Chemical Shift Assignment of the Transmembrane Helices of DsbB, a 20-kDa Integral Membrane Enzyme, by 3D Magic-angle Spinning NMR Spectroscopy

Overview

Journal Protein Sci

Specialty Biochemistry

Date 2008 Jan 30

PMID 18227427

Citations 40

Authors

Ying Li

Deborah A Berthold

Robert B Gennis

Chad M Rienstra

Affiliations

Soon will be listed here.

Abstract

The Escherichia coli inner membrane enzyme DsbB catalyzes disulfide bond formation in periplasmic proteins, by transferring electrons to ubiquinone from DsbA, which in turn directly oxidizes cysteines in substrate proteins. We have previously shown that DsbB can be prepared in a state that gives highly resolved magic-angle spinning (MAS) NMR spectra. Here we report sequential 13C and 15N chemical shift assignments for the majority of the residues in the transmembrane helices, achieved by three-dimensional (3D) correlation experiments on a uniformly 13C, 15N-labeled sample at 750-MHz 1H frequency. We also present a four-dimensional (4D) correlation spectrum, which confirms assignments in some highly congested regions of the 3D spectra. Overall, our results show the potential to assign larger membrane proteins using 3D and 4D correlation experiments and form the basis of further structural and dynamical studies of DsbB by MAS NMR.

Citing Articles

Efficient and stable reconstitution of the ABC transporter BmrA for solid-state NMR studies.

Kunert B, Gardiennet C, Lacabanne D, Calles-Garcia D, Falson P, Jault J Front Mol Biosci. 2015; 1:5.

PMID: 25988146 PMC: 4428385. DOI: 10.3389/fmolb.2014.00005.

Membrane proteins in their native habitat as seen by solid-state NMR spectroscopy.

Brown L, Ladizhansky V Protein Sci. 2015; 24(9):1333-46.

PMID: 25973959 PMC: 4570529. DOI: 10.1002/pro.2700.

On the problem of resonance assignments in solid state NMR of uniformly ¹⁵N,¹³C-labeled proteins.

Tycko R J Magn Reson. 2015; 253:166-72.

PMID: 25797013 PMC: 4371143. DOI: 10.1016/j.jmr.2015.02.006.

Lipid bilayer-bound conformation of an integral membrane beta barrel protein by multidimensional MAS NMR.

Eddy M, Su Y, Silvers R, Andreas L, Clark L, Wagner G J Biomol NMR. 2015; 61(3-4):299-310.

PMID: 25634301 PMC: 4398622. DOI: 10.1007/s10858-015-9903-1.

NMR structures of membrane proteins in phospholipid bilayers.

Radoicic J, Lu G, Opella S Q Rev Biophys. 2014; 47(3):249-83.

PMID: 25032938 PMC: 4153756. DOI: 10.1017/S0033583514000080.

References

Byrne B, Iwata S . Membrane protein complexes. Curr Opin Struct Biol. 2002; 12(2):239-43. DOI: 10.1016/s0959-440x(02)00316-0. View

Lundstrom K . Structural genomics of GPCRs. Trends Biotechnol. 2005; 23(2):103-8. DOI: 10.1016/j.tibtech.2004.12.006. View

Cornilescu G, Delaglio F, Bax A . Protein backbone angle restraints from searching a database for chemical shift and sequence homology. J Biomol NMR. 1999; 13(3):289-302. DOI: 10.1023/a:1008392405740. View

Kadokura H, Katzen F, Beckwith J . Protein disulfide bond formation in prokaryotes. Annu Rev Biochem. 2003; 72:111-35. DOI: 10.1146/annurev.biochem.72.121801.161459. View

McDermott A . Structural and dynamic studies of proteins by solid-state NMR spectroscopy: rapid movement forward. Curr Opin Struct Biol. 2004; 14(5):554-61. DOI: 10.1016/j.sbi.2004.09.007. View

Ohashi R, Takegoshi K . Asymmetric (13)C-(13)C polarization transfer under dipolar-assisted rotational resonance in magic-angle spinning NMR. J Chem Phys. 2006; 125(21):214503. DOI: 10.1063/1.2364503. View

Opella S, Marassi F . Structure determination of membrane proteins by NMR spectroscopy. Chem Rev. 2004; 104(8):3587-606. PMC: 3270942. DOI: 10.1021/cr0304121. View

Frericks H, Zhou D, Yap L, Gennis R, Rienstra C . Magic-angle spinning solid-state NMR of a 144 kDa membrane protein complex: E. coli cytochrome bo3 oxidase. J Biomol NMR. 2006; 36(1):55-71. DOI: 10.1007/s10858-006-9070-5. View

Inaba K, Takahashi Y, Ito K . Reactivities of quinone-free DsbB from Escherichia coli. J Biol Chem. 2005; 280(38):33035-44. DOI: 10.1074/jbc.M506189200. View

10.

Liu Y, Engelman D, Gerstein M . Genomic analysis of membrane protein families: abundance and conserved motifs. Genome Biol. 2002; 3(10):research0054. PMC: 134483. DOI: 10.1186/gb-2002-3-10-research0054. View

11.

van Gammeren A, Hulsbergen F, Hollander J, de Groot H . Residual backbone and side-chain 13C and 15N resonance assignments of the intrinsic transmembrane light-harvesting 2 protein complex by solid-state Magic Angle Spinning NMR spectroscopy. J Biomol NMR. 2005; 31(4):279-93. DOI: 10.1007/s10858-005-1604-8. View

12.

von Heijne G . Membrane-protein topology. Nat Rev Mol Cell Biol. 2006; 7(12):909-18. DOI: 10.1038/nrm2063. View

13.

Morcombe C, Zilm K . Chemical shift referencing in MAS solid state NMR. J Magn Reson. 2003; 162(2):479-86. DOI: 10.1016/s1090-7807(03)00082-x. View

14.

Pintacuda G, Giraud N, Pierattelli R, Bockmann A, Bertini I, Emsley L . Solid-state NMR spectroscopy of a paramagnetic protein: assignment and study of human dimeric oxidized CuII-ZnII superoxide dismutase (SOD). Angew Chem Int Ed Engl. 2006; 46(7):1079-82. DOI: 10.1002/anie.200603093. View

15.

Inaba K, Takahashi Y, Fujieda N, Kano K, Miyoshi H, Ito K . DsbB elicits a red-shift of bound ubiquinone during the catalysis of DsbA oxidation. J Biol Chem. 2003; 279(8):6761-8. DOI: 10.1074/jbc.M310765200. View

16.

Fernandez C, Wider G . TROSY in NMR studies of the structure and function of large biological macromolecules. Curr Opin Struct Biol. 2003; 13(5):570-80. DOI: 10.1016/j.sbi.2003.09.009. View

17.

Zhou D, Shea J, Nieuwkoop A, Franks W, Wylie B, Mullen C . Solid-state protein-structure determination with proton-detected triple-resonance 3D magic-angle-spinning NMR spectroscopy. Angew Chem Int Ed Engl. 2007; 46(44):8380-3. PMC: 2790053. DOI: 10.1002/anie.200702905. View

18.

Pervushin K, Riek R, Wider G, Wuthrich K . Attenuated T2 relaxation by mutual cancellation of dipole-dipole coupling and chemical shift anisotropy indicates an avenue to NMR structures of very large biological macromolecules in solution. Proc Natl Acad Sci U S A. 1997; 94(23):12366-71. PMC: 24947. DOI: 10.1073/pnas.94.23.12366. View

19.

Seavey B, Farr E, Westler W, Markley J . A relational database for sequence-specific protein NMR data. J Biomol NMR. 1991; 1(3):217-36. DOI: 10.1007/BF01875516. View

20.

Franks W, Kloepper K, Wylie B, Rienstra C . Four-dimensional heteronuclear correlation experiments for chemical shift assignment of solid proteins. J Biomol NMR. 2007; 39(2):107-31. DOI: 10.1007/s10858-007-9179-1. View