Integrated top-down and bottom-up proteomics mass spectrometry for the characterization of endogenous ribosomal protein heterogeneity

Ying Zhang; Qinghua Cai; Yuxiang Luo; Yu Zhang; Huilin Li

doi:10.1016/j.jpha.2022.11.003

Volume 13 Issue 1

Jan. 2023

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Ying Zhang, Qinghua Cai, Yuxiang Luo, Yu Zhang, Huilin Li. Integrated top-down and bottom-up proteomics mass spectrometry for the characterization of endogenous ribosomal protein heterogeneity[J]. Journal of Pharmaceutical Analysis, 2023, 13(1): 63-72. doi: 10.1016/j.jpha.2022.11.003

Citation:

Ying Zhang, Qinghua Cai, Yuxiang Luo, Yu Zhang, Huilin Li. Integrated top-down and bottom-up proteomics mass spectrometry for the characterization of endogenous ribosomal protein heterogeneity[J]. Journal of Pharmaceutical Analysis, 2023, 13(1): 63-72. doi: 10.1016/j.jpha.2022.11.003

Citation:

Ying Zhang, Qinghua Cai, Yuxiang Luo, Yu Zhang, Huilin Li. Integrated top-down and bottom-up proteomics mass spectrometry for the characterization of endogenous ribosomal protein heterogeneity[J]. Journal of Pharmaceutical Analysis, 2023, 13(1): 63-72. doi: 10.1016/j.jpha.2022.11.003

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Integrated top-down and bottom-up proteomics mass spectrometry for the characterization of endogenous ribosomal protein heterogeneity

doi: 10.1016/j.jpha.2022.11.003

a. School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, 510006, China;
b. Henan Engineering Laboratory for Mammary Bioreactor, School of Life Sciences, Henan University, Kaifeng, Henan, 475004, China;
c. The Shennong Laboratory, Zhengzhou, 450002, China;
d. Guangdong Key Laboratory of Chiral Molecule and Drug Discovery, School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou, 510006, China

Funds:

This work was supported in part by the National Natural Science Foundation of China (Grant Nos.: 91953102 and 81872836), Natural Science Foundation of Guangdong Province, China (Grant Nos.: 2019A1515011265 and 2022A1515010965), the Fundamental Research Funds for Sun Yat-sen University, China (Grant No.: 19ykzd26), and Open Project Funding of the State Key Laboratory of Crop Stress Adaptation and Improvement (Grant No.: 2020KF05). Huilin Li would like to thank the Pearl River Talent Recruitment Program for support.

Received Date: Jul. 26, 2022
Accepted Date: Nov. 08, 2022
Rev Recd Date: Nov. 07, 2022
Publish Date: Nov. 14, 2022

Abstract

Abstract

Ribosomes are abundant, large RNA-protein complexes that are the sites of all protein synthesis in cells. Defects in ribosomal proteins (RPs), including proteoforms arising from genetic variations, alternative splicing of RNA transcripts, post-translational modifications and alterations of protein expression level, have been linked to a diverse range of diseases, including cancer and aging. Comprehensive characterization of ribosomal proteoforms is challenging but important for the discovery of potential disease biomarkers or protein targets. In the present work, using E. coli 70S RPs as an example, we first developed a top-down proteomics approach on a Waters Synapt G2 Si mass spectrometry (MS) system, and then applied it to the HeLa 80S ribosome. The results were complemented by a bottom-up approach. In total, 50 out of 55 RPs were identified using the top-down approach. Among these, more than 30 RPs were found to have their N-terminal methionine removed. Additional modifications such as methylation, acetylation, and hydroxylation were also observed, and the modification sites were identified by bottom-up MS. In a HeLa 80S ribosomal sample, we identified 98 ribosomal proteoforms, among which multiple truncated 80S ribosomal proteoforms were observed, the type of information which is often overlooked by bottom-up experiments. Although their relevance to diseases is not yet known, the integration of top-down and bottom-up proteomics approaches paves the way for the discovery of proteoform-specific disease biomarkers or targets.
- Ribosomal proteins,
- Top-down MS,
- Bottom-up MS,
- Proteoforms

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References(80)

References

S. Pechmann, F. Willmund, J. Frydman, The ribosome as a hub for protein quality control, Mol. Cell 49 (2013) 411-421

K. Ikeuchi, T. Izawa, T. Inada, Recent progress on the molecular mechanism of quality controls induced by ribosome stalling, Front. Genet. 9 (2019), 743

J.C. Bowman, A.S. Petrov, M. Frenkel-Pinter, et al., Root of the tree: the significance, evolution, and origins of the ribosome, Chem. Rev. 120 (2020) 4848-4878

C. Pena, E. Hurt, V.G. Panse, Eukaryotic ribosome assembly, transport and quality control, Nat. Struct. Mol. Biol. 24 (2017) 689-699

A.S. Petrov, B. Gulen, A.M. Norris, et al., History of the ribosome and the origin of translation, Proc. Natl. Acad. Sci. USA 112 (2015) 15396-15401

A.V. Korobeinikova, M.B. Garber, G.M. Gongadze, Ribosomal proteins: structure, function, and evolution, Biochemistry (Mosc.) 77 (2012) 562-574

J. Kang, N. Brajanovski, K.T. Chan, et al., Ribosomal proteins and human diseases: molecular mechanisms and targeted therapy, Signal Transduct. Targeted Ther. 6 (2021), 323

Z. Turi, M. Lacey, M. Mistrik, et al., Impaired ribosome biogenesis: mechanisms and relevance to cancer and aging, Aging 11 (2019) 2512-2540

I. Boria, P. Quarello, F. Avondo, et al., A new database for ribosomal protein genes which are mutated in Diamond-Blackfan anemia, Hum. Mutat. 29 (2008) E263-E270

H.T. Gazda, M.R. Sheen, A. Vlachos, et al., Ribosomal protein L5 and L11 mutations are associated with cleft palate and abnormal thumbs in Diamond-Blackfan anemia patients, Am. J. Hum. Genet. 83 (2008) 769-780

I. Orsolic, S. Bursac, D. Jurada, et al., Cancer-associated mutations in the ribosomal protein L5 gene dysregulate the HDM2/p53-mediated ribosome biogenesis checkpoint, Oncogene 39 (2020) 3443-3457

R.J. Weatheritt, T. Sterne-Weiler, B.J. Blencowe, The ribosome-engaged landscape of alternative splicing, Nat. Struct. Mol. Biol. 23 (2016) 1117-1123

Y. Zhang, J. Qian, C. Gu, et al., Alternative splicing and cancer: a systematic review, Signal Transduct. Targeted Ther. 6 (2021), 78

D. Simsek, M. Barna, An emerging role for the ribosome as a nexus for post-translational modifications, Curr. Opin. Cell Biol. 45 (2017) 92-101

C. Petibon, M.M. Ghulam, M. Catala, et al., Regulation of ribosomal protein genes: an ordered anarchy, Wiley Interdiscip Rev RNA 12 (2021), e1632

L.M. Smith, N.L. Kelleher, Proteoform: a single term describing protein complexity, Nat. Methods 10 (2013) 186-187

R. Aebersold, J.N. Agar, I.J. Amster, et al., How many human proteoforms are there?, Nat. Chem. Biol. 14 (2018) 206-214

L.M. Smith, N.L. Kelleher, Proteoforms as the next proteomics currency, Science 359 (2018) 1106-1107

G. Millan-Zambrano, A. Burton, A.J. Bannister, et al., Histone post-translational modifications-cause and consequence of genome function, Nat. Rev. Genet. 23 (2022) 563-580

R. Aebersold, M. Mann, Mass spectrometry-based proteomics, Nature 422 (2003) 198-207

J.R. Yates, C.I. Ruse, A. Nakorchevsky, Proteomics by mass spectrometry: approaches, advances, and applications, Annu. Rev. Biomed. Eng. 11 (2009) 49-79

J.B. Muller, P.E. Geyer, A.R. Colaco, et al., The proteome landscape of the kingdoms of life, Nature 582 (2020) 592-596

R. Aebersold, M. Mann, Mass-spectrometric exploration of proteome structure and function, Nature 537 (2016) 347-355

B.T. Chait, Chemistry. mass spectrometry: bottom-up or top-down?, Science 314 (2006) 65-66

N.L. Kelleher, Peer reviewed: top-down proteomics, Anal. Chem. 76 (2004) 196A-203A

J.C. Tran, L. Zamdborg, D.R. Ahlf, et al., Mapping intact protein isoforms in discovery mode using top-down proteomics, Nature 480 (2011) 254-258

T.K. Toby, L. Fornelli, N.L. Kelleher, Progress in top-down proteomics and the analysis of proteoforms, Annu. Rev. Anal. Chem. Palo Alto Calif 9 (2016) 499-519

L.M. Smith, J.N. Agar, J. Chamot-Rooke, et al., The human proteoform project: defining the human proteome, Sci. Adv. 7 (2021), eabk0734

A.D. Catherman, O.S. Skinner, N.L. Kelleher, Top down proteomics: facts and perspectives, Biochem. Biophys. Res. Commun. 445 (2014) 683-693

J.A. Melby, D.S. Roberts, E.J. Larson, et al., Novel strategies to address the challenges in top-down proteomics, J. Am. Soc. Mass Spectrom. 32 (2021) 1278-1294

S.J. Hardy, C.G. Kurland, P. Voynow, et al., The ribosomal proteins of Escherichia coli. I. purification of the 30S ribosomal proteins, Biochemistry 8 (1969) 2897-2905

T.J. El-Baba, S.A. Raab, R.P. Buckley, et al., Thermal analysis of a mixture of ribosomal proteins by vT-ESI-MS: toward a parallel approach for characterizing the stabilitome, Anal. Chem. 93 (2021) 8484-8492

B. Burton, M.T. Zimmermann, R.L. Jernigan, et al., A computational investigation on the connection between dynamics properties of ribosomal proteins and ribosome assembly, PLoS Comput. Biol. 8 (2012), e1002530

A.T. Gudkov, The L7/L12 ribosomal domain of the ribosome: structural and functional studies, FEBS Lett. 407 (1997) 253-256

L. Tsiatsiani, A.J.R. Heck, Proteomics beyond trypsin, FEBS J. 282 (2015) 2612-2626

C. Chabanet, M. Yvon, Prediction of peptide retention time in reversed-phase high-performance liquid chromatography, J. Chromatogr. 599 (1992) 211-225

C.N. Chang, M. Schwartz, F.N. Chang, Identification and characterization of a new methylated amino acid in ribosomal protein L33 of Escherichia coli, Biochem. Biophys. Res. Commun. 73 (1976) 233-239

E.J. Dupree, M. Jayathirtha, H. Yorkey, et al., A critical review of bottom-up proteomics: the good, the bad, and the future of this field, Proteomes 8 (2020), 14

P.T. Wingfield, N-terminal methionine processing, Curr. Protoc. Protein Sci. 88 (2017) 6.14.1-6.14.3

H. Demirci, S.T. Gregory, A.E. Dahlberg, et al., Multiple-site trimethylation of ribosomal protein L11 by the PrmA methyltransferase, Structure 16 (2008) 1059-1066

M.J. Suh, D.M. Hamburg, S.T. Gregory, et al., Extending ribosomal protein identifications to unsequenced bacterial strains using matrix-assisted laser desorption/ionization mass spectrometry, Proteomics 5 (2005) 4818-4831

W.E. Running, S. Ravipaty, J.A. Karty, et al., A top-down/bottom-up study of the ribosomal proteins of Caulobacter crescentus, J. Proteome Res. 6 (2007) 337-347

J. Lhoest, C. Colson, Cold-sensitive ribosome assembly in an Escherichia coli mutant lacking a single methyl group in ribosomal protein L3, Eur. J. Biochem. 121 (1981) 33-37

D.M. Cameron, S.T. Gregory, J. Thompson, et al., Thermus thermophilus L11 methyltransferase, PrmA, is dispensable for growth and preferentially modifies free ribosomal protein L11 prior to ribosome assembly, J. Bacteriol. 186 (2004) 5819-5825

N. Brot, W.P. Tate, C.T. Caskey, et al., The requirement for ribosomal proteins L7 and L12 in peptide-chain termination, Proc. Natl. Acad. Sci. USA 71 (1974) 89-92

A.V. Oleinikov, G.G. Jokhadze, R.R. Traut, A single-headed dimer of Escherichia coli ribosomal protein L7/L12 supports protein synthesis, Proc. Natl. Acad. Sci. USA 95 (1998) 4215-4218

I. Pettersson, C.G. Kurland, Ribosomal protein L7/L12 is required for optimal translation, Proc. Natl. Acad. Sci. USA 77 (1980) 4007-4010

F.N. Chang, Temperature-dependent variation in the extent of methylation of ribosomal proteins L7 and L12 in Escherichia coli, J. Bacteriol. 135 (1978) 1165-1166

W. Ge, A. Wolf, T. Feng, et al., Oxygenase-catalyzed ribosome hydroxylation occurs in prokaryotes and humans, Nat. Chem. Biol. 8 (2012) 960-962

C. DeBoever, Y. Tanigawa, M.E. Lindholm, et al., Medical relevance of protein-truncating variants across 337,205 individuals in the UK Biobank study, Nat. Commun. 9 (2018), 1612

J. Vlasak, R. Ionescu, Fragmentation of monoclonal antibodies, mAbs 3 (2011) 253-263

W. Yu, J.E. Vath, M.C. Huberty, et al., Identification of the facile gas-phase cleavage of the Asp-Pro and Asp-Xxx peptide bonds in matrix-assisted laser desorption time-of-flight mass spectrometry, Anal. Chem. 65 (1993) 3015-3023

M.I. Lerman, A.S. Spirin, L.P. Gavrilova, et al., Studies on the structure of ribosomes: II. Stepwise dissociation of protein from ribosomes by caesium chloride and the re-assembly of ribosome-like particles, J. Mol. Biol. 15 (1966) 268-281

G.M. Blaha, S. Diggs, T.K. Tam, et al., The Effects of Ribosomal Proteins uS2, uS3, and uS4 on Transcription, 2022. https://doi.org/10.1096/fasebj.2022.36.S1.L7615

H. Khatter, A.G. Myasnikov, S.K. Natchiar, et al., Structure of the human 80S ribosome, Nature 520 (2015) 640-645

M. van de Waterbeemd, S. Tamara, K.L. Fort, et al., Dissecting ribosomal particles throughout the kingdoms of life using advanced hybrid mass spectrometry methods, Nat. Commun. 9 (2018), 2493

M.A. Rivas, M. Pirinen, D.F. Conrad, et al., Human genomics. Effect of predicted protein-truncating genetic variants on the human transcriptome, Science 348 (2015) 666-669

A.D. Neverov, I.I. Artamonova, R.N. Nurtdinov, et al., Alternative splicing and protein function, BMC Bioinf. 6 (2005), 266

X. Xie, P. Guo, H. Yu, et al., Ribosomal proteins: insight into molecular roles and functions in hepatocellular carcinoma, Oncogene 37 (2018) 277-285

W. Wang, S. Nag, X. Zhang, et al., Ribosomal proteins and human diseases: pathogenesis, molecular mechanisms, and therapeutic implications, Med. Res. Rev. 35 (2015) 225-285

S. Challa, B.R. Khulpateea, T. Nandu, et al., Ribosome ADP-ribosylation inhibits translation and maintains proteostasis in cancers, Cell 184 (2021) 4531-4546.e26

A. Pecoraro, M. Pagano, G. Russo, et al., Ribosome biogenesis and cancer: overview on ribosomal proteins, Int. J. Mol. Sci. 22 (2021), 5496

J. Xie, W. Zhang, X. Liang, et al., RpL32 promotes lung cancer progression by facilitating p53 degradation, Mol. Ther. Nucleic Acids 21 (2020) 75-85

C. Li, M. Ge, D. Chen, et al., RPL21 siRNA blocks proliferation in pancreatic cancer cells by inhibiting DNA replication and inducing G1 arrest and apoptosis, Front. Oncol. 10 (2020), 1730

R.Y. Ebright, S. Lee, B.S. Wittner, et al., Deregulation of ribosomal protein expression and translation promotes breast cancer metastasis, Science 367 (2020) 1468-1473

S. N. Slimane, V. Marcel, T. Fenouil, et al., Ribosome biogenesis alterations in colorectal cancer, Cells 9 (2020), 2361

A. Bee, Y.Q. Ke, S. Forootan, et al., Ribosomal protein L19 is a prognostic marker for human prostate cancer, Clin. Cancer Res. 12 (2006) 2061-2065

S. Muro, Y. Miyake, H. Kato, et al., Serum anti-60S ribosomal protein L29 antibody as a novel prognostic marker for unresectable pancreatic cancer, Digestion 91 (2015) 164-173

C. Li, M. Ge, Y. Yin, et al., Silencing expression of ribosomal protein L26 and L29 by RNA interfering inhibits proliferation of human pancreatic cancer PANC-1 cells, Mol. Cell. Biochem. 370 (2012) 127-139

T. Ota, Y. Suzuki, T. Nishikawa, et al., Complete sequencing and characterization of 21,243 full-length human cDNAs, Nat. Genet. 36 (2004) 40-45

A. Labriet, E. Levesque, E. Cecchin, et al., Germline variability and tumor expression level of ribosomal protein gene RPL28 are associated with survival of metastatic colorectal cancer patients, Sci. Rep. 9 (2019), 13008

M.I. Yavor, T.V. Pomozov, S.N. Kirillov, et al., High performance gridless ion mirrors for multi-reflection time-of-flight and electrostatic trap mass analyzers, Int. J. Mass Spectrom. 426 (2018) 1-11

K. Richardson, J. Hoyes, A novel multipass oa-TOF mass spectrometer, Int. J. Mass Spectrom. 377 (2015) 309-315

X. Shen, T. Xu, B. Hakkila, et al., Capillary zone electrophoresis-electron-capture collision-induced dissociation on a quadrupole time-of-flight mass spectrometer for top-down characterization of intact proteins, J. Am. Soc. Mass Spectrom. 32 (2021) 1361-1369

M.R. Mehaffey, Q. Xia, J.S. Brodbelt, Uniting native capillary electrophoresis and multistage ultraviolet photodissociation mass spectrometry for online separation and characterization of Escherichia coli ribosomal proteins and protein complexes, Anal. Chem. 92 (2020) 15202-15211

K.A. Brown, C. Anderson, L. Reilly, et al., Proteomic analysis of the functional inward rectifier potassium channel (kir) 2.1 reveals several novel phosphorylation sites, Biochemistry 60 (2021) 3292-3301

D.S. Roberts, B. Chen, T.N. Tiambeng, et al., Reproducible large-scale synthesis of surface silanized nanoparticles as an enabling nanoproteomics platform: enrichment of the human heart phosphoproteome, Nano Res. 12 (2019) 1473-1481

L.V. Schaffer, R.J. Millikin, M.R. Shortreed, et al., Improving proteoform identifications in complex systems through integration of bottom-up and top-down data, J. Proteome Res. 19 (2020) 3510-3517

A.J. Cesnik, M.R. Shortreed, L.V. Schaffer, et al., Proteoform Suite: Software for constructing, quantifying, and visualizing proteoform families, J. Proteome Res. 17 (2018) 568-578

D.B. Lima, M. Dupre, M. Duchateau, et al., ProteoCombiner: integrating bottom-up with top-down proteomics data for improved proteoform assessment, Bioinformatics 37 (2021) 2206-2208

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