Solid‐State NMR of Matrix Metalloproteinase 12: An Approach Complementary to Solution NMR

Balayssac, Stéphane; Bertini, Ivano; Fälber, Katja; Fragai, Marco; Jehle, Stefan; Lelli, Moreno; Luchinat, Claudio; Oschkinat, Hartmut; Yeo, Kwon Joo

doi:10.1002/cbic.200600408

Cited by 42 publications

(30 citation statements)

References 13 publications

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“…The different 3D crystals of ArtP were directly used to record 13 C, 13 C PDSD correlation spectra. In all spectra, narrow lines comparable to other crystalline protein preparations [48][49][50] were observed. The expected two tyrosine and ten proline signal sets were easily identified in the spectra of ArtP (Table 2).…”

Section: Resultsmentioning

confidence: 77%

A MAS NMR Study of the Bacterial ABC Transporter ArtMP

et al. 2010

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ATP-binding cassette (ABC) transport systems facilitate the translocation of substances, like amino acids, across cell membranes energised by ATP hydrolysis. This work describes first structural studies on the ABC transporter ArtMP from Geobacillus stearothermophilus in native lipid environment by magic-angle spinning NMR spectroscopy. The 2D crystals of ArtMP and 3D crystals of isolated ArtP were prepared in different nucleotide-bound or -unbound states. From selectively (13)C,(15)N-labelled ArtP, several sequence-specific assignments were obtained, most of which could be transferred to spectra of ArtMP. Residues Tyr133 and Pro134 protrude directly into the ATP-binding pocket at the interface of the ArtP subunits, and hence, are sensitive monitors for structural changes during nucleotide binding and hydrolysis. Distinct sets of NMR shifts were obtained for ArtP with different phosphorylation states of the ligand. Indications were found for an asymmetric or inhomogeneous state of the ArtP dimer bound with triphosphorylated nucleotides. With this investigation, a model system was established for screening all functional states occurring in one ABC transporter in native lipid environment.

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Section: Resultsmentioning

confidence: 77%

A MAS NMR Study of the Bacterial ABC Transporter ArtMP

et al. 2010

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“…The 13 C-13 C PDSD spectra obtained with the 2 dilution schemes (diluted CoMMP-12 and diluted ZnMMP-12 samples) were recorded, assigned, and compared with those of fully enriched CoMMP-12 and ZnMMP-12 (38,45), respectively. Representative portions of these spectra are shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Paramagnetic shifts in solid-state NMR of proteins to elicit structural information

Balayssac

Bertini

Bhaumik

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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The recent observation of pseudocontact shifts (pcs) in 13 C highresolution solid-state NMR of paramagnetic proteins opens the way to their application as structural restraints. Here, by investigating a microcrystalline sample of cobalt(II)-substituted matrix metalloproteinase 12 [CoMMP-12 (159 AA, 17.5 kDa)], it is shown that a combined strategy of protein labeling and dilution of the paramagnetic species (i.e., 13 C-, 15 N-labeled CoMMP-12 diluted in unlabeled ZnMMP-12, and 13 C-, 15 N-labeled ZnMMP-12 diluted in unlabeled CoMMP-12) allows one to easily separate the pcs contributions originated from the protein internal metal (intramolecular pcs) from those due to the metals in neighboring proteins in the crystal lattice (intermolecular pcs) and that both can be used for structural purposes. It is demonstrated that intramolecular pcs are significant structural restraints helpful in increasing both precision and accuracy of the structure, which is a need in solid-state structural biology nowadays. Furthermore, intermolecular pcs provide unique information on positions and orientations of neighboring protein molecules in the solid phase.S olid-state NMR (SSNMR) on biomolecules is a rapidly growing technique, with an increased interest based on its ability to determine protein structures in the solid phase (1, 2) and to permit the study of noncrystalline biomolecular systems such as membrane proteins (3) and fibrils (4, 5).Limitations in biomolecular structural determination through SSNMR are due to the difficulties in obtaining a large number of restraints to be used for structural purposes (4, 6-9). Most of the structural information is obtained through distance restraints, analogous to nuclear Overhauser effects (NOE) in solution NMR, which in the solid state are obtained through experiments such as proton-driven spin diffusion (PDSD) (1, 6, 10), and CHHC (11), whereas specifically designed sequences can be applied on short peptides (4,(12)(13)(14). The ability to obtain a large number of distance restraints is hampered by the reduced resolution of SSNMR spectra, which increases the amount of ambiguities in the assigned restraints (15), whereas relayed transfer (10, 16) and the effects of the dipolar truncation (17) affect the accuracy of these restraints. These problems have been tackled by working on samples prepared with selective labeling schemes (1, 6) and, more recently, on uniformly labeled proteins with the help of software able to provide automated PDSD/ CHHC assignment and on dealing with a large number of ambiguous restraints (10,15,18). However, even when additional dihedral angle restraints from backbone chemical shiftsthrough Chemical Shift Index (CSI) (19) or TALOS (20) programs-are used, the size of the affordable proteins has been, up to now, limited to small systems (Ͻ100 aa) (10,15,18). In this work, we show how SSNMR paramagnetic restraints such as pseudocontact shifts (pcs) can be used as additional sources of restraints for protein structural determination, even providing information abou...

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“…[32,33] Proton chemical-shift evolution can of course be implemented in the indirect dimension of 13 C direct-detection NMR experiments, as proposed for the study of proteins [34][35][36][37] and nucleic acids. [38,39] Finally, the development of strategies to overcome problems related to 13 C detection in solution, such as homonuclear decoupling, [10,22,40] which have initially been stimulated by experiments developed for solid-state applications, [41,42] as well as common and complementary aspects of 13 C detection in solution and in the solid state, [13,17,[43][44][45] are now creating a basis for cross-fertilization between these two areas of biomolecular NMR [46] that can be eventually combined to tackle challenging systems. [43] The main focus of the experiments developed so far has been the identification of otherwise undetectable NMR resonances and sequence-specific assignment of the polypeptidic chain.…”

Section: Introductionmentioning

confidence: 98%

Exclusively Heteronuclear NMR Experiments to Obtain Structural and Dynamic Information on Proteins

et al. 2010

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Provided that (13)C-detected NMR experiments are either preferable or complementary to (1)H detection, we report here tools to determine C(alpha)-C', C'-N, and C(alpha)-H(alpha) residual dipolar couplings on the basis of the CON experiment. The coupling constants determined on ubiquitin are consistent with the subset measured with the (1)H-detected HNCO sequences. Since the utilization of residual dipolar couplings may depend on the mobility of the involved nuclei, we also provide tools to measure longitudinal and transverse relaxation rates of N and C'. This new set of experiments is a further development of a whole strategy based on (13)C direct-detection NMR spectroscopy for the study of biological macromolecules.

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Solid‐State NMR of Matrix Metalloproteinase 12: An Approach Complementary to Solution NMR

Cited by 42 publications

References 13 publications

A MAS NMR Study of the Bacterial ABC Transporter ArtMP

A MAS NMR Study of the Bacterial ABC Transporter ArtMP

Paramagnetic shifts in solid-state NMR of proteins to elicit structural information

Exclusively Heteronuclear NMR Experiments to Obtain Structural and Dynamic Information on Proteins

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