Abstract We present the de novo resonance assignments for the crystalline 33 kDa C-terminal domain of the Ure2 prion using an optimized set of five 3D solid-state NMR spectra. We obtained, using a single uniformly 13C, 15N labeled protein sample, sequential chemical-shift information for 74% of the N, Cα, Cβ triples, and for 80% of further side-chain resonances for these spin systems. We describe the procedures and protocols devised, and discuss possibilities and limitations of the assignment of this largest protein assigned today by solid-state NMR, and for which no solution-state NMR shifts were available. A comparison of the NMR chemical shifts with crystallographic data reveals that regions with high crystallographic B-factors are particularly difficult to assign. While the secondary structure elements derived from the chemical shift data correspond mainly to those present in the X-ray crystal structure, we detect an additional helical element and structural variability in the protein crystal, most probably originating from the different molecules in the asymmetric unit, with the observation of doubled resonances in several parts, including entire stretches, of the protein. Our results provide the point of departure towards an atomic-resolution structural analysis of the C-terminal Ure2p domain in the context of the full-length prion fibrils.
Extensive de novo solid-state NMR assignments of the 33*kDa C-terminal domain of the Ure2 prion.
Extensive de novo solid-state NMR assignments of the 33*kDa C-terminal domain of the Ure2 prion.
Extensive de novo solid-state NMR assignments of the 33*kDa C-terminal domain of the Ure2 prion.
J Biomol NMR. 2011 Jul 31;
Authors: Habenstein B, Wasmer C, Bousset L, Sourigues Y, Schütz A, Loquet A, Meier BH, Melki R, Böckmann A
We present the de novo resonance assignments for the crystalline 33*kDa C-terminal domain of the Ure2 prion using an optimized set of five 3D solid-state NMR spectra. We obtained, using a single uniformly (13)C, (15)N labeled...
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08-02-2011 11:40 AM
The Core of Ure2p Prion Fibrils Is Formed by the N-Terminal Segment in a Parallel Cross-? Structure: Evidence from Solid-State NMR.
The Core of Ure2p Prion Fibrils Is Formed by the N-Terminal Segment in a Parallel Cross-? Structure: Evidence from Solid-State NMR.
The Core of Ure2p Prion Fibrils Is Formed by the N-Terminal Segment in a Parallel Cross-? Structure: Evidence from Solid-State NMR.
J Mol Biol. 2011 Apr 8;
Authors: Kryndushkin DS, Wickner RB, Tycko R
Intracellular fibril formation by Ure2p produces the non-Mendelian genetic element in Saccharomyces cerevisiae, making Ure2p a prion protein. We show that solid-state NMR spectra of full-length Ure2p fibrils, seeded...
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04-19-2011 11:01 PM
NMR assignments of the N-terminal domain of Nephila clavipes spidroin 1.
NMR assignments of the N-terminal domain of Nephila clavipes spidroin 1.
NMR assignments of the N-terminal domain of Nephila clavipes spidroin 1.
Biomol NMR Assign. 2010 Dec 10;
Authors: Parnham S, Gaines WA, Duggan BM, Marcotte WR, Hennig M
The building blocks of spider dragline silk are two fibrous proteins secreted from the major ampullate gland named spidroins 1 and 2 (MaSp1, MaSp2). These proteins consist of a large central domain composed of approximately 100 tandem copies of a 35-40 amino acid repeat sequence. Non-repetitive N and...
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12-15-2010 12:03 PM
[NMR paper] Magic-angle spinning solid-state NMR spectroscopy of the beta1 immunoglobulin binding domain of protein G (GB1): 15N and 13C chemical shift assignments and conformational analysis.
Magic-angle spinning solid-state NMR spectroscopy of the beta1 immunoglobulin binding domain of protein G (GB1): 15N and 13C chemical shift assignments and conformational analysis.
Related Articles Magic-angle spinning solid-state NMR spectroscopy of the beta1 immunoglobulin binding domain of protein G (GB1): 15N and 13C chemical shift assignments and conformational analysis.
J Am Chem Soc. 2005 Sep 7;127(35):12291-305
Authors: Franks WT, Zhou DH, Wylie BJ, Money BG, Graesser DT, Frericks HL, Sahota G, Rienstra CM
Magic-angle spinning...
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12-01-2010 06:56 PM
[NMR paper] Prion protein interaction with the C-terminal SH3 domain of Grb2 studied using NMR an
Prion protein interaction with the C-terminal SH3 domain of Grb2 studied using NMR and optical spectroscopy.
Related Articles Prion protein interaction with the C-terminal SH3 domain of Grb2 studied using NMR and optical spectroscopy.
Biochemistry. 2004 Aug 17;43(32):10393-9
Authors: Lysek DA, Wüthrich K
Transmissible spongiform encephalopathies have been observed exclusively in organisms expressing the host-encoded prion protein (PrP). The function of the cellular isoform of PrP found in healthy organisms has so far not been identified,...
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11-24-2010 10:01 PM
[NMR paper] Backbone 1H and 15N resonance assignments of the N-terminal SH3 domain of drk in fold
Backbone 1H and 15N resonance assignments of the N-terminal SH3 domain of drk in folded and unfolded states using enhanced-sensitivity pulsed field gradient NMR techniques.
Related Articles Backbone 1H and 15N resonance assignments of the N-terminal SH3 domain of drk in folded and unfolded states using enhanced-sensitivity pulsed field gradient NMR techniques.
J Biomol NMR. 1994 Nov;4(6):845-58
Authors: Zhang O, Kay LE, Olivier JP, Forman-Kay JD
The backbone 1H and 15N resonances of the N-terminal SH3 domain of the Drosophila signaling adapter...
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08-22-2010 03:29 AM
[NMR paper] Complete 15N and 1H NMR assignments for the amino-terminal domain of the phage 434 re
Complete 15N and 1H NMR assignments for the amino-terminal domain of the phage 434 repressor in the urea-unfolded form.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www.pubmedcentral.nih.gov-corehtml-pmc-pmcgifs-pubmed-pmc.gif Related Articles Complete 15N and 1H NMR assignments for the amino-terminal domain of the phage 434 repressor in the urea-unfolded form.
Proc Natl Acad Sci U S A. 1992 May 15;89(10):4397-401
Authors: Neri D, Wider G, Wüthrich K
The amino-terminal domain of the phage 434 repressor consisting of residues 1-69...
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08-21-2010 11:41 PM
[NMR paper] Selective and extensive 13C labeling of a membrane protein for solid-state NMR invest
Selective and extensive 13C labeling of a membrane protein for solid-state NMR investigations.
Related Articles Selective and extensive 13C labeling of a membrane protein for solid-state NMR investigations.
J Biomol NMR. 1999 May;14(1):71-4
Authors: Hong M, Jakes K
The selective and extensive 13C labeling of mostly hydrophobic amino acid residues in a 25 kDa membrane protein, the colicin Ia channel domain, is reported. The novel 13C labeling approach takes advantage of the amino acid biosynthetic pathways in bacteria and suppresses the...
Extensive de novo solid-state NMR assignments of the 33 kDa C-terminal domain of the Ure2 prion
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