NMR Solution Structure

The reductase (MMOR) is a 39.7kDa protein that contains both FAD and a 6282] cluster in a single polypeptide chain (Lund and Dalton, 1985 Prince and Patel, 1986 Fox et al., 1989 Gassner and Lippard, 1999). The two cofactors are commonly found in proteins that are involved in electron transfer in other oxygenase systems, therefore MMOR is proposed to 

FIGURE 6. NMR solution structure of MMOB isolated from M. trichosporium OB3b. (Adapted from Chang et al., 1999). 

All ehemieal, spectroscopic, and kinetic studies of the MMO systems eondueted thus far are consistent with the formation of specific, high affinity complexes between the components. The points of interactions between MMOH and the other two components were first defined using chemical cross-linking procedures. MMOB is readily eross-linked to the MMOH a-subunit while MMOR cross-links to the p-subunit when a water-soluble carbodiimide is reacted with a stoichiometrie mixture of components in solution (Fox et al., 1991). The endogenous tryptophan fluorescence of MMOH and the eoncentration dependence of the MMO components in 

In the MMO OB3b system, the values could be used to predict the concentration dependence of the MMOB enhancement on the rate of the multiple turnover reaction. The fit to the experimental data predicts that the maximum rate is attained when a stoichiometric ternary complex (based on active site concentration) is established. Excess MMOB is inhibitory, apparently due to the formation of inactive MMOB-MMOR and MMOB-MMOB complexes, or perhaps binding of MMOB in the MMOR binding site. Cross-linking experiments were used to demonstrate the formation of each of these inhibitory complexes. Component complexes also play a significant role during the single turnover reaction as described below. 

---

MV Botuyan, A Toy-Palmer, J Chung, RC Blake II, P Beroza, DA Case. NMR solution structure of Cu(I) rusticyanm from Thiobacillus feiTooxidans Structural basis of the extreme acid stability and redox potential. J Mol Biol 263 752-767, 1996. 

NMR Assignments, Structural Constraints, and Structural Parameters for the Available NMR Solution Structures of Iron-Sulfur Proteins"- ... 

Tab. 2.8 Comparison of selected backbone torsion angles characteristic of the y-peptide 2.5-heli-cal backbone extracted from NMR solution structure of y -hexapeptide 141 and solid-state structure of y -tetrapeptide 146 [200, 205, 207]...

Mayer KL, Stone Ml. NMR solution structure and receptor peptide binding of the CC chemokine eotaxin-2. Biochemistry 2000 39 8382-95. 

Moy FJ, Chanda PK, Chen JM, Cosmi S, Edris W, Skotnicki JS, Wilhelm J, Powers P. NMR solution structure of the catalytic fragment of human fibroblast collagenase complexed with a sulfonamide derivative of a hydroxamic acid compound. Biochemistry 1999 38 7085-7096. 

Shimamoto, S., et al. (2007). NMR solution structure of lipocalin-type prostaglandin D synthase Evidence for partial overlapping of catalytic pocket and retinoic acid-binding pocket within the central cavity. /. Biol. Chem. (in press)... 

Tyukhtenko S, Tiburu EK, Deshmukh L, Vinogradova O, Janero DR, Makriyannis A (2009) NMR solution structure of human cannabinoid receptor-1 helix 7/8 peptide candidate electrostatic interactions and microdomain formation. Biochem Biophys Res Commun 390 441 146... 

Fernandez C, Wuthrich K (2003) NMR solution structure determination of membrane proteins reconstituted in detergent micelles. FEBS Lett 555 144-150... 

In addition to intrastrand cross-link described above, ds-DDP can also form an interstrand cross-link representing less than 10% of the total lesions [8,74]. The NMR solution structure of the duplex d(CCTCG CTCTC)-d(GAGAG CGAGG) containing a single inter-... 

These solution NMR and X-ray crystallographic findings have been contradicted by X-ray structures solved by Rypniewski et al.32 The results show a reduced active site unchanged from the oxidized state and let these authors to propose a five-coordinate copper ion that exists throughout the oxidation and reduction process. In 2001 the Protein Data Bank listed 39 X-ray crystallographic and NMR solution structures for CuZnSOD, including oxidized, reduced, genetically modified, and other species with or without attached substrates or substrate mimics such as azide ion. The reader is advised to search the Protein Data Bank for additional and more up-to-date structural depositions and search the literature for further discussion of mechanism. 

The NMR solution structure of Ras GDP [216] is very similar to the X-ray structure [22], except the position of helix 2, which matches neither with the corresponding helix in the GDP nor in the GppNHp X-ray structure, respectively. The authors argue that such energetically finely balanced conformational... 

Recently, Sivakolundu and Mabrouk published an NMR solution structure of horse heart ferrocytochrome c in which the heme contains an Fe(II) ion. This solution structure was the first in which the cytochrome c protein was dissolved in a nonaqueous solvent a solvent mix of 70% water and 30% acetonitrile (ACN). The data obtained from the NMR study are deposited in the protein data bank (PDB) as (1) ILCl, the minimized average NMR structure and (2) 1LC2, the 30 lowest energy NMR structures. 

One important visualization technique that users should acquire is the ability to transform two-dimensional stereoviews into three dimensions. The stereoviews may be found in visualization software such as MDL Chime and RasMol.They are also found in journal articles in Science or Nature and others, especially those describing new biomolecule X-ray crystallographic or NMR solution structures. The following hints for learning to transform the stereoviews come from the extremely helpful website http //www.usm.mame. edu/ rhodes/OHelp/StereoView.html. 

The reference 13 author compared the N-domain NaTK -ATPase X-ray crystallographic structure with NMR solution structures also published in 2003... 

Recent examples of successful peptide-ligand based discoveries of drug-like peptidomimetics include the discovery of SST antagonists, or the discovery of non-peptidic antagonists of the recently deorphanized urotensin II receptor at Sanofi-Aventis. ° As illustrated in Fig. 3, Flohr etal. used 3D models of the NMR solution structure of cyclic peptide derivatives of Urotensin II as a template for virtual 3D pharmacophore searches which resulted into non-peptidic candidates for lead optimization. 

M. O. Ebert, A. Luther, H. K. Huynh, R. Krishnamurthy, A. Eschenmoser, and B. Jaun, NMR solution structure of the duplex formed by self-pairing of a-L-arabinopyranosyl-(4, 2 )-(CGAATTCG), Helv. Chim. Acta, 85 (2002) 4055 1073. 

The former is a protein of 14.7 kDa involved in the multienzyme nucleotide excision repair (NER) pathway with a determined NMR solution structure . In this protein, the Zn + possesses rather a structural than a catalytic role. Zn NMR spectra were acquired using a rather sophisticated probe (for details, see Reference 87) and operating at temperatures 5-250 K. Data acquisition was performed with the application of spin-echo methods for enhanced sensitivity . Specifically, experiments were carried out at 25 K using a combination of CP (cross-polarization) and spikelet echo pulse sequences which provide a considerable increase in signal-to-noise ratio (of the order of 30) relative to a classical quadrupole echo pulse sequence. The proton field strength applied to the above measurements was 60 kHz with a matching field of 20 kHz for zinc and a contact time... 

---