Nuclear magnetic resonance study of side-chain conformation of phenylalanine residue in [Met5]-enkephalin: solvent, pH, and temperature dependence.

[Met5]-Enkephalin and N-acetylphenylalanine methylamide containing (2S,3S)-[2,3-2H2]Phe were synthesized 270 MHz 1H NMR spectra of the normal and selectively deuterated species were analysed. The lower-field and higher-field beta-proton signals of the Phe4 residue of [Met5]-enkephalin were unambiguously assigned to the pro-S and pro-R protons, respectively. The same assignments apply to N-acetylphenylalanine methylamide in polar organic solvents and in 2H2O, but the alternative assignments apply in C2HCl3. For [Met5]-enkephalin, the vicinal spin coupling constants 3JalphabetaS and 3 JalphabetaR and the rotamer populations around the Calpha-Cbeta bond were determined in a variety of solvents. From the pH and temperature dependences of rotamer populations of [Met5]-enkephalin, the side-chain conformation of the Phe residue in 2H2O solution was found to be considerably different from that in (C2H3)2SO solution. Rotamer populations of the Phe4 residue of [Met5]-enkephalin in organic solvents depend on solvent polarity. As compared with the reference model molecule of N-acetylphenylalanine methylamide, the rotamer populations of Phe4 of [Met5]-enkephalin are affected possibly by steric repulsion with other residues; the rotamer I is primarily favored but the rotamer II is appreciably destabilized in weakly polar solvents.     

Different rotamer populations around the C-5C-6 bond for α- and β-d-galactopyranosides through the combined interaction of the gauche and anomeric effects: A 300-MHz 1H-n.m.r. and mndo study

A 300-MHz ¹H-n.m.r. study of methyl 2,3,4-tri-O-methyl-ga- (1) and β-d-galactopyranoside-6-(dimethyl phosphate) (3), using various solvents, shows that the gauche (gg) rotamer populations about the C-5C-6 bond are are the same in all solvents, whereas those of the gauche(trans) (gt) and trans(gauche) (tg, O-5 and O-6 trans) rotamers are solvent dependent. The tg population increases with decreasing polarity of the solvent, which is attributed to an increased electrostatic repulsion between O-5 and O-6 in apolar solvents. The tg population of 3 is larger than that of 1 and the same difference is observed in the corresponding compounds (2 and 4) which have a trigonal-bipyramidal five-coördinated phosphorus (P^v) at position 6 and which have a higher electron density at O-6. These differences in rotamer populations are due to an effect additional to that of the coulombic effect between O-5 and O-6. That these differences are caused by a combination of the gauche and anomeric effects is supported by the finding that the tg population increases with increasing pK_a of the group at C-1. The results of the n.m.r. measurements (in CCl₄) are reproduced fairly accurately by MNDO calculations on model systems. The solvent dependence of the rotamer population around the C-5ẋC-6 bond is a good criterion for the assignment of the H-6S,6R resonances since, for galactopyranosides, J_5,6S increases and J_5,6R decreases as the polarity of the solvent decreases.     

Quantitative two-dimensional nmr study of dermenkephalin,a highly potent and selective δ-opioid peptide

Dermenkephalin, H-Tyr-(D ) Met-Phe-His-Leu-Met-Asp-NH₂, a highly potent and selective δ-opioid peptide isolated from frog skin, was studied in DMSO-d₆ solution by two-dimensional nmr spectroscopy, including the determination of NH temperature coefficients, the evaluation of ³J coupling constants from phase-sensitive correlated spectroscopy (COSY) and the volumes of nuclear Overhauser effect (NOE) correlations. The two-dimensional NOE spectroscopy (NOESY) spectrum of dermenkephalin revealed sequential, medium-, and long-range effects. To put this information on a quantitative basis, special attention was devoted to J cross-peak suppression, quantification of the NOE volumes and analysis of the overlaps, normalization of the NOEs against diagonal peaks and H_ββ′ geminal interactions. Although most of the dihedral angles deduced from the ³J_Nα coupling constants together with several N_iα_i and α_iN_{i + 1} NOEs pointed to a partially extended peptide backbone, several N_i N_{i + 1} NOEs and β_i N_{i + 1} interactions argued in favor of a folded structure. Moreover, several long-range correlations of strong intensities were found that supported a close spatial proximity between the side chains of D -Met² and Met⁶, Tyr¹ and His⁴, Tyr¹ and Asp⁷, and His⁴ and the C-terminal amide group. In Phe, the g^? rotamer in the side chain is deduced from the ³J_αβ coupling constants and αβ and Nβ NOE correlations. Whereas the amide proton dependency was not indicative of stable hydrogen bonds, the nonuniform values of the temperature coefficient may reflect an equilibrium mixture of folded and extended conformers. The overall data should provide realistic starting models for energy minimization and modelization studies. © 1993 John Wiley & Sons, Inc.     

Relating side-chain mobility in proteins to rotameric transitions: Insights from molecular dynamics simulations and NMR

The dynamic aspect of proteins is fundamental to understanding protein stability and function. One of the goals of NMR studies of side-chain dynamics in proteins is to relate spin relaxation rates to discrete conformational states and the timescales of interconversion between those states. Reported here is a physical analysis of side-chain dynamics that occur on a timescale commensurate with monitoring by ²H spin relaxation within methyl groups. Motivated by observations made from tens-of-nanoseconds long MD simulations on the small protein eglin c in explicit solvent, we propose a simple molecular mechanics-based model for the motions of side-chain methyl groups. By using a Boltzmann distribution within rotamers, and by considering the transitions between different rotamer states, the model semi-quantitatively correlates the population of rotamer states with ‘model-free’ order parameters typically fitted from NMR relaxation experiments. Two easy-to-use, analytical expressions are given for converting S²_axis’ values (order parameter for C–CH₃ bond) into side-chain rotamer populations. These predict that S²_axis’ values below 0.8 result from population of more than one rotameric state. The relations are shown to predict rotameric sampling with reasonable accuracy on the ps–ns timescale for eglin c and are validated for longer timescales on ubiquitin, for which side-chain residual dipolar coupling (RDC) data have been collected.     

Stereospecific assignments of 1H-nmr methyl lines and conformation of valyl residues in the lac repressor headpiece

Two-dimensional ¹H-nmr methods are described to obtain information on the sidechain conformation of valyl residues of the lac repressor headpiece and to assign the resonances of their methyl groups stereospecifically. The spin–spin coupling constants (J_αβ) between C^αand C^β protons are obtained from two-dimensional correlated spectroscopy experiments. Large values for J_αβ(10–12 Hz) corresponding to trans orientations for these protons (g⁺ conformation) are found for all valyl residues in α-helical segments. For these valyl residues, the distance between one methyl group (γ₁)and the valyl amide proton is much shorter than for the other methyl group, so that stereospecific resonance assignments follow from relative intensities of the corresponding cross peaks in a two-dimensional nuclear Overhauser enhancement spectrum. Thus, streospecific assignments could be made for the methyl groups of Val 9, 20, 23, and 38 (of a total of eight valyl residues).     

Nuclear magnetic resonance analyses of side chain conformations of histidine and aromatic amino acid derivatives. Solvent and pH dependence

Stereoselectively beta-deuterated species were synthesized of Ac-His-NHMe, Ac-His-OEt, Ac-His-OH and H-His-NHMe, which are useful as models of histidine residues in peptides. From the spectral comparison of 1H n.m.r., the beta-proton resonances of the normal species were unambiguously assigned. In (C2H3)2SO, C2(2)H5O2H, C2H3O2H, and C5(2)H5N solution and in aqueous solution, the lower-field and higher-field components of beta-proton resonances of the four histidine derivatives are assigned to the pro-R and pro-S protons, respectively. The alternative assignments apply for Ac-His-NHMe, Ac-His-OEt and Ac-His-OH in non-polar solvents such as C2HCl3. Vicinal coupling constants 3J alpha beta S and 3J alpha beta R were obtained for calculating the fractional populations of rotamers about the C alpha-C beta bond. The rotamer populations depend little on the ionization states of the alpha-amino and carboxyl groups or the imidazole ring. The rotamer populations depend significantly on the solvent polarity, similar to those of Phe, Tyr and Trp derivatives. For the two beta-proton resonances of His, Phe, Tyr, and Trp derivatives in a variety of solvents, linear relationships are found between the differences in chemical shifts and the differences in vicinal coupling constants.     

The role of helices 5 and 6 on the human β1-adrenoceptor activation mechanism

The human β₁-adrenoceptor (β₁AR) is a G-protein-coupled receptor (GPCR) involved in sympathetic system regulation through agonist-induced activation. The conserved CWXP-motif in helix 6 (rotamer toggle switch) is one of the most important activation switches in Class A GPCRs. In order to investigate how the agonist binding disturbs this switch, we carried out molecular dynamics simulations of a hβ₁AR model in the apo and R-noradrenaline-bound forms. The results show that the agonist binding changes the β₁-angle distribution of Cys336, Trp337 and Phe341 residues and increases the helix 6 bending. Overall, we provide a functional hβ₁AR model, showing how the rotamer toggle switch mechanism works at atomic level.     

1H-N.m.r. and c.d. investigation of the histidine side chain conformation in small peptides

Three series of model peptides containing histidine have been examined by ¹H-n.m.r. and c.d. spectroscopy: X-His peptides with X = Gly, Ala, Leu; His-X peptides with X = Gly, Ala, Leu, Ser, Lys, Phe, Tyr; and Pro-His-X peptides with X = Gly; Ala; Leu; Val; Phe; Tyr, C.d. spectra were obtained for pH values between 1 and 11 to give titration curves [θ] vs. pH; ¹H-n.m.r. spectra were recorded at four selected pH values corresponding to defined ionic species. ¹H-n.m.r. spectra in Me₂SO of the NH₃⁺, Imid⁺, COO^? ionic state (pH 4.5) were also obtained. The histidine side chain conformation in the various peptides and the changing ionic states is reflected in the ³J_αβ,β′ coupling constants, the Δδ ββ′ anisochrony values and the c.d. histidine chromophore contribution at 215 nm, and qualitative and semiquantitative correlations can be established between these parameters. Whereas the histidine side chain conformation is quite different in each of the three series, and varies with the ionic state and environment, it is practically identical for each peptide within a series: the nature of the X-residue does not exert any influence on the histidine side chain conformational behaviour. Thus, the classical rotamer distribution R I > R II > R III which is due to steric factors is usually observed unless specific intramolecular interactions such as hydrogen or ionic bonds override these.     

Conformational dependence of the vicinal proton coupling constant for the Cα?Cβ bond in peptides

We use the nmr data concerning the C^αH? C^βH fragment in eight peptides with rigid side chains to parametrize a Karplus correlation between the vicinal proton J_αβ coupling constant and the dihedral angle θ. When considering molecules containing the fragment C^αH^α? C^βH^βH^β′, the three-dimensional structure of the model peptides does not need to be known with accurate precision, since each set of J_αβ and J_αβ′ coupling constants is then related to the coefficients of the Karplus equation. A good correlation is observed with the Karplus equation, which is in substantial agreement with the J_αβ coupling constants reported for rigid as well as rotating C^α? C^β bonds in peptides.     

Analysis of side-chain rotamers in transmembrane proteins

We measured the frequency of side-chain rotamers in 14 alpha-helical and 16 beta-barrel membrane protein structures and found that the membrane environment considerably perturbs the rotamer frequencies compared to soluble proteins. Although there are limited experimental data, we found statistically significant changes in rotamer preferences depending on the residue environment. Rotamer distributions were influenced by whether the residues were lipid or protein facing, and whether the residues were found near the N- or C-terminus. Hydrogen-bonding interactions with the helical backbone perturbs the rotamer populations of Ser and His. Trp and Tyr favor side-chain conformations that allow their side chains to extend their polar atoms out of the membrane core, thereby aligning the side-chain polarity gradient with the polarity gradient of the membrane. Our results demonstrate how the membrane environment influences protein structures, providing information that will be useful in the structure prediction and design of transmembrane proteins.     

Topographic analysis of the S7 binding subsite of the tachykinin neurokinin-1 receptor

Conformationally and configurationally restricted rotameric probes of phenylalanine have been incorporated in the sequence of substance P (SP)—Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH₂—for analyzing the binding pockets of Phe⁷ (S₇) and Phe⁸ (S₈), in the neurokinin-1 receptor. These analogues of phenylalanine are (2S, 3R)- and (2S, 3S)-indanylglycines, E- and Z-α, β-dehydrophenylalanines, and 2(S)-α, β-cyclopropylphenylalanines [Δ^E Phe, Δ^ZPhe, ▿^E2(S)Phe, and ▿^Z2(S)Phe]. Binding data obtained with either conformationally (Ing diastereoisomers) or configurationally (Δ^EPhe, Δ^ZPhe) probes have unveiled large differences in the binding potencies of these rotameric probes. With the support of nmr data and energy calculations done on these SP-substituted analogues, we attempt to answer questions inherent to such study. First, none of these six probes prevents the formation of bioactive conformation(s) of the backbone of SP. Second, both diastereoisomers (S, S) and (S, R) of indanylglycine preferentially adopt, in the sequence of SP, the gauche (−) and trans side-chain orientations, respectively, as previously postulated from energy calculations with model peptides. However, in solution, the difference in energy between these rotamers included in the sequence of SP, compared to model peptides, is smaller since the other rotamer can be detected in [(2S, 3R) Ing⁷]SP. Finally, from this study we can hypothesize that the large variations observed in the affinities of Phe⁷ substituted analogues of SP must come from steric hindrance in the S₇ binding site, which drastically restricts the space filling around the C_α (SINGLE BOND) C_β bond of residue 7. © 1996 John Wiley & Sons, Inc.     

Design of Peptides Using α,β-Dehydro-residues: Synthesis,Crystal Structure and Molecular Conformation of N-Boc-L-Val-ΔPhe–ΔPhe-L-Ala-OCH3

To obtain general rules of peptide design using α,β-dehydro-residues, a sequence with two consecutive ΔPhe-residues, Boc-L -Val-ΔPhe–ΔPhe- L -Ala-OCH₃, was synthesized by azlactone method in solution phase. The peptide was crystallized from its solution in an acetone/water mixture (70:30) in space group P6₁ with a=b=14.912(3) Å, c= 25.548(5) Å, V=4912.0(6) ų. The structure was determined by direct methods and refined by a full matrix least-squares procedure to an R value of 0.079 for 2891 observed [I?3σ(I)] reflections. The backbone torsion angles ?₁=?54(1)°, ψ₁= 129(1)°, ω₁=?177(1)°, ?₂ =57(1)°, ψ₂=15(1)°, ω₂ =?170(1)°, ?₃=80(1)°, ψ₃ =7(2)°, ω₃=?177(1)°, ?₄ =?108(1)° and ψ^T₄=?34 (1)° suggest that the peptide adopts a folded conformation with two overlapping β-turns of types II and III′. These turns are stabilized by two intramolecular hydrogen bonds between the CO of the Boc group and the NH of ΔPhe³ and the CO of Val¹ and the NH of Ala⁴. The torsion angles of ΔPhe² and ΔPhe³ side chains are similar and indicate that the two ΔPhe residues are essentially planar. The folded molecules form head-to- tail intermolecular hydrogen bonds giving rise to continuous helical columns which run parallel to the c-axis. This structure established the formation of two β-turns of types II and III′ respectively for sequences containing two consecutive ΔPhe residues at (i+2) and (i+3) positions with a branched β-carbon residue at one end of the tetrapeptide.     

Conformational Similarity Indices Between Different Residues in Proteins and α-Helix Propensities

Abstract

Various amino acid similarity matrices have been derived using data on physicochemical properties and molecular evolution. Conformational similarity indices, CS_XX′, between different residues are computed here using the distribution of the main-chain and side-chain torsion angles and the values have been used to cluster amino acids in proteins. A subset of these parameters, CS_AX′ indicates the extent of similarity in the main-chain and side-chain conformations (φ ψ and χ₁) of different residues (X) with Ala (A) and is found to have strong correlation with α-helix propensities. However, no subset of CS_XX′ provides any linear relationship with β-sheet propensities, suggesting that the conformational feature favouring the location of a residue in an a-helix is different from the one favouring the β-sheet. Conformationally similar residues (close CS_AX values) have similar steric framework of the side-chain (linear/branched, aliphatic/aromatic), irrespective of the polarity or hydrophobicity. Cooperative nucleation of helix may be facile for a contiguous stretch of residues with high overall CS_AX values.     

Effect of Short- and Long-Range Interactions on Trp Rotamer Populations Determined by Site-Directed Tryptophan Fluorescence of Tear Lipocalin

In the lipocalin family, the conserved interaction between the main α-helix and the β-strand H is an ideal model to study protein side chain dynamics. Site-directed tryptophan fluorescence (SDTF) has successfully elucidated tryptophan rotamers at positions along the main alpha helical segment of tear lipocalin (TL). The rotamers assigned by fluorescent lifetimes of Trp residues corroborate the restriction expected based on secondary structure. Steric conflict constrains Trp residues to two (t, g ⁻) of three possible χ₁ (t, g ⁻, g ⁺) canonical rotamers. In this study, investigation focused on the interplay between rotamers for a single amino acid position, Trp 130 on the α-helix and amino acids Val 113 and Leu 115 on the H strand, i.e. long range interactions. Trp130 was substituted for Phe by point mutation (F130W). Mutations at positions 113 and 115 with combinations of Gly, Ala, Phe residues alter the rotamer distribution of Trp130. Mutations, which do not distort local structure, retain two rotamers (two lifetimes) populated in varying proportions. Replacement of either long range partner with a small amino acid, V113A or L115A, eliminates the dominance of the t rotamer. However, a mutation that distorts local structure around Trp130 adds a third fluorescence lifetime component. The results indicate that the energetics of long-range interactions with Trp 130 further tune rotamer populations. Diminished interactions, evident in W130G113A115, result in about a 22% increase of α-helix content. The data support a hierarchic model of protein folding. Initially the secondary structure is formed by short-range interactions. TL has non-native α-helix intermediates at this stage. Then, the long-range interactions produce the native fold, in which TL shows α-helix to β-sheet transitions. The SDTF method is a valuable tool to assess long-range interaction energies through rotamer distribution as well as the characterization of low-populated rotameric states of functionally important excited protein states.     

Role of two consecutive α,β-dehydrophenylalanines in peptide structure: Crystal and molecular structure of Boc-Leu-ΔPhe-ΔPhe-Ala-Phe-NHMe

An N^α-protected model pentapeptide containing two consecutive ΔPhe residues, Boc-Leu-ΔPhe-ΔPhe-Ala-Phe-NHMe, has been synthesized by solution methods and fully characterized. ¹H-nmr studies provided evidence for the occurrence of a significant population of a conformer having three consecutive, intramolecularly H-bonded β-bends in solution. The solid state structure has been determined by x-ray diffraction methods. The crystals grown from aqueous methanol are orthorhombic, space group P2₁2₁2₁, a = 11.503(2), b = 16.554(2), c = 22.107(3) Å, V = 4209(1) Å,³ and Z = 4. The x-ray data were collected on a CAD4 diffractometer using CuK_a radiation (λ = 1.5418 Å). The structure was determined using direct methods and refined by full-matrix least-squares procedure. The R factor is 5.3%. The molecule is characterized by a right handed 3₁₀-helical conformation (〈ϕ〉 = −68.2°, 〈ψ〉 = −26.3°), which is made up of two consecutive type III β-bends and one type I β-bend. In the solid state the helical molecules are aligned head-to-tail, thus forming long rod like structures. A comparison with other peptide structures containing consecutive ΔPhe residues is also provided. The present study confirms that the -ΔPhe-ΔPhe-sequence can be accommodated in helical structures. © 1997 John Wiley & Sons, Inc. Biopoly 42: 373–382, 1997     

1H-1H and 13C-13C vicinal coupling constants and amino acid side chain conformation in peptides

The vicinal coupling constants ¹³C′-¹³C^γ were measured in aspartic acid and phenylalanine (85 % ¹³C enrichment) as free amino acids and in the peptides Asp-Pro and Gly-Pro-Phe. These coupling constants used in connection with those between the α -and the β-protons provide the unambiguous assignment of rotamers I and II in the Asp and Phe side chains. The method is generally applicable to other amino acids and residues even in large peptides. A possible set of J_g^c,c and J_t^c,c values is proposed for the use of carbon 13-carbon 13 vicinal coupling constants in the side chain conformational studies of amino acid residues with a free carboxyl group.     

Synthesis and conformational study of peptides possessing helically arranged ΔZPhe side chains

Z-Dehydrophenylalanine (Δ^zPhe) possessing four oligopeptides, Boc-(L -Ala-Δ^zPhe-Aib)_n-OCH₃ (n = 1–4: Boc, t-butoxycarbonyl; Aib, α-aminoisobutyric acid), were synthesized, and their solution conformations were investigated by ¹H-nmr, ir, uv, and CD spectroscopy and theoretical CD calculation. ¹H-nmr (the solvent accessibility of NH groups) and ir studies indicated that all the NH groups except for those belonging to the N-terminal L -Ala-Δ^zPhe moiety participate in intramolecular hydrogen bonding in chloroform. This suggests that the peptides n = 2–4 have a 4 → 1 hydrogen-bonding pattern characteristic of 3₁₀-helical structures. The uv spectra of all these peptides recorded in chloroform and in trimethyl phosphate showed an intense maximum around 276 nm assigned to the Δ^zPhe chromophores. The corresponding CD spectra of the peptides n = 2–4 showed exciton couplets with a negative peak at longer wavelengths, whereas that of the peptide n = 1 showed only weak signals. Theoretical CD spectra were calculated for the peptides n = 2–4 of several helical conformations, on the basis of exciton chirality method. This calculation indicated that the three peptides form a helical conformation deviating from the perfect 3₁₀-helix that contains three residues per turn, and that their side chains of Δ^z Phe residues are arranged regularly along the helix. The center-to-center distance between the nearest phenyl pair(s) was estimated to be ～ 5.5 Å. The chemical shifts of the Δ^zPhe side-chain protons (H^β and aromatic H) for the peptides n = 2–4 indicated anisotropic shielding effect of neighboring phenyl group(s); the effect also supports a regular arrangement of the Δ^z Phe side chains along the helical axis. © 1993 John Wiley & Sons, Inc.     

NMR evidence for a type I β-turn in (Pro2)-tetrapeptides and interdependence of cis:Trans isomerism,ring flexibility,and backbone conformation

Tetrapeptides with proline in position 2, asparagine or leucine in position 3, and glycine in positions 1 and 4, with end groups free or blocked on the N-terminal side, were studied in their various ionic states in ²H₂O and in Me₂SO-d₆ by ¹H- and ¹³C-nmr. In order to clarify or refine some details, successive substitutions of the residues in these peptides with amino acids enriched to 85% in ¹³C, or to 85% ¹³C plus 97% ²H were carried out. The ¹H and ¹³C chemical shifts as well as the ¹H-¹H, ¹³C-¹³C, and ¹³C-¹H coupling constants and the signal intensities show strong similarity of behavior between the tetrapeptides of asparagine and leucine. The main conformational characteristics are (1) the almost total stabilization of the trans conformer in the type I β-turn structure when the peptide is in the zwitterion state dissolved in Me₂SO. This is deduced from the ³J and the ³J coupling constants, which both furnish a dihedral angle of ?³ = ?90°, and from the positive value of the temperature coefficient of the glycine-4 amide protons, which suggests a type 4 → 1 hydrogen bond; (2) the evolution of cis and trans isomer fractions which change with the ionic state of the peptides in Me₂SO, whereas they remain constant in aqueous solution; and (3) the conformation of the pyrrolidine ring as it follows the variations in cis:trans isomer populations together with the side-chain rotamer fractions of the residue in position 3. In the β-turn conformation the isomer cis is less abundant and the pyrrolidine ring is more flexible; this explains the perfect accommodation of the proline residue in position 2 of a bend. The interdependence of these phenomena where interactive forces play a predominant role underlines the importance of cooperative effects in the molecule. The results also suggest that the cis isomer of proline can adapt itself just as well as the trans isomer to position 2 of a type I β-turn.     

The structure of Λ-β1-[Co(R,R-picchxn)(S-tyr)]Br2 · 3.5H2O and solution configuration of Λ-β1-[Co(R,R-picchxn)(aromatic S-aminoacidate)] complexes: model systems for the investigation of non-covalent aromatic interactions

Intramolecular non-covalent interactions involving aromatic residues in the ternary species Λ-β₁-[Co(R,R-picchxn)(S-aa)]²⁺ (aa=Tyr, OMe-tyr or Phe) have been investigated. Such interactions are important to discriminatory processes associated with molecular recognition in chemical and biochemical systems. The single-crystal X-ray study of Λ-β₁-[Co(R,R-picchxn)(S-tyr)]Br₂ · 3.5H₂O demonstrates the influence of intramolecular π-π and bifurcated NH-π interactions in determining the molecular conformation of the complex cation in the solid state. The Co(III) complexes synthesised are diamagnetic, and have been fully assigned in solution using multidimensional NMR techniques. Remarkably, the solid state conformation observed for Λ-β₁-[Co(R,R-picchxn)(S-tyr)]²⁺ has been shown to predominate in solutions of all the complexes, as evidenced by appropriate rOe correlations. ¹H NMR measurements carried out in order to determine equilibrium rotamer distributions confirm the dominance of this conformer in solution. NMR measurements also show that rotamer populations are relatively unchanged at elevated temperatures and in a variety of solvents. The results of this detailed study, which demonstrate the significance of non-covalent interactions involving aromatic residues to the determination of the molecular conformation, serve to highlight the suitability of these simple ternary Co(III) complexes to act as models for such interactions.     

A 13C-NMR study on the conformational and dynamical properties of a cereal seed storage protein,C-hordein,and its model peptides

We have recorded the ¹³C CP-MAS and DD-MAS nmr spectra of dry and hydrated barley storage protein, C-hordein, as a model for wheat S-poor prolamins, together with those of model synthetic peptides (Pro)₂(Gln)₆(I) and (Pro-Gln-Gln-Pro-Phe-Pro-Gln-Gln)₃(II) under dry or hydrated conditions. The spectral features of C-hordein as well as these peptides were appreciably different from each other depending on the extent of hydration, reflecting different domains that adopt different types of conformations as well as dynamics. In particular, considerable proportions of the peak intensities were lost in the CP-MAS spectra, and well-resolved ¹³C-nmr signals emerged in DD-MAS nmr spectra owing to acquisition of molecular motions by swelling. It was shown that local β-turn or (Pro)_n type II conformation is more preferable for individual Pro residues and β-sheet type conformation is dominant for individual Gln residues in the dry and hydrated systems. In addition, two types of Gln environments are originated in C-hordein that differ in their mobility. Further, ¹³C spin-lattice relaxation times (T₁'s) of C- hordein and peptide II were reduced by more than one order of magnitude by hydration, reflecting the presence of well-swollen molecular chains. In contrast, theT₁ values of peptide I upon hydration remained one third of those in the dry state. Carbon-resolved proton spin-lattice relaxation times in the rotating frame (T_1ρ's) were also decreased by about 50% upon hydration, although these parameters were less sensitive as compared to T₁ values. In addition, the ¹³C-nmr signals of the aromatic side chain of Phe residues disappeared on hydration owing to interference between the frequency of the acquired flip-flop motion and the proton decoupling frequency. This information gives a new insight into establishing the structural properties of the studied protein system. A model may be put forward for a gel-type structure in which the more rigid part of the system involves intermolecular hydrogen-bonded Gln side chains as well as some hydrophobic “pockets” involving Pro and Phe residues. The liquid-like domain is characterized by considerable backbone and side-chain motion as well as rapid ring-puckering motion in Pro residues. © 1997 John Wiley & Sons, Inc.