0.616 | 4.57 | ER, membrane bound |
IPI00109061 | Tubulin beta-4 chain homolog | 0 | −0.406 | 4.78 | Cytoplasmic |
IPI00110827 | Actin, alpha skeletal muscle | 0 | −0.232 | 5.23 | Cytoplasmic |
IPI00110850 | Actin, cytoplasmic 1 | 0 | −0.2 | 5.29 | Cytoplasmic |
IPI00111908 | Predicted: carbamoyl-phosphate synthetase 1 | 0 | −0.12 | 6.42 | Cytoplasmic |
IPI00117348 | Tubulin alpha-2 chain | 0 | −0.23 | 4.94 | Cytoplasmic |
IPI00117914 | Arginase 1 | 0 | −0.187 | 6.52 | Cytoplasmic |
IPI00134746 | Argininosuccinate synthase | 0 | −0.361 | 8.36 | Cytoplasmic |
IPI00117978a | Cytochrome c oxidase subunit IV isoform 1, mitochondrial precursor | 1 | −0.412 | 9.25 | Mitochondrial inner membrane |
IPI00315794a | Cytochrome b5 outer mitochondrial membrane isoform precursor | 1 | −0.602 | 4.79 | Mitochondrial outer membrane |
IPI00122547a | Voltage-dependent anion-selective channel protein 2 | 0 | −0.223 | 7.44 | Outer mitochondrial membrane |
IPI00124790a | Polyposis locus protein 1-like 1 | 3 | 0.375 | 6.82 | Integral to membrane |
IPI00138406a | Ras-related protein Rap-1A | 0 | −0.375 | 6.39 | Membrane |
IPI00121788 | Peroxiredoxin 1 | 0 | −0.221 | 8.26 | Microbody |
IPI00131366 | Keratin, type II cytoskeletal 6B | 0 | −0.488 | 8.32 | Sarcolemma |
aMembrane proteins with one or more predicted trans-membrane origins or validated by references.
Biological Networks Gene Ontology [
Biological pathways analysis based on KEGG pathway database was performed with an analysis toolkit—WebGestalt (
Significantly enriched KEGG pathways for mouse liver microsome proteins (
To ascertain the coverage of our dataset with the enriched pathways or biological processes, the KEGG search service was used to map our dataset on KEGG pathways. Two of the aforementioned enriched KEGG pathways (metabolism of xenobiotics and ribosome) were related to the well-known function and composition of the microsome (Figure
Proteins involved in KEGG pathways. (a) Metabolism of xenobiotics. (b) Ribosome.
KEGG pathway | Protein ID | Protein name | MS-identified proteins | |
---|---|---|---|---|
A. Metabolism of exnobiotics | EC:1.14.14.1 | IPI00128287 | Cytochrome P450 1A2 | + |
IPI00123964 | Cytochrome P450 2A5 | + | ||
IPI00116572 | Cytochrome P450 2D9 | + | ||
IPI00323908 | Cytochrome P450 2D10 | + | ||
IPI00321644 | Cytochrome P450 2D26 | + | ||
IPI00114779 | Cytochrome P450 2C38 | + | ||
IPI00308328 | Cytochrome P450 2F2 | + | ||
EC:2.5.1.18 | IPI00331322 | Microsomal glutathione S-transferase 1 | + | |
EC:1.1.1.1 | IPI00221400 | Alcohol dehydrogenase A chain | + | |
B. Ribosome | Small subunit | IPI00135640 | 26S protease regulatory subunit 8 | + |
IPI00125971 | 26S protease regulatory subunit S10B | + | ||
IPI00331092 | 40S ribosomal protein S4, X isoform | + | ||
IPI00116908 | Similar to 40s ribosomal protein S12 | + | ||
IPI00322562 | 40S ribosomal protein S14 | + | ||
IPI00465880 | 40S ribosomal protein S17 | + | ||
IPI00113241 | 40S ribosomal protein S19 | + | ||
IPI00123604 | 40S ribosomal protein SA | + | ||
IPI00314950 | 60S acidic ribosomal protein P0 | + | ||
Large subunit | IPI00331461 | 60S ribosomal protein L11 | + | |
IPI00849793 | 60S ribosomal protein L12 | + | ||
IPI00222546 | 60S ribosomal protein L22 | + | ||
IPI00139780 | 60S ribosomal protein L23 | + |
Metabolism of xenobiotics by cytochrome P450 pathway, and ribosome map views of identified proteins. The two enriched metabolic pathway maps were generated by KEGG, which incorporated the proteomic data into the KEGG pathway maps. All of the genes in mouse are colored; the genes contained in the protein dataset are red.
Proteome analysis of the cell membrane-bound organelles is a daunting task mainly because of (a) isolation of membrane that is free from nonconstituents and (b) solubilization of membrane proteins in a manner amenable to isoelectric focusing [
Hydrophobicity and relatively low abundance causes a challenge for proteomic technology to separate and identify membrane proteins. The hydrophobicity of proteins is frequently expressed as GRAVY scores (
Microsome-sealed vesicles could be converted into flat membrane sheets with cisternal contents that were released effectively with the treatment solution (100 mM Na2CO3; 0°C). It appears to be as effective as the low detergent procedure in selectively releasing microsomal content. In the current study, some proteins that were identified from Na2CO3-extracted fraction were classified as membrane associated mainly based on published reports, even though their predicted transmembrane domains (TMDs) did not suggest a membrane origin. The observations point out the fact that structure alone may not be the deciding factor, as far as the association of proteins with cell membrane is concerned. First, the proteins may be bound to the membrane simply to perform their functional obligations. Consequently, they could become part of complexes involving membrane proteins and may not depart from them easily under the conditions of sample preparation. For example, many enzymes were identified in the extracted membrane fraction, such as Cis-retinol androgen dehydrogenase 1 (short-chain dehydrogenase family). It is anchored to the ER membrane facing the cytoplasm by an N-terminal signaling sequence of 22 residues and takes part in the membrane-associated retinoid metabolism [
Two datasets from 1DE and 2DE are part of the complete protein composition of microsomes. A bioinformatics analysis of the two datasets combined offers more information. For an overview of the proteomic data and comprehending their biological importance, biological networks GO (BinGO) (
The KEGG pathway database integrates current knowledge on molecular interaction networks in biological processes. To gain a broad understanding of our dataset, WebGestalt (a web-based gene set analysis toolkit) was used to map the identified proteins to KEGG pathways. The results showed that 112 of the total proteins were associated with one or more KEGG pathways. Meanwhile, 97 of 112 target proteins (87%) fell into 34 KEGG pathways; they were specifically enriched (
Collectively, the bioinformatics analysis via enrichment analysis of GO annotation and KEGG pathways derived meaning from the proteomic data and assisted in the understanding of the function of liver at the subcellular level.
Mammalian liver microsome proteomes have been studied by several groups [
Comparison of the current study with the literature data [
Current study | Ref. [ | Ref. [ | Ref. [ | |
---|---|---|---|---|
Species | Mouse | Mouse | Mouse | Rat |
Sample | Liver microsome | Liver microsome | Liver microsome | ER, Golgi |
Pretreatment | None | Phenobarbital | Phenobarbital or 3-methylcholanthrene | None |
Sample preparation | Subfractionated by differential ultra-centrifugation + sucrose gradient centrifugation + Na2CO3 | Subfractionated by differential ultra-centrifugation | Differential ultracentrifugation | Subfractionated by differential ultra-centrifugation + density gradient centrifugation |
Protein separation | 2DE, 1DE | 1DE | 2DE | 1DE |
1D/2D-Gel Stain | Coomassie brilliant blue (2DE; 1DE) | — | Silver stain | — |
Protein identification | MS/MS | MS/MS | PMF | MS/MS |
Protein spots on 2D-Gel | 514 | — | 1100 | — |
Proteins identified in 2D-Gel | 183 | — | 139 | — |
Proteins identified in 1DE | 99 | 519 | — | 832 (ER) |
Proteins identified in 2-D LC | — | 1410 | — | — |
Proteins identified in 3-D LC | — | 3703 | — | — |
Total identified proteins | 259 | 4142 | Unspecified | 832 (ER) |
Membrane proteins | 2DE: 41 (41/183 = 22%) | 1DE: 138 (138/519 = 27%) | Unspecified | 183 (183/832 = 22%) |
Protein superfamily | ||||
P450 family members | 10 | 29 | 2 | 11 |
Ribosomal proteins | 13 | 16 | Unspecified | 45 |
UDP glycosyltransferases, UGTs | 6 | 8 | Unspecified | 3 |
Tubulins | 11 | 5 | Unspecified | 2 |
Short-chain dehydrogenase/reductase | 32 | 9 | Unspecified | 56 |
Protein disulfide isomerase | 2 | 4 | Unspecified | 1 |
Comparison of selected proteins between the current study and the literature data [
Protein | Current study | Ref. [ | Ref. [ | Ref. [ |
---|---|---|---|---|
P450 family members | 2D9, 2A5, 2C38, 1A2, 2D11, 2F2, 2D10, 2D26 | 2C37 | 17A1, 20A1, 2B2, 2J3, 4A1, 4A8, 4F1, 4F4, 4V3, 8B1, NA2 | |
GRP-170 | Hypoxia upregulated protein 1 | 170 kDa glucose regulated protein | — | |
Endoplasmin | Endoplasmin | Tumor rejection antigen gp96 | ||
Serotransferrin | Serotransferrin | Transferrin | — | |
78 kDa glucose-regulated protein | 78 kDa glucose-regulated protein | 78 kDa glucose-regulated protein | — | |
Stress-induced phosphoprotein 1 | — | Stress-induced phosphoprotein 1 | — | |
Calreticulin family | Calnexin | Calreticulin | — | |
Protein disulfide-isomerase | Protein disulfide-isomerase precursor (PDI) | Protein disulfide-isomerase precursor (PDI) | Similar to disulfide isomerase | |
Glucose-regulated protein similar to ER-60 protease | — | Glucose-regulated protein similar to ER-60 protease | — | |
Erp58 | — | Erp58 | — | |
Vitamin D-binding protein | — | Vitamin D-binding protein | — | |
Tubulins | Tubulin beta-4, alpha-1, alpha-2, alpha-4, beta-5, beta-2C, beta 2 | Tubulin alpha | Tubulin alpha 6 | |
Fibrinogen | — | Fibrinogen, gamma polypeptide | — | |
Serine protease inhibitor | — | Similar to serine protease inhibitor 1–4 | — | |
Argininosuccinate synthetase 1 | Argininosuccinate synthetase 1 | Argininosuccinate synthetase 1 | — | |
Interferon-inducible GTPase | — | Interferon-inducible GTPase | ||
Progesterone receptor membrane component | Progesterone receptor membrane component | Progesterone receptor membrane component | — | |
Major urinary protein 2 | Major urinary protein 2 | Major urinary protein 2 | — | |
Superoxide dismutase I | — | Superoxide dismutase I | — | |
Ribosomal proteins | 26S protease regulatory subunit 8, S10B; | Unspecified | 40S Ribosomal Protein S10, S12, S18, S20, S21, S23, S24, S25, S26, S27, S29, S30, S6, S9 | |
UDP glycosyltransferases, UGTs | UDP-glucuronosyltransferase 2B5, 2B36, 1A5 | Unspecified | UDP-Glucuronosyltransferase 1A7 | |
UDP-glucuronosyltransferase 1-1 precursor | UDP-Glucuronosyltransferase GTNA2 | |||
UDP-glucuronosyltransferase 1-2 precursor | ||||
Short-chain dehydrogenase/reductase | Glyceraldehyde-3-phosphate dehydrogenase | Unspecified | Glyceraldehyde 3-phosphate dehydrogenase | |
Oxidoreductase HTATIP2 NADH-ubiquinone oxidoreductase 24 kDa subunit | Unspecified | Oxidoreductase ero1-L endoplasmic oxidoreductase 1 Beta |
No protein list was obtained from [
The preparation of liver microsomes was optimized. The data presented here demonstrated that 1DE and 2DE are complementary approaches to analyze the intracellular microsomes that contain considerable numbers of highly hydrophobic membrane proteins. An integrated bioinformatics analysis of all of the microsome proteins identified with 1DE and 2DE can provide a relatively complete understanding of the protein composition and cellular function of the target microsome organelles. The information presented here will be useful for successful analysis of other membranous organelles. Our data will assist in understanding the function of liver and are an important reference for subsequent analysis of liver disease-related microsome proteins for biomarker discovery and mechanism clarification of a liver disease.
Acetonitrile
Biological Networks Gene Ontology
Dithiothreitol
One-dimensional gel electrophoresis
Two-dimensional gel electrophoresis
Endoplasmic reticulum
Gene ontology
Grand average of hydropathy
Human Liver Proteome Project
Isoelectric focusing
Kyoto Encyclopedia of Genes and Genomes
Liquid chromatography
Mass spectrometry
Tandem mass spectrometry
Quadrupole-time of flight
Reverse phase
Transmembrane domains.
This work was supported by China National Haman Liver Proteome Project (Grant no. 2004 BA711A18) and The National Basic Research Program of China (Grant No. 2011CB910704).