Mass Spectrometry-Based Proteomic and Immunoproteomic Analyses of the Candida albicans Hyphal Secretome Reveal Diagnostic Biomarker Candidates for Invasive Candidiasis
Abstract
:1. Introduction
2. Materials and Methods
2.1. Study Population and Serum Samples
2.2. Isolation of C. albicans Hyphal Secreted Proteins
2.3. Protein Separation by Sodium Dodecyl Sulfate-Polyacrylamide gel Electrophoresis (SDS-PAGE)
2.4. Protein Identification by Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS) Analysis
2.5. Indirect Enzyme-Linked Immunosorbent Assay (ELISA)
2.6. Immunoproteomic Analysis or SERPA
2.7. Bioinformatic Analysis
3. Results
3.1. Isolation of the C. albicans Hyphal Secretome
3.2. Gel-Free LC-MS/MS Analysis of the C. albicans Hyphal Secretome
3.3. Serologic Responses to the C. albicans Hyphal Secretome in ncIC, cIC and Non-IC Patients
4. Discussion
4.1. The C. albicans Hyphal Secretome Comprises Proteins Involved in Interaction with Host and Immunogenic Proteins
4.2. The C. albicans Hyphal Immunosecretome Recognized by IgG Antibodies Differs between IC and Non-IC Patients
4.3. IgG Antibodies to Bgl2, Eno1, Pgk1, Glx3, Pra1, Sap5 and Tdh3 Are IC Diagnostic Biomarker Candidates
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Cellular Location | No. of Identified Proteins | Protein Names |
---|---|---|
Extracellular region, cell surface, cell wall, and cell periphery | 37 | Ade8, Als1, Bgl2, Cht1, Cht3, Cip1, Coi1, Ece1, Ecm33, Gcy1, Grp2, Hex1, Kre9, Mal2, Mdg1, Met15, Mp65, Nit3, Ofr1, Orf19.1394, Orf19.3053, Orf19.6809, Orf19.6867, Orf19.7322, Pga12, Pga4, Pra1, Pst2, Rbe1, Rbp1, Rhd3, Sah1, Sap10, Sap8, Slk19, Sol3, Xyl2 |
Intracellular region | 107 | Ade17, Aha1, Ams1, Anb1, Arf2, Arg1, Arg3, Arg4, Aro2, Asc1, Bfr1, Bud7, Cit1, Cmd1, Cpr6, Crm1, Cyc1, Dtd2, Ecm4, Egd1, Eif4e, Erg10, Erg20, Etr1, Fbp1, Fum11, Gcv3, Grs1, Grx3, Het1, His1, Hom2, Hta3, Hxk2, Idh1, Kex1, Krs1, Lat1, Lsc1, Lys21, Mca1, Mcr1, Mdh1, Mmd1, Mrf1, Mxr1, Npt1, Ntf2, Orf19.1355, Orf19.1448.1, Orf19.1738.1, Orf19.1815, Orf19.2930, Orf19.3319, Orf19.3681, Orf19.3932, Orf19.4150, Orf19.4382, Orf19.4898, Orf19.518, Orf19.5322, Orf19.5943.1, Orf19.5961, Orf19.6559, Orf19.6596, Orf19.6701, Orf19.6872, Orf19.7152, Orf19.7196, Orf19.7214, Orf19.7297, Orf19.7330, Orf19.7368, Orf19.7404, Orf19.7531, Orf19.7578, Orf19.86, Orf19.904, Pol30, Rdi1, Rnr21, Rpl10a, Rpl12, Rpl30, Rpp0, Rps12, Rps19a, Rps21b, Rps22a, Sbp1, Sec14, Skp1, Sno1, Snz1, Sod2, Sod3, Sub2, Sui1, Tfs1, Thi4, Tif1, Tub1, Tub2, Tup1, Uba1, Yhb1, Ykt6 |
Both locations | 93 | Aat21, Abp1, Acb1, Aco1, Ahp1, Ape3, Atp1, Atp2, Bmh1, Cam1, Cat1, Cdc19, Cdc3, Cof1, Cyp1, Cyp5, Eft2, Egd2, Emp24, Eng1, Eno1, Fba1, Fdh1, Gdh3, Gnd1, Gpd2, Gph1, Gpm1, Gsp1, Hem13, Hsp12, Hsp70, Hsp90, Ino1, Ipp1, Kel1, Leu2, Lpd1, Lsp1, Mdh1-1, Met6, Mir1, Mlc1, Mnt1, Orf19.1085, Orf19.1862, Orf19.1946, Orf19.3915, Orf19.4395, Orf19.4597, Orf19.5342, Pdc11, Pet9, Pfy1, Pgi1, Pgk1, Phr2, Pin3, Pmm1, Por1, Prx1, Pst3, Rho1, Rib3, Rpl14, Rpl6, Rps20, Sam2, Sap4, Sap5, Sec4, Sim1, Smt3, Sod1, Spe3, Ssa2, Ssb1, Tal1, Tdh3, Tma19, Tpi1, Tpm2, Trr1, Trx1, Tsa1, Ttr1, Ubi3, Ugp1, Utr2, Vps21, Xog1, Ynk1, Ypt1 |
Immunoreactive C. albicans Hyphal Secreted Proteins | IgG Antibody- Reactivity Levels a | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Protein Name | Protein Description | LC-MS/MS | MALDI-TOF-MS | ncIC | cIC | Non- IC | ||||
No. of Peptides | Ranking According to NSAF | NSAF | No. of Matched/ Unmatched Peptides | Mascot Score | % of Sequence Coverage | |||||
Bgl2 | Cell wall 1,3-beta- glucosyltransferase | 8 | 55 | 0.004 | 10/58 | 213 | 20 | +++ | +++ | − |
Eno1 | Enolase | 30 | 1 | 0.08 | 9/61 | 157 | 32 | ++++ | +++ | + |
Glx3 | Glutathione- independent glyoxalase | NI b | NI b | NI b | 12/56 | 276 | 61 | +++ | +++ | − |
Pgk1 | Phosphoglycerate kinase | 38 | 2 | 0.04 | 21/47 | 460 | 48 | +++ | ++ | + |
Pra1 | pH regulated antigen | 10 | 12 | 0.01 | 11/57 | 240 | 29 | ND | +++++ | +++ |
9/59 | 218 | 16 | ||||||||
10/58 | 229 | 20 | ||||||||
7/61 | 75 | 8 | ||||||||
7/60 | 115 | 11 | ||||||||
5/63 | 96 | 7 | ||||||||
Sap5 | Secreted aspartyl proteinase | 26 | 3 | 0.03 | 14/54 | 272 | 39 | ++ | +++ | ++ |
28/43 | 815 | 60 | ||||||||
39/38 | 1220 | 67 | ||||||||
6/62 | 60 | 18 | ||||||||
Tdh3 | Glyceraldehyde-3- phosphate | 27 | 9 | 0.02 | 16/53 | 140 | 51 | − | + | − |
Standard Name | Description | NetNGlyc 1.0 Server Prediction | NetNGlyc 4.0 Server Prediction | NSAF | No. Peptides | Mr |
---|---|---|---|---|---|---|
Rbt4 | Pry family protein | no | yes | 0.19 | 11 | 37.4 |
Mp65 | Cell surface mannoprotein | no | yes | 0.15 | 13 | 39.3 |
Sun41 | Cell wall glycosidase | yes | yes | 0.12 | 11 | 43.7 |
Tos1 | Protein similar to alpha agglutinin anchor subunit | no | yes | 0.10 | 15 | 49.4 |
Ecm33 | GPI-anchored cell wall protein | yes | yes | 0.03 | 6 | 43.5 |
Cht3 | Major chitinase | no | yes | 0.03 | 7 | 60.0 |
Pra1 | Cell surface protein that sequesters zinc from host tissue | yes | yes | 0.02 | 7 | 33.1 |
Glx3 | Glutathione-independent glyoxalase | yes | no | 0.02 | 12 | 25.8 |
Sim1 | Adhesin-like protein | yes | yes | 0.01 | 9 | 39.4 |
Scw11 | Cell wall protein | yes | yes | 0.01 | 10 | 54.4 |
Standard Namea | Description a | Cell Localizationb | Signal Peptide a | References c |
---|---|---|---|---|
Aco1 | Aconitase | Shared | No | [25,77] |
Ade17 | 5-Aminoimidazole-4-carboxamide ribotide transformylase | Intracellular | No | [25] |
Als1 | Cell-surface adhesin | Extracellular | yes | [54] |
Asc1 | 40S ribosomal subunit similar to G-beta subunits | Intracellular | No | [25] |
Atp1 | ATP synthase alpha subunit | Shared | No | [25] |
Atp2 | F1 beta subunit of F1F0 ATPase complex | Shared | No | [25] |
Bgl2 | Cell wall 1,3-beta-glucosyltransferase | Extracellular | Yes | [24] |
Cdc19 | Pyruvate kinase at yeast cell surface | Shared | No | [77] |
Ece1 | Extent of cell elongation protein | Extracellular | yes | [54] |
Ecm33 | GPI-anchored cell wall protein | Extracellular | yes | [54] |
Eft2 | Elongation factor 2 | Shared | No | [77] |
Eno1 | Enolase | Shared | No | [50,53,78] |
Fba1 | Fructose-bisphosphate aldolase | Shared | No | [25] |
Gnd1 | 6-phosphogluconate dehydrogenase | Shared | No | [29] |
Gpm1 | Phosphoglycerate mutase | Shared | No | [25,77] |
Grp2 | NAD(H)-linked methylglyoxal oxidoreductase involved in regulation of methylglyoxal and pyruvate levels | Extracellular | No | [25] |
Hem13 | Coproporphyrinogen III oxidase | Shared | No | [25] |
Hsp70 | Putative Hsp70 chaperone | Shared | No | [25,53] |
Hsp90 | Essential chaperone | Shared | No | [25,53] |
Hxk2 | Hexokinase II | Intracellular | No | [25] |
Ino1 | Inositol-1-phosphate synthase | Shared | No | [25] |
Ipp1 | Putative inorganic pyrophosphatase | Shared | No | [25] |
Mdh1 | Mitochondrial malate dehydrogenase | Intracellular | Yes | [25] |
Mdh1-1 | Predicted malate dehydrogenase precursor | Shared | No | |
Met6 | 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase (methionine synthase) | Shared | No | [49,77] |
Mp65 | Cell surface mannoprotein | Extracellular | Yes | [79,80] |
Msi3 | Essential HSP70 family protein | Intracellular | No | |
orf19.7196 | Putative vacuolar protease | Intracellular | Yes | [33] |
orf19.7214 | Glucan 1,3-beta-glucosidase | Intracellular | No | [54] |
Pdc11 | Pyruvate decarboxylase | Shared | No | [25] |
Pga4 | GPI-anchored cell surface protein | Extracellular | Yes | [54] |
Pgi1 | Glucose-6-phosphate isomerase | Shared | No | [25] |
Pgk1 | Phosphoglycerate kinase | Shared | No | [50,53] |
Por1 | Mitochondrial outer membrane porin | Shared | No | [25] |
Pra1 | pH regulated antigen | Extracellular | Yes | [54,81] |
Prx1 | Thioredoxin peroxidase | Shared | No | [33] |
Sah1 | S-adenosyl-L-homocysteine hydrolase | Extracellular | No | [25] |
Slk19 | Alkaline-induced protein of plasma membrane | Extracellular | no | [54] |
Ssa2 | HSP70 family chaperone | Shared | No | [25,54] |
Ssb1 | HSP70 family heat shock protein | Shared | No | [25,32] |
Tal1 | Transaldolase | Shared | No | [33] |
Tdh3 | NAD-linked glyceraldehyde-3-phosphate dehydrogenase | Shared | No | [25,50,69] |
Tif | Translation initiation factor | Intracellular | No | [25] |
Tpi1 | Triose-phosphate isomerase | Shared | No | [25,32] |
Tsa1 | TSA/alkyl hydroperoxide peroxidase C (AhPC) family protein | Shared | No | [27] |
Utr2 | Putative GPI anchored cell wall glycosidase | Extracellular | yes | [54] |
Xog1 | Exo-1,3-beta-glucanase | Shared | Yes | [54] |
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Vaz, C.; Pitarch, A.; Gómez-Molero, E.; Amador-García, A.; Weig, M.; Bader, O.; Monteoliva, L.; Gil, C. Mass Spectrometry-Based Proteomic and Immunoproteomic Analyses of the Candida albicans Hyphal Secretome Reveal Diagnostic Biomarker Candidates for Invasive Candidiasis. J. Fungi 2021, 7, 501. https://0-doi-org.brum.beds.ac.uk/10.3390/jof7070501
Vaz C, Pitarch A, Gómez-Molero E, Amador-García A, Weig M, Bader O, Monteoliva L, Gil C. Mass Spectrometry-Based Proteomic and Immunoproteomic Analyses of the Candida albicans Hyphal Secretome Reveal Diagnostic Biomarker Candidates for Invasive Candidiasis. Journal of Fungi. 2021; 7(7):501. https://0-doi-org.brum.beds.ac.uk/10.3390/jof7070501
Chicago/Turabian StyleVaz, Catarina, Aida Pitarch, Emilia Gómez-Molero, Ahinara Amador-García, Michael Weig, Oliver Bader, Lucía Monteoliva, and Concha Gil. 2021. "Mass Spectrometry-Based Proteomic and Immunoproteomic Analyses of the Candida albicans Hyphal Secretome Reveal Diagnostic Biomarker Candidates for Invasive Candidiasis" Journal of Fungi 7, no. 7: 501. https://0-doi-org.brum.beds.ac.uk/10.3390/jof7070501