Abstract
RNA nanotechnology often feature protein RNA complexes. The interaction between proteins and large RNAs are difficult to study using traditional structure-based methods like NMR or X-ray crystallography. RCAP, an approach that uses reversible-cross-linking affinity purification method coupled with mass spectrometry, has been developed to map regions within proteins that contact RNA. This chapter details how RCAP is applied to map protein–RNA contacts within virions.
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References
Kao CC, Ni P, Hema M, Huang X, Dragnea B (2011) The coat protein leads the way: an update on basic and applied studies with the Brome mosaic virus coat protein. Mol Plant Pathol 12(4):403–412. doi:10.1111/j.1364-3703.2010.00678.x
Noueiry AO, Ahlquist P (2003) Brome mosaic virus RNA replication: revealing the role of the host in RNA virus replication. Annu Rev Phytopathol 41:77–98. doi:10.1146/annurev.phyto. 41.052002.095717
Running WE, Ni P, Kao CC, Reilly JP (2012) Chemical reactivity of brome mosaic virus capsid protein. J Mol Biol 423(1):79–95. doi:10.1016/j.jmb.2012.06.031
Lucas RW, Larson SB, McPherson A (2002) The crystallographic structure of brome mosaic virus. J Mol Biol 317(1):95–108. doi:10.1006/jmbi.2001.5389
Ni P, Wang Z, Ma X, Das NC, Sokol P, Chiu W, Dragnea B, Hagan M, Kao CC (2012) An examination of the electrostatic interactions between the N-terminal tail of the Brome Mosaic Virus coat protein and encapsidated RNAs. J Mol Biol 419(5):284–300. doi:10.1016/j.jmb.2012.03.023
Vaughan R, Running W, Qi R, Kao CC (2012) Mapping protein-RNA interactions. Virus Adapt Treat 4:29–41. doi:http://dx.doi.org/10.2147/VAAT.S31299
Leitner A, Walzthoeni T, Kahraman A, Herzog F, Rinner O, Beck M, Aebersold R (2010) Probing native protein structures by chemical cross-linking, mass spectrometry, and bioinformatics. Mol Cell Proteomics 9(8):1634–1649. doi:10.1074/mcp. R000001-MCP201
Park AY, Robinson CV (2011) Protein-nucleic acid complexes and the role of mass spectrometry in their structure determination. Crit Rev Biochem Mol Biol 46(2):152–164. doi:10.3109/10409238.2011.559451
Kim YC, Russell WK, Ranjith-Kumar CT, Thomson M, Russell DH, Kao CC (2005) Functional analysis of RNA binding by the hepatitis C virus RNA-dependent RNA polymerase. J Biol Chem 280(45):38011–38019. doi:10.1074/jbc.M508145200, M508145200 [pii]
Hema M, Murali A, Ni P, Vaughan RC, Fujisaki K, Tsvetkova I, Dragnea B, Kao CC (2010) Effects of amino-acid substitutions in the Brome mosaic virus capsid protein on RNA encapsidation. Mol Plant Microbe Interact 23(11):1433–1447. doi:10.1094/MPMI-05-10-0118
Hwang J, Huang L, Cordek DG, Vaughan R, Reynolds SL, Kihara G, Raney KD, Kao CC, Cameron CE (2010) Hepatitis C virus nonstructural protein 5A: biochemical characterization of a novel structural class of RNA-binding proteins. J Virol 84(24):12480–12491. doi:10.1128/JVI. 01319-10
Ranjith-Kumar CT, Duffy KE, Jordan JL, Eaton-Bassiri A, Vaughan R, Hoose SA, Lamb RJ, Sarisky RT, Kao CC (2008) Single-stranded oligonucleotides can inhibit cytokine production induced by human Toll-like receptor 3. Mol Cell Biol 28(14):4507–4519. doi:10.1128/MCB. 00308-08
Ranjith-Kumar CT, Murali A, Dong W, Srisathiyanarayanan D, Vaughan R, Ortiz-Alacantara J, Bhardwaj K, Li X, Li P, Kao CC (2009) Agonist and antagonist recognition by RIG-I, a cytoplasmic innate immunity receptor. J Biol Chem 284(2):1155–1165. doi:10.1074/jbc.M806219200
Vaughan R, Fan B, You JS, Kao CC (2012) Identification and functional characterization of the nascent RNA contacting residues of the hepatitis C virus RNA-dependent RNA polymerase. RNA 18(8):1541–1552. doi:10.1261/rna.031914.111
Yi G, Vaughan RC, Yarbrough I, Dharmaiah S, Kao CC (2009) RNA binding by the brome mosaic virus capsid protein and the regulation of viral RNA accumulation. J Mol Biol 391(2):314–326. doi:10.1016/j.jmb.2009.05.065, doi:S0022-2836(09)00647-0 [pii]
Jones S, Daley DT, Luscombe NM, Berman HM, Thornton JM (2001) Protein-RNA interactions: a structural analysis. Nucleic Acids Res 29(4):943–954
Metz B, Kersten GF, Hoogerhout P, Brugghe HF, Timmermans HA, de Jong A, Meiring H, ten Hove J, Hennink WE, Crommelin DJ, Jiskoot W (2004) Identification of formaldehyde-induced modifications in proteins: reactions with model peptides. J Biol Chem 279(8):6235–6243. doi:10.1074/jbc.M310752200
Niranjanakumari S, Lasda E, Brazas R, Garcia-Blanco MA (2002) Reversible cross-linking combined with immunoprecipitation to study RNA-protein interactions in vivo. Methods 26(2):182–190, doi:10.1016/S1046-2023(02)00021-XS1046-2023(02)00021-X [pii]
Lu K, Ye W, Zhou L, Collins LB, Chen X, Gold A, Ball LM, Swenberg JA (2010) Structural characterization of formaldehyde-induced cross-links between amino acids and deoxynucleosides and their oligomers. J Am Chem Soc 132(10):3388–3399. doi:10.1021/ja908282f
Toth J, Biggin MD (2000) The specificity of protein-DNA crosslinking by formaldehyde: in vitro and in drosophila embryos. Nucleic Acids Res 28(2):e4
Barlow JJ, Mathias AP, Williamson R, Gammack DB (1963) A simple method for the quantitative isolation of undegraded high molecular weight ribonucleic acid. Biochem Biophyl Res Commun 13:61–66
Cathala G, Savouret JF, Mendez B, West BL, Karin M, Martial JA, Baxter JD (1983) A method for isolation of intact, translationally active ribonucleic acid. DNA 2(4):329–335
Vaughan R, Tragesser B, Ni P, Ma X, Dragnea B, Kao CC (2014) The tripartite virions of the brome mosaic virus have distinct physical properties that affect the timing of the infection process. J Virol 88(11):6483–6491. doi:10.1128/JVI. 00377-14
Seidler J, Zinn N, Boehm ME, Lehmann WD (2010) De novo sequencing of peptides by MS/MS. Proteomics 10(4):634–649. doi:10.1002/pmic.200900459
Koenig T, Menze BH, Kirchner M, Monigatti F, Parker KC, Patterson T, Steen JJ, Hamprecht FA, Steen H (2008) Robust prediction of the MASCOT score for an improved quality assessment in mass spectrometric proteomics. J Proteome Res 7(9):3708–3717. doi:10.1021/pr700859x
Perkins DN, Pappin DJ, Creasy DM, Cottrell JS (1999) Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20(18):3551–3567. doi:10.1002/(SICI)1522-2683(19991201)20:18<3551::AID-ELPS3551>3.0.CO;2-2
Clauser KR, Baker P, Burlingame AL (1999) Role of accurate mass measurement (+/- 10 ppm) in protein identification strategies employing MS or MS/MS and database searching. Anal Chem 71(14):2871–2882
Artimo P, Jonnalagedda M, Arnold K, Baratin D, Csardi G, de Castro E, Duvaud S, Flegel V, Fortier A, Gasteiger E, Grosdidier A, Hernandez C, Ioannidis V, Kuznetsov D, Liechti R, Moretti S, Mostaguir K, Redaschi N, Rossier G, Xenarios I, Stockinger H (2012) ExPASy: SIB bioinformatics resource portal. Nucleic Acids Res 40(Web Server issue):W597–603. doi:10.1093/nar/gks400
Wilkins MR, Gasteiger E, Bairoch A, Sanchez JC, Williams KL, Appel RD, Hochstrasser DF (1999) Protein identification and analysis tools in the ExPASy server. Methods Mol Biol 112:531–552
Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera–a visualization system for exploratory research and analysis. J Comput Chem 25(13):1605–1612. doi:10.1002/jcc.20084
Perez-Vargas J, Vaughan RC, Houser C, Hastie KM, Kao CC, Nemerow GR (2014) Isolation and characterization of the DNA and protein binding activities of adenovirus protein. J Virol 88(16):9287–9296
Janecki DJ, Beardsley RL, Reilly JP (2005) Probing protein tertiary structure with amidination. Anal Chem 77(22):7274–7281. doi:10.1021/ac050891z
Liu X, Reilly JP (2009) Correlating the chemical modification of Escherichia coli ribosomal proteins with crystal structure data. J Proteome Res 8(10):4466–4478. doi:10.1021/pr9002382
Running WE, Reilly JP (2009) Ribosomal proteins of Deinococcus radiodurans: their solvent accessibility and reactivity. J Proteome Res 8(3):1228–1246. doi:10.1021/pr800544y
Deval J, D’Abramo CM, Zhao Z, McCormick S, Coutsinos D, Hess S, Kvaratskhelia M, Gotte M (2007) High resolution footprinting of the hepatitis C virus polymerase NS5B in complex with RNA. J Biol Chem 282(23):16907–16916. doi:10.1074/jbc.M701973200
Lundblad RL (2005) Chemical reagents for protein modification, 3rd edn. CRC Press, Boca Raton, FL
Kannan N, Schneider TD, Vishveshwara S (2000) Logos for amino-acid preferences in different backbone packing density regions of protein structural classes. Acta Crystallogr D Biol Crystallogr 56(Pt 9):1156–1165
Inman JK, Perham RN, DuBois GC, Appella E (1983) Amidination. Methods Enzymol 91:559–569
Carven GJ, Stern LJ (2005) Probing the ligand-induced conformational change in HLA-DR1 by selective chemical modification and mass spectrometric mapping. Biochemistry 44(42):13625–13637. doi:10.1021/bi050972p
Lauber MA, Reilly JP (2011) Structural analysis of a prokaryotic ribosome using a novel amidinating cross-linker and mass spectrometry. J Proteome Res 10(8):3604–3616. doi:10.1021/pr200260n
Shepherd CM, Borelli IA, Lander G, Natarajan P, Siddavanahalli V, Bajaj C, Johnson JE, Brooks CL III, Reddy VS (2006) VIPERdb: a relational database for structural virology. Nucleic Acids Res 34(Database issue):D386–D389. doi:10.1093/nar/gkj032
Acknowledgement
This work was supported by a grant from the NIH NIAID 1R01AI090280. We thank C.T. Ranjith-Kumar, William Running, James Reilly, and Jonathan Karty for helpful discussions and reagents used to determine the conditions for the protocols in this work.
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Vaughan, R.C., Kao, C.C. (2015). Mapping Protein–RNA Interactions by RCAP, RNA-Cross-Linking and Peptide Fingerprinting. In: Guo, P., Haque, F. (eds) RNA Nanotechnology and Therapeutics. Methods in Molecular Biology, vol 1297. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2562-9_16
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DOI: https://doi.org/10.1007/978-1-4939-2562-9_16
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