Separation of cross-relaxation and J cross-peaks in 2D rotating-frame NMR spectroscopy

Kessler, Horst; Griesinger, Christian; Kerssebaum, Rainer; Wagner, Karl G.; Ernst, R. R.

doi:10.1021/ja00236a063

Cited by 371 publications

(171 citation statements)

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“…The digital resolution in I Abbreviations: AMPCPP, adenosine S-(a,P-methylene)triphosphate; C(CO)NH, side-chain carbon to nitrogen (via carbonyl) correlation; CT, constant time: DGSA, distance geometry-simulated annealing; DIPSI, decoupling in the presence of scalar interactions; FID, free induction decay; GARP, globally optimized alternating-phase rectangular pulses; HMBC, heteronuclear multiple bond correlation; HMQC, heteronuclear multiple-quantum correlation; HSQC, heteronuclear single-quantum correlation; INEPT, insensitive nuclear enhancement by polarization transfer; NMR, nuclear magnetic resonance: NOE, nuclear Overhauser effect; NOESY, nuclear Overhauser effect spectroscopy: 2D, two dimensional; 3D, three dimensional; RF, radiofrequency; ROESY, rotating frame Overhauser effect spectroscopy; TOCSY, total correlation spectroscopy; TPPI, time-proportional phase incrementation; TSP, sodium 3-(trimethylsilyl)propionate-2,2,3,3-d4. (Kumar et al, 1980) and 2D ROESY experiments with a spin-lock time of 70 ms (Kessler et al, 1987) were carried out with unlabeled MutT (1.5 mM) in 2H20 containing the other components as described above. A 3D IH-13C NOESY-HSQC experiment was recorded at 500 MHz with a mixing time of 150 ms using the sequence of Muhandiram et al (1993) on 13C/ I5N-labeled MutT in D20.…”

Section: Methodsmentioning

confidence: 99%

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Abeygunawardana¹,

Weber²,

Gittis³

et al. 1995

Biochemistry

124

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The MutT enzyme (129 residues) catalyzes the hydrolysis of normal and mutagenic nucleoside triphosphates, such as 8-oxo-dGTP, by substitution at the rarely attacked P-P, to yield NMP and pyrophosphate. Previous heteronuclear NMR studies of MutT have shown the secondary structure to consist of a five-stranded mixed P-sheet connected by the loop I-a-helix I-loop I1 motif, by two tight tums, and by loop 111, and terminated by loop IV-a-helix I1 (4) restraints and 34 backbone hydrogen bond restraints were used to determine the tertiary structure ofMutT by distance geometry, simulated annealing, and energy minimization with the program X-PLOR. The structure is globular and compact with the parallel portion of the P-sheet sandwiched between the two a-helices, forming an a f , ! ? fold. The essential divalent cation has previously been shown to bind near residues Gly-37, Gly-38, Lys-39, and Glu-57, and nucleotides have been shown to bind near residues Leu-54 and Val-58 by NMR relaxation methods [Frick et al. (1995) Biochemistry 34, 5577-55861. The tertiary structure of MutT shows these residues to be near each other along the loop I-helix I region of the enzyme. A cluster of five glutamate residues (41,53, 56,57, and 98) form a patch of strongly negative electrostatic potential likely constituting the metal binding site. This site is contiguous with a deep cleft between P-strands A, C, and D and loop I which may contribute to the nucleotide binding site. This location of the active site is consistent with mutagenesis studies and with sequence homologies among MutT-like pyrophosphohydrolases.The MutT enzyme, a pyrophosphohydrolase of 129 residues, catalyzes the unusual hydrolysis of nucleoside and deoxynucleoside triphosphates (NTP) by nucleophilic substitution at the rarely attacked ,!?-phosphorus, yielding pyrophosphate and a nucleotide (NMP) as products (Bhatnagar et al., 1991;Weber et al., 1992). Like other enzymes which catalyze substitution at the electron-rich ,!?-phosphorus, this enzyme requires two divalent cations for activity (Frick et al., 1994). The biological role of this enzyme is to prevent ' This work was supported in part by National Institutes of Health Grants DK28616 (to A.S.M.), GM18649 (to M.J.B.), and GM36358 (which supports A.G.G.).A complete listing of the distance restraints derived from the NOE data has been deposited at the Brookhaven Protein Data Bank (Chemistry Department, Brookhaven National Laboratory, Upton, NJ) together with the atomic coordinates of the family of 15 acceptable structures (file name IMUT).

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Section: Methodsmentioning

confidence: 99%

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Abeygunawardana¹,

Weber²,

Gittis³

et al. 1995

Biochemistry

124

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“…Finally, information regarding interproton distances was obtained by means of rotating-frame cross-relaxation twodimensional experiments of the ROESY type [15] using the pulsed spin-lock method of Kessler et al [16]. These experiments were performed at 2°C where the resolution of NH protons was best.…”

Section: Nuclear Magnetic Resonancementioning

confidence: 99%

Secondary structure of the Arg‐Gly‐Asp recognition site in proteins involved in cell‐surface adhesion

Reed

Hull

Lieth

et al. 1988

European Journal of Biochemistry

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The primary sequence Arg‐Gly‐Asp has been found in a number of proteins which bind to cell surface receptors. Studies with synthetic peptides have shown that the presence of charged side chains alone is not sufficient to confer binding activity. Application of folding algorithms to proteins and peptides having similar sequences indicates that binding activity is strongly correlated with the presence of two or more closely spaced residues that each have a high probability of initiating a β‐bend. Circular dichroic studies on the hexapeptide GRGDSP, whose sequence is contained in fibronectin and which also shows binding activity, demonstrate that it adopts an unusual conformation in aqueous solution. 1H‐NMR spectra of the peptide in aqueous solution show that the two amide hydrogens of Asp4 and Ser5 exchange very slowly. Computer‐assisted modeling using restrained molecular dynamics and energy minimization results in conformations that include two β‐bends of type III‐III or III‐I (hydrogen bonds 4 → 1 and 5 → 2), fully consistent with constraints imposed by 1H‐ and 13C‐NMR data. It is suggested that this unusual secondary structure provides an additional specificity determinant.

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“…The spectra in mixed water solvent (90% H20 + 10% :H20 ) include ID IH recorded ,aith P1T pulse sequence [11] and 2D nuclear Overhauser enhancement spectroscopy (NOESY) [12] with 1T detection pulse and a mixing time of 200 ms. The 2D experiments in 2H20 include clean total correlation spectroscopy (clean TOCSY) [13] with a mixing time of 100 ms, NOESY with a mixing time of 120 ms and rotating frame nuclear Overhauser enhancement spectroscopy (ROESY) [14] with a mixing time of lI/(1 ms. Temperature of 10°C was used in most NMR experiments, though 1D experiments were carried out in the range 5 40°( ".…”

Section: Methodsmentioning

confidence: 99%

Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

Chary

Govil

et al. 1995

FEBS Letters

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Separation of cross-relaxation and J cross-peaks in 2D rotating-frame NMR spectroscopy

Cited by 371 publications

References 0 publications

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Secondary structure of the Arg‐Gly‐Asp recognition site in proteins involved in cell‐surface adhesion

Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

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Separation of cross-relaxation and J cross-peaks in 2D rotating-frame NMR spectroscopy

Cited by 371 publications

References 0 publications

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Solution Structure of the MutT Enzyme, a Nucleoside Triphosphate Pyrophosphohydrolase

Secondary structure of the Arg‐Gly‐Asp recognition site in proteins involved in cell‐surface adhesion

Evidence for A+(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC

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Evidence for A⁺(anti)–G(syn) mismatched base–pairing in d–GGTAAGCGTACC