Paramagnetic Labeling of Proteins and Pseudocontact Shift in Structural Biology

Abstract

Abstract:

The interactions between unpaired electrons and nuclear spins in proteins, usually represented in paramagnetic relaxation enhancement (PRE), pseudocontact shift (PCS) and residual dipolar couplings (RDC), provide rich sources of structural restraints. These paramagnetic effects are valuable in structure determination of protein and protein-ligand complex as well as in elucidation of protein dynamics. Because most proteins do not have paramagnetic centers, generation of paramagnetic restraints usually relies on site-specific labeling of proteins with paramagnetic ions. In this paper, efforts towards paramagnetic labeling of proteins, especially with lanthanide ions, are summarized, and PCS, as a powerful NMR spectroscopic tool for structural biology, is discussed.

Key words: NMR, paramagnetic labeling, protein, lanthanide ion, pseudocontact shift

CLC Number:

O482.53

YANG Yin，CHEN Jia-liang，SU Xun-cheng*.

Paramagnetic Labeling of Proteins and Pseudocontact Shift in Structural Biology

[J]. Chinese Journal of Magnetic Resonance.

References

[1] Bertini I, Luchinat C, Parigi G. Paramagnetic constraints: an aid for quick solution structure determination of paramagnetic metalloproteins[J]. Concepts Magn Reson, 2002, 14(4): 259－286.

[2] Bertini I, Luchinat C, Parigi G. Magnetic susceptibility in paramagnetic NMR[J]. Prog NMR Spectr, 2002, 40: 249－273.

[3] Clore G M, Iwahara J. Theory, practice, and applications of paramagnetic relaxation enhancement for the characterization of transient low-population states of biological macromolecules and their complexes[J]. Chem Rev, 2009, 109(9): 4 108－ 4 139.

[4] Otting G. Protein NMR using paramagnetic ions[J]. Annu Rev Biophys, 2010, 39: 387－405.

[5] Liu Zhu(刘主), Tang Chun(唐淳). Paramagnetic relaxation enhancement-a tool for visualizing transient protein structures(顺磁弛豫增强技术与蛋白质瞬态结构)[J]. Chinese J Magn Reson(波谱学杂志), 2011, 28(3): 301－316.

[6] Banci L, Bertini I, Bren K L, et al. The use of pseudocontact shifts to refine solution structures of paramagnetic metalloproteins: Met80Ala cyano-cytochrome c as an example[J]. J Biol Inorg Chem, 1996, 1(2): 117－126.

[7] Banci L, Bertini I, Savellini G G, et al. Pseudocontact shifts as constraints for energy minimization and molecular dynamics calculations on solution structures of paramagnetic metalloproteins[J]. Proteins, 1997, 29(1): 68－76.

[8] Schmitz C, Stanton-Cook M J, Su X C, et al. Numbat: an interactive software tool for fitting Δχ-tensors to molecular coordinates using pseudocontact shifts[J]. J Biomol NMR, 2008 41(3): 179－189.

[9] Yang Y, Li Q F, Cao C, et al. Site-specific labeling of proteins with a chemically stable, high-affinity tag for protein study[J]. Eur J Chem, 2013, 19(3): 1 097－1 103.

[10] Allegrozzi M, Bertini I, Janik M B L, et al. Lanthanide-induced pseudocontact shifts for solution structure refinements of macromolecules in shells up to 40 Å from the metal ion[J]. J Am Chem Soc, 2000, 122(17): 4 154－4 161.

[11] Su X C, Otting G. Paramagnetic labelling of proteins and oligonucleotides for NMR[J]. J Biomol NMR, 2010, 46(1): 101－112.

[12] Edwards T E, Sigurdsson S T. Site-specific incorporation of nitroxide spin-labels into 2'-positions of nucleic acids[J]. Nat Protoc, 2007, 2(8): 1 954－1 962.

[13] Shelke S A, Sigurdsson S T. Effect of N3 modifications on the affinity of spin label ç for abasic sites in duplex DNA[J]. Chembiochem, 2012, 13(5): 684－690.

[14] Jakobsen U, Shelke S A, Vogel S, et al. Site-directed spin-labeling of nucleic acids by click chemistry: detection of abasic sites in duplex DNA by EPR spectroscopy[J]. J Am Chem Soc, 2010, 132(30): 10 424－10 428.

[15] Lee L, Sykes B D. Proton nuclear magnetic resonance determination of the sequential ytterbium replacement of calcium in carp parvalbumin[J]. Biochemistry, 1981, 20(5): 1 156－1 162.

[16] Pintacuda G, Keniry M A, Huber T, et al. Fast structure-based assignment of ¹⁵N HSQC spectra of selectively ¹⁵N-labeled paramagnetic proteins[J]. J Am Chem Soc, 2004, 126(9): 2 963－2 970.

[17] Keniry M A, Park A Y, Owen E A, et al. Structure of the theta subunit of Escherichia coli DNA polymerase III in complex with the epsilon subunit[J]. J Bacteriol, 2006, 188(12): 4 464－4 473.

[18] Biekofsky R R, Muskett F W, Schmidt J M, et al. NMR approaches for monitoring domain orientations in calcium-binding proteins in solution using partial replacement of Ca²⁺ by Tb³⁺[J]. FEBS Lett, 1999, 460(3): 519－526.

[19] Bertini I－8 021., Gelis I, Katsaros N, et al. Tuning the affinity for lanthanides of calcium binding proteins[J]. Biochemistry, 2003, 42(26): 8 011

[20] Li S, Yang W, Maniccia A W, et al. Rational design of a conformation-switchable Ca²⁺- and Tb³⁺-binding protein without the use of multiple coupled metal-binding sites[J]. FEBS J, 2008, 275(20): 5 048－5 061.

[21] Ma C, Opella S J. Lanthanide ions bind specifically to an added “EF-hand” and orient a membrane protein in micelles for solution NMR spectroscopy[J]. J Magn Reson, 2000, 146(2): 381－384.

[22] Nitz M, Franz K J, Maglathlin R L, et al. A powerful combinatorial screen to identify high-affinity terbium(III)-binding peptides[J]. Chembiochem, 2003, 4(4): 272－276.

[23] Wöhnert J, Franz K J, Nitz M, et al. Protein alignment by a coexpressed lanthanide-binding tag for the measurement of residual dipolar couplings[J]. J Am Chem Soc, 2003, 125(44): 13 338－13 339.

[24] Martin L J, Hähnke M J, Nitz M, et al. Double-lanthanide-binding tags: design, photophysical properties, and NMR applications[J]. J Am Chem Soc, 2007, 129(22): 7 106－7 113.

[25] Barthelmes K, Reynolds A M, Peisach E, et al. Engineering encodable lanthanide-binding tags into loop regions of proteins[J]. J Am Chem Soc, 2011, 133(4): 808－819.

[26] Su X C, Huber T, Dixon N E, et al. Site-specific labelling of proteins with a rigid lanthanide-binding tag[J]. Chembiochem, 2006, 7(10): 1 599－1 604.

[27] Su X C, McAndrew K, Huber T, et al. Lanthanide-binding peptides for NMR measurements of residual dipolar couplings and paramagnetic effects from multiple angles[J]. J Am Chem Soc, 2008, 130(5): 1 681－1 687.

[28] Saio T, Ogura K, Yokochi M, et al. Two-point anchoring of a lanthanide-binding peptide to a target protein enhances the paramagnetic anisotropic effect[J]. J Biomol NMR, 2009, 44(3): 157－166.

[29] Pintacuda G, Moshref A, Leonchiks A, et al. Site-specific labelling with a metal chelator for protein-structure refinement[J]. J Biomol NMR, 2004, 29(3): 351－361.

[30] Ikegami T, Verdier L, Sakhaii P, et al. Novel techniques for weak alignment of proteins in solution using chemical tags coordinating lanthanide ions[J]. J Biomol NMR, 2004, 29(3): 339－349.

[31] Leonov A, Voigt B, Rodriguez-Castañeda F, et al. Convenient synthesis of multifunctional EDTA-based chiral metal chelates substituted with an S-mesylcysteine[J]. Eur J Chem, 2005, 11(11): 3 342－3 348.

[32] Haberz P, Rodriguez-Castañeda F, Junker J, et al. Two new chiral EDTA-based metal chelates for weak alignment of proteins in solution－1 278.[J]. Org Lett, 2006, 8(7): 1 275

[33] Prudêncio M, Rohovec J, Peters J A, et al. A caged lanthanide complex as a paramagnetic shift agent for protein NMR[J]. Eur J Chem, 2004, 10(13): 3 252－3 260.

[34] Peters F, Maestre-Martinez M, Leonov A, et al. Cys-Ph-TAHA: a lanthanide binding tag for RDC and PCS enhanced protein NMR[J]. J Biomol NMR, 2011, 51(3): 329－337.

[35] Su X C, Man B, Beeren S, et al. A dipicolinic acid tag for rigid lanthanide tagging of proteins and paramagnetic NMR spectroscopy[J]. J Am Chem Soc, 2008, 130(32): 10 486－10 487.

[36] Swarbrick J D, Ung P, Su X C, et al. Engineering of a bis-chelator motif into a protein α-helix for rigid lanthanide binding and paramagnetic NMR spectroscopy[J]. Chem Commun (Camb), 2011, 47(26): 7 368－7 370.

[37] Swarbrick J D, Ung P, Chhabra S, et al. An iminodiacetic acid based lanthanide binding tag for paramagnetic exchange NMR spectroscopy[J]. Angew Chem Int Ed Engl, 2011, 50(19): 4 403－4 406.

[38] Yagi H, Maleckis A, Otting G. A systematic study of labelling an α-helix in a protein with a lanthanide using IDA-SH or NTA-SH tags[J]. J Biomol NMR, 2013, 55(2): 157－166.

[39] Man B, Su X C, Liang H, et al. 3-Mercapto-2,6-pyridinedicarboxylic acid: a small lanthanide-binding tag for protein studies by NMR spectroscopy[J]. Eur J Chem, 2010, 16(12): 3 827－3 832.

[40] Jia X, Maleckis A, Huber T, et al. 4,4'-dithiobisdipicolinic acid: a small and convenient lanthanide binding tag for protein NMR spectroscopy[J]. Eur J Chem, 2011, 17(24): 6 830－6 836.

[41] Keizers P H, Saragliadis A, Hiruma Y, et al. Design, synthesis, and evaluation of a lanthanide chelating protein probe: CLaNP-5 yields predictable paramagnetic effects independent of environment[J]. J Am Chem Soc, 2008, 130(44): 14 802－14 812.

[42] Häussinger D, Huang J R, Grzesiek S. DOTA-M8: An extremely rigid, high-affinity lanthanide chelating tag for PCS NMR spectroscopy[J]. J Am Chem Soc, 2009, 131(41): 14 761－14 767.

[43] Graham B, Loh C T, Swarbrick J D, et al. DOTA-amide lanthanide tag for reliable generation of pseudocontact shifts in protein NMR spectra[J]. Bioconjug Chem, 2011, 22(10): 2 118－2 125.

[44] Li Q F, Yang Y, Maleckis A, et al. Thiol-ene reaction: a versatile tool in site-specific labelling of proteins with chemically inert tags for paramagnetic NMR[J]. Chem Commun (Camb), 2012, 48(21): 2 704－2 706.

[45] Su X C, Liang H, Loscha K V, et al. [Ln(DPA)₃]^3– is a convenient paramagnetic shift reagent for protein NMR studies[J]. J Am Chem Soc, 2009, 131(30): 10 352－10 353.

[46] Jia X, Yagi H, Su X C, et al. Engineering [Ln(DPA)₃]^3– binding sites in proteins: a widely applicable method for tagging proteins with lanthanide ions[J].J Biomol NMR, 2011, 50(4): 411－420.

[47] Wei Z, Yang Y, Li Q F, et al. Noncovalent tagging proteins with paramagnetic lanthanide complexes for protein study[J]. Eur J Chem, 2013, 19(18): 5 758－5 764.

[48] Wang L, Xie J, Schultz P G. Expanding the genetic code[J]. Annu Rev Biophys Biomol Struct, 2006, 35: 225－249.

[49] Nguyen T H, Ozawa K, Stanton-Cook M, et al. Generation of pseudocontact shifts in protein NMR spectra with a genetically encoded cobalt(II)-binding amino acid[J]. Angew Chem Int Ed Engl, 2011, 50(3): 692－694.

[50] Loh C T, Ozawa K, Tuck K L, et al. Lanthanide tags for site-specific ligation to an unnatural amino acid and generation of pseudocontact shifts in proteins[J]. Bioconjug Chem, 2013, 24(2): 260－268.

[51] Banci L, Bertini I, Gori Savellini G G, et al. Pseudocontact shifts as constraints for energy minimization and molecular dynamic calculations on solution structures of paramagnetic metalloproteins[J]. Proteins Struct Funct Genet, 1997, 29(1): 68－76.

[52] Banci L, Bertini I, Cavallaro G, et al. Paramagnetism-based restraints for Xplor-NIH[J]. J Biomol NMR, 2004, 28(3): 249－261.

[53] Yagi H, Pilla K B, Maleckis A, et al. Protein fold determination from backbone amide pseudocontact shifts generated by lanthanide tags at multiple sites[J]. Structure, 2013, 21(6): 883－890.

[54] Bertini I, Del Bianco C, Gelis I, et al. Experimentally exploring the conformational space sampled by domain reorientation in calmodulin[J]. Proc Natl Acad Sci USA, 2004, 101(18): 6 841－6 846.

[55] Bertini I, Giachetti A, Luchinat C, et al. Conformational space of flexible biological macromolecules from average data[J]. J Am Chem Soc, 2010, 132(38): 13 553－13 558.

[56] Bertini I, Gupta Y K, Luchinat C, et al. Paramagnetism-based NMR restraints provide maximum allowed probabilities for the different conformations of partially independent protein domains[J]. J Am Chem Soc, 2007, 129(42): 12 786－12 794.

[57] Pintacuda G, Park A Y, Keniry M A, et al. Lanthanide labeling offers fast NMR approach to 3D structure determinations of protein-protein complexes[J]. J Am Chem Soc, 2006, 128(11): 3 696－3 702.

[58] Saio T, Yokochi M, Kumeta H, et al. PCS-based structure determination of protein-protein complexes[J]. J Biomol NMR, 2010, 46(4): 271－280.

[59] Ruan Ke(阮科), Gao Jia(高佳), Ma Rong-sheng(马荣声). NMR in fragment based lead discovery(基于片断的先导化合物发现中的核磁共振).[J] Chinese J Magn Reson(波谱学杂志)，2012, 29(2): 164－181.

[60] John M, Pintacuda G, Park A Y, et al. Structure determination of protein-ligand complexes by transferred paramagnetic shifts[J]. J Am Chem Soc, 2006, 128(39): 12 910－12 916.

[61] Balogh E, Wu D, Zhou G, et al. NMR second site screening for structure determination of ligands bound in the hydrophobic pocket of HIV-1 gp41[J]. J Am Chem Soc, 2009, 131(8): 2 821－2 823.

[62] Saio T, Ogura K, Shimizu K, et al. An NMR strategy for fragment-based ligand screening utilizing a paramagnetic lanthanide probe[J]. J Biomol NMR, 2011, 51(3): 395－408.

[63] de la Cruz L, Nguyen TH, Ozawa K, et al. Binding of low molecular weight inhibitors promotes large conformational changes in the dengue virus NS2B-NS3 protease: fold analysis by pseudocontact shifts[J]. J Am Chem Soc, 2011, 133(47):19 205－19 215.

[64] Guan J Y, Keizers P H, Liu W M, et al. Small-molecule binding sites on proteins established by paramagnetic NMR spectroscopy[J]. J Am Chem Soc. 2013, 135(15): 5 859－5 868.

[65] Wang Ya-qiang(王亚强), Li Cong-gang(李从刚), Pielak G J. In-cell protein magnetic resonance spectroscopy(细胞内蛋白质核磁共振).[J] Chinese J Magn Reson(波谱学杂志), 2012, 29(4): 475－487.

[66] Li J P, Pilla K B, L Q F, et al. Magnic angle spinning NMR structure determination of proteins from pseudocontact shifts[J]. J Am Chem Soc. 2013, 135(22): 8 294－8 303.