Investigating diproline segments in proteins: occurrences, conformation and classification

The covalent linkage between the side‐chain and the backbone nitrogen atom of proline leads to the formation of the five‐membered pyrrolidine ring and hence restriction of the backbone torsional angle ? to values of ?60 °± 30° for the L ‐proline. Diproline segments constitute a chain fragment with considerably reduced conformational choices. In the current study, the conformational states for the diproline segment ( ^L Pro‐ ^L Pro) found in proteins has been investigated with an emphasis on the cis and trans states for the Pro‐Pro peptide bond. The occurrence of diproline segments in turns and other secondary structures has been studied and compared to that of Xaa‐Pro‐Yaa segments in proteins which gives us a better understanding on the restriction imposed on other residues by the diproline segment and the single proline residue. The study indicates that P_II–P_II and P_II–α are the most favorable conformational states for the diproline segment. The analysis on Xaa‐Pro‐Yaa sequences reveals that the Xaa‐Pro peptide bond exists preferably as the trans conformer rather than the cis conformer. The present study may lead to a better understanding of the behavior of proline occurring in diproline segments which can facilitate various designed diproline‐based synthetic templates for biological and structural studies. © 2011 Wiley Periodicals, Inc. Biopolymers 97: 54–64, 2012.     

Solid-Phase Synthesis and NMR Structural Studies of the Marine Antibacterial Cyclic Tetrapeptide: Cyclo[GSPE]

Twelve-membered head-to-tail cyclic tetrapeptides (CTPs) are rigid molecules found in nature and possess a diverse range of biological activities. A possible reason may be due to their ability to adopt rigid conformations in solution mimicking reverse-turns. Reverse-turns are common structural motifs which serve as molecular recognition sites in many protein-receptor interactions. In this paper, we describe the solid-phase synthesis of the antibacterial cyclic tetrapeptide cyclo[Gly-Ser-Pro-Glu] (cyclo[GSPE]), first isolated from the Ruegeria strain of marine bacteria by Mitova et al. (J Nat Prod 67:1178–1181, 2004). Our NMR experiments in H₂O:D₂O:DMSO (18:1:1) revealed that it possessed three conformations in an approximate ratio of 4:2:1 based on NMR amide peak intensities. 2D NMR studies and computer calculations revealed that the major conformer adopted a reverse-turn conformation and have ω torsion angles twisted by up to 2°, with two transoid amide bonds between Gly-Ser, Pro-Glu and two cisoid amide bonds between Ser-Pro, Glu-Gly in a cis–trans-cis–trans (ctct) pattern. This supports previous reports that majority of CTPs adopt a ctct pattern when dissolved in hydrogen-bond disrupting solvents (Che and Marshall in J Med Chem 49:111–124, 2006 and references cited therein). An ensemble of ten lowest-energy-minimised 3D structures generated using XPLOR-NIH software revealed that cyclo[GSPE] possessed a rigid backbone ring scaffold. The remaining two minor conformers were present in quantities too low for NMR structural studies.     

Energetically unfavorable amide conformations for N6‐acetyllysine side chains in refined protein structures

The reversible acetylation of lysine to form N6‐acetyllysine in the regulation of protein function is a hallmark of epigenetics. Acetylation of the positively charged amino group of the lysine side chain generates a neutral N‐alkylacetamide moiety that serves as a molecular “switch” for the modulation of protein function and protein–protein interactions. We now report the analysis of 381 N6‐acetyllysine side chain amide conformations as found in 79 protein crystal structures and 11 protein NMR structures deposited in the Protein Data Bank (PDB) of the Research Collaboratory for Structural Bioinformatics. We find that only 74.3% of N6‐acetyllysine residues in protein crystal structures and 46.5% in protein NMR structures contain amide groups with energetically preferred trans or generously trans conformations. Surprisingly, 17.6% of N6‐acetyllysine residues in protein crystal structures and 5.3% in protein NMR structures contain amide groups with energetically unfavorable cis or generously cis conformations. Even more surprisingly, 8.1% of N6‐acetyllysine residues in protein crystal structures and 48.2% in NMR structures contain amide groups with energetically prohibitive twisted conformations that approach the transition state structure for cis‐trans isomerization. In contrast, 109 unique N‐alkylacetamide groups contained in 84 highly accurate small molecule crystal structures retrieved from the Cambridge Structural Database exclusively adopt energetically preferred trans conformations. Therefore, we conclude that cis and twisted N6‐acetyllysine amides in protein structures deposited in the PDB are erroneously modeled due to their energetically unfavorable or prohibitive conformations. Proteins 2013; © 2012 Wiley Periodicals, Inc.     

Nuclear magnetic resonance spectra of poly(N-alkylamino acid)s

220-MHz NMR spectra of various poly (N-alkylamino acid)s are investigated. Spectra of polysarcosine recorded in various solvents showed fine splittings of the methyl and methylene bands. Comparing the spectrum with that of its model compound, the fine structure of the methyl band of polysarcosine was assigned to four dyad sequences of the cis–trans isomeric state of the main chain amide bonds. Also the methylene band was roughly divided into cis and trans bands. From the temperature dependence of the spectra of polysarcosine, a double coalescence phenomenon was observed, in which the four dyad peaks coalesced into two peaks corresponding to cis and trans, then the two peaks coalesced into one peak. Further, the approximate value of the free energy for the internal rotation of the main chain amide bond was estimated. NMR spectra of various poly(N-alkylglycine)s in methylene chloride solution were also obtained. From the comparsion of their methylene bands, the introduction of the bulky N-alkyl groups was found to increase the cis content of the amide bond.     

Conformational energy studies on N-methylated analogs of thyrotropin releasing hormone, enkephalin, and luteinizing hormone-releasing hormone

Empirical conformational energy calculations have been carried out for N-methyl derivatives of alanine and phenylalanine dipeptide models and N-methyl-substituted active analogs of three biologically active peptides, namely thyrotropin-releasing hormone (TRH), enkephalin (ENK), and luteinizing hormone-releasing hormone (LHRH). The isoenergetic contour maps and the local dipeptide minima obtained, when the peptide bond (ω) preceding the N-methylated residue is in the trans configuration show that (1) N-methylation constricts the conformational freedom of both the ith and (i + 1)th residues; (2), the lowest energy position for both residues occurs around ? = ?135° ± 5° and ψ = 75° ± 5°, and (3) the α_L conformational state is the second lowest energy state for the (i + 1)th residue, whereas for the ith residue the C₅ (extended) conformation is second lowest in energy. When the peptide bond (ω_i) is in the cis configuration the ith residue is energetically forbidden in the range ? = 0° to 180° and ψ = ?180° to +180°. Conformations of low energy for ω_i = 0° are found to be similar to those obtained for the trans peptide bond. In all the model systems (irrespective of cis or trans), the α_R conformational state is energetically very high. Significant deviations from planarity are found for the peptide bond when the amide hydrogen is replaced by a methyl group. Two low-energy conformers are found for [(N-Me)His²]TRH. These conformers differ only in the ? and ψ values at the (N-Me)His² residue. Among the different low-energy conformers found for each of the ENK analogs [D -Ala²,(N-Me)Phe⁴, Met⁵]ENK amide and [D -Ala²,(N-Me)Met⁵]ENK amide, one low-energy conformer was found to be common for both analogs with respect to the side-chain orientations. The stability of the low-energy structures is discussed in the light of the activity of other analogs. Two low-energy conformers were found for [(N-Me)Leu⁷]LHRH. These conformations differ in the types of bend around the positions 6 and 7 of LHRH. One bend type is eliminated when the active analog [D -Ala⁶,(M-Me)Leu⁷]LHRH is considered.     

Cation binding cyclic peptides composed of imino acid residues

Cyclo(L -Pro-Sar)_n (n = 2–4) with moderate flexibility and hydrophobicity of molecular structure was synthesized, and the characteristics of these cyclic peptides and their metal complexes in acetonitrile were investigated in connection with the residual properties using ¹³C-nmr measurements. The cyclic tetrapeptide cyclo(L -Pro-Sar)₂ showed a sterically hindered phenomenon in acetonitrile in which the amide backbone adopted a cis-trans-cis-trans sequence. The cyclic hexapeptide cyclo(L -Pro-Sar)₃ existed as a mixture of several conformers whose interconversion is slow on the nmr time scale, including cis-cis-trans and/or cis-trans-trans arrangement of the Sar-Pro bond. Finally, it was demonstrated that the cyclic octapeptide cyclo(L -Pro-Sar)₄ behaved as a mixture of multiple conformers which allowed for cis-trans isomerism about the Pro-Sar peptide bond, of which 20–30% had the all-cis Sar-Pro bond isomer and the remaining 70–80% had one (or more) cis Sar-Pro bond isomer. ¹³C-nmr spectra also demonstrated that cyclo(L -Pro-Sar)_n (n = 3,4) formed a 1:1 ion complex whose conformation was characterized by an all-trans peptide bond in the presence of excess metal salt. Cation binding studies, using CD measurements, established that the ion selectivity of cyclo(L -Pro-Sar)₄ in acetonitrile decreased in the order, Ba²⁺ > Ca²⁺ > Na⁺ > Mg²⁺ > Li⁺.     

Removal of the 5‐nitro‐2‐pyridine‐sulfenyl protecting group from selenocysteine and cysteine by ascorbolysis

We previously reported on a method for the facile removal of 4‐methoxybenzyl and acetamidomethyl protecting groups from cysteine (Cys) and selenocysteine (Sec) using 2,2′‐dithiobis‐5‐nitropyridine dissolved in trifluoroacetic acid, with or without thioanisole. The use of this reaction mixture removes the protecting group and replaces it with a 2‐thio(5‐nitropyridyl) (5‐Npys) group. This results in either a mixed selenosulfide bond or disulfide bond (depending on the use of Sec or Cys), which can subsequently be reduced by thiolysis. A major disadvantage of thiolysis is that excess thiol must be used to drive the reaction to completion and then removed before using the Cys‐containing or Sec‐containing peptide in further applications. Here, we report a further advancement of this method as we have found that ascorbate at pH 4.5 and 25 °C will reduce the selenosulfide to the selenol. Ascorbolysis of the mixed disulfide between Cys and 5‐Npys is much less efficient but can be accomplished at higher concentrations of ascorbate at pH 7 and 37 °C with extended reaction times. We envision that our improved method will allow for in situ reactions with alkylating agents and electrophiles without the need for further purification, as well as a number of other applications. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Cyclo(D-Pro-L-Pro-D-Pro-L-Pro): Structural properties and cis/trans isomerization of the cyclotetrapeptide backbone

Combinations of L - and D -proline residues are useful compounds for finding new structures and properties of cyclic peptides. This is demonstrated with one striking example, the cyclic tetrapeptide c(D -Pro-L -Pro-D -Pro-L -Pro). For this molecule composed of strictly alternating D - and L -configurated residues, a highly symmetrical structure is expected, which should be an optically inactive meso-form. Cyclization of the enantiomeric pure linear precursor D -Pro-L -Pro-D -Pro-L -Pro, however, yields a racemic mixture of two enantiomeric cyclotetrapeptides, both with twofold symmetry and a cis–trans–cis–trans sequence of the peptide bonds. Remarkably, this formation of a racemate was not caused by racemization, but by cis/trans isomerization of all peptide bonds in the ring. This process may occur in the linear precursor during the ring formation (cyclization of conformers with trans–cis–trans or cis–trans–cis arrangement of the amide bonds) as well as in the enantiomeric pure cyclic tetrapeptide at higher temperature. In the latter case, an all-cis structure should exist as the intermediate, which can form a cis–trans–cis–trans sequence in two equivalent ways, leading finally to two enantiomeric cyclotetrapeptides. In the first one, the cis peptide bonds are attributed to the L -residues and the trans peptide bonds to the D -residues; in the second one, the cis bonds belong to the D and the trans bonds to the L -residues. The mixture of these two enantiomers does not crystallize in the racemic form, but in enantiomeric pure separate crystals. The structural properties could be proved by ¹H- and ¹³C-nmr spectroscopy and x-ray analysis. The cis/trans isomerization process was confirmed by optical rotation measurements and CD spectroscopy, as well as DREIDING model studies. Calorimetric measurements in the solid state suggest the existence of the expected all-cis intermediate. The backbone conformation of the 12-membered medium-sized ring shows only slight deviations—up to 6° —from the planarity of the peptide bonds. On the other hand, the four pyrrolidine rings show different types of puckering of the C_γ or the C_β atoms.     

13C- and 1H-nmr evidence for all-cis peptide bonds in cyclic tetrapeptide cyclo(L-Pro-Sar)2

Conformations of the cyclic tetrapeptide cyclo(L -Pro-Sar)₂ in solution were studied by ¹H- and ¹³C-nmr spectrometry and model building. The nmr data provide definite evidence that this cyclic peptide exists chiefly in two conformations, namely, a C₂-symmetric conformation and an asymmetric structure. The former was demonstrated to be predominant in polar solvents (100% in Me₂SO-d₆). This structure contains all cis-peptide bond linkages and all trans′ Pro C^α?CO bonds. It represents the first cyclic tetrapeptide in which all four peptide bonds have been found in the cis-conformation. As the polarity of the solvent decreases, the population of C₂-symmetric conformers decreases (88% in CD₃CN and 65% in CDCl₃). At the same time, a minor asymmetric conformer, characterized by cis-cis-cis-trans peptide bond sequences (two cis Sar-Pro bonds, one cis Pro-Sar bond, and one trans Pro-Sar bond), is seen to increase (9% in CD₃CN and 30% in CDCl₃). A proposed predominant conformation in solution for cyclo(L -Pro-Sar)₂ was compared with a crystal structure, as reported in an accompanying paper. Both structures show striking overall similarities.     

Synthesis and conformational analysis of an expanded cyclic ketoxime‐hexapeptide

In this work the synthesis of a linear hexapeptide with a hydroxylamine functionality at the N‐terminus and a ketone instead of the carboxylic acid at the C‐terminus is described. Cyclization by ketoxime formation yields the 19‐membered ring‐expanded cyclic hexapeptide cyclo[Goly‐Val‐Ala‐Pro‐Leu‐Kly] which adopts a main conformer with two intramolecular hydrogen bonds. The hydrolytic stability of a ketoxime lies between the inert amide and the labile imine. The substitution of an amide bond for an iminium bond transforms the irreversible macrocyclization into a reversible process, but macrocyclic imines are difficult to isolate because they are prone to hydrolysis. The enhanced chemical stability of the ketoxime justifies its application in ligation protocols. The detailed NMR analysis of a ketoxime linkage presented here identifies its local conformational preferences in a constrained peptide environment. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Comprehensive Chiroptical Study of Proline‐Containing Diamide Compounds

The continuously growing interest in the understanding of peptide folding led to the conformational investigation of methylamides of N‐acetyl‐amino acids as diamide models. Here we report the results of detailed conformational analysis on Ac‐Pro‐NHMe and Ac‐β‐HPro‐NHMe diamides. These compounds were analyzed by experimental and computational methods, the conformational distributions obtained by Density Functional Theory (DFT) calculations for isolated and solvated diamide compounds are discussed. The conformational preference of proline‐containing diamide compounds as a function of the ambience was observed by a number of chiroptical spectroscopic techniques, such as vibrational circular dichroism (VCD), electronic circular dichroism (ECD), Raman optical activity (ROA) spectroscopy, and additionally by single crystal X‐ray diffraction analyses. Based on a comparison between Ac‐Pro‐NHMe and Ac‐β‐HPro‐NHMe, one can conclude that due to the greater conformational freedom of the β‐HPro derivative, Ac‐β‐HPro‐NHMe shows different behavior in solid‐ and solution‐phase, as well. Ac‐β‐HPro‐NHMe tends to form cis Ac‐β‐HPro amide conformation in water, dichloromethane, and acetonitrile in contrast to its α‐Pro analog. On the other hand, the crystal structure of the β‐HPro compound cannot be related to any of the conformers obtained in vacuum and solution while the X‐ray structure of Ac‐Pro‐NHMe was identified as tα_L–, which is a trans Ac‐Pro amide containing conformer also predominant in polar solvents. Chirality 26:228–242, 2014. © 2014 Wiley Periodicals, Inc.     

trans-Stilbene degradation by Arthrobacter sp. SL3 cells

trans-Stilbene degradation was examined by the reaction using resting cells of microorganisms isolated through the enrichment culture using trans-stilbene. The strain SL3, showing the highest trans-stilbene-degrading activity, was identified as Arthrobacter sp. One of the reaction products was identified to be cis,cis-muconic acid. Arthrobacter sp. SL3 cells also transformed benzaldehyde, benzoic acid and catechol into cis,cis-muconic acid, suggesting that one benzene ring of trans-stilbene was converted into cis,cis-muconic acid via benzaldehyde formed by its C_α=C_β bond cleavage.     

Trans-cis amide bond isomerization in fulleroprolines

The ¹H NMR study of fulleroproline derivative Ac-Fpr-OtBu and its Pro analogue Ac-l -Pro-OtBu over a range of temperatures in toluene-d₈ solution has enabled the comparison of their equilibrium and activation parameters for the trans/cis interconversion around the amide partial double bond. © 1998 European Peptide Society and John Wiley & Sons, Ltd.     

Vibrational circular dichroism as a probe of solid‐state organisation of derivatives of cyclic β‐amino acids: Cis‐ and trans‐2‐aminocyclobutane‐1‐carboxylic acid

Peptide models built from cis‐ and trans‐2‐aminocyclobutane‐1‐carboxylic acids (ACBCs) are studied in the solid phase by combining Fourier‐transform infrared spectroscopy (FTIR) absorption spectroscopy, vibrational circular dichroism (VCD), and quantum chemical calculations using density functional theory (DFT). The studied systems are N‐tert‐butyloxycarbonyl (Boc) derivatives of 2‐aminocyclobutanecarboxylic acid (ACBC) benzylamides, namely Boc?(cis‐ACBC)?NH?Bn and Boc?(trans‐ACBC)?NH?Bn. These two diastereomers show very different VCD signatures and intensities, which of the trans‐ACBC derivative being one order of magnitude larger in the region of the ν (CO) stretch. The spectral signature of the cis‐ACBC derivative is satisfactorily reproduced by that of the monomer extracted from the solid‐state geometry of related ACBC derivatives, which shows that no long‐range effects are implicated for this system. In terms of hydrogen bonds, the geometry of this monomer is intermediate between the C6 and C8 structures (exhibiting a 6‐ or 8‐membered cyclic NH?O hydrogen bond) previously evidenced in the gas phase. The benzyl group must be in an extended geometry to reproduce satisfactorily the shape of the VCD spectrum in the ν (CO) range, which qualifies VCD as a potential probe of dispersion interaction. In contrast, reproducing the IR and VCD spectrum of the trans‐ACBC derivative requires clusters larger than four units, exhibiting strong intermolecular H‐bonding patterns. A qualitative agreement is obtained for a tetramer, although the intensity enhancement is not reproduced. These results underline the sensitivity of VCD to the long‐range organisation in the crystal.     

Sensory rhodopsin-I as a bidirectional switch: opposite conformational changes from the same photoisomerization

The phototaxis receptor sensory rhodopsin I (SRI) exists in two protein conformations, each of which is converted to the other by light absorption by the protein's retinylidene chromophore. One conformer inhibits a histidine-kinase attached to its bound transducer HtrI and its formation induces attractant motility responses, whereas the other conformer activates the kinase and its formation induces repellent responses. We performed Fourier transform infrared spectroscopy with temperature, pH, and mutation-induced shifts in the conformer equilibrium, and found that both conformers when present in the unphotolyzed dark state contain an all-trans retinal configuration that is photoisomerized to 13-cis, i.e., the same photoisomerization causes the opposite conformational change in the photointerconvertible pair of conformers depending on which conformer is present in the dark. Therefore, switching between the protein global conformations that define the two conformers is independent of the direction of isomerization. Insights into this phenomenon are gained from analysis of the evolution of the receptor from light-driven proton pumps, which use similar conformers for transport. The versatility of the conformational changes of microbial rhodopsins, including conformer interexchangeability in the photocycle as shown here, is likely a significant factor in the evolution of the diverse functionality of this protein family.     

Solid-state geometry and conformation of linear,diastereoisomeric oligoprolines

We have carried out a systematic analysis of the solid-state conformational preferences of a number of linear homo-oligoprolines (to the tetramer) by ir absorption and x-ray diffraction. The peptides present different chiral sequences (tacticities), various types (urethane and amide) of N-protecting groups, and free and blocked C-termini (which imply different capabilities of forming H-bonds). The following conclusions can be drawn: (i) values for the geometry of the prolyl residue and the peptide bond in the cis and in the trans conformations are proposed; (ii) in general the conformational angles φ and ψ in the linear homo-oligoprolines have values appropriate for the polyproline II structure (conformation F); (iii) the pyrrolidine ring shows various types of puckering with no apparent relation to the backbone conformation; (iv) Pro-Pro peptide bonds generally take the trans conformation, the few cases of cis conformation being formed by Pro residues of different chirality; (v) the single H-bond donor — OH, when present, is always bonded to H-acceptors, which can be either the urethane or the amide or the peptide carbonyl but never the carbonyl group of the — COOH moiety.     

Nmr studies of the molecular conformations in the linear oligopeptides H-(L-Ala)n-L-Pro-OH

The molecular conformations of the linear oligopeptides H-(L -Ala)_n-L -Pro-OH, with n = 1,2 and 3, have been investigated. ¹³C nmr observation of the equilibrium between the cis and trans forms of the Ala-Pro peptide bond indicated the occurrence of nonrandom conformations in solutions of these flexible peptides. The formation of the nonrandom species containing the cis form of the Ala-Pro bond was found to depend on the deprotonation of the carboxylic acid group of proline, the solvent, and the ionic strength in aqueous solution. The influence of intramolecular hydrogen bonding on the relative conformational energies of the species containing the cis and trans Ala-Pro peptide bond was studied by comparison of the peptides H-(Ala)_n-Pro-OH with analogous molecules where hydrogen bond formation was excluded by the covalent structure. In earlier work a hydrogen bond between the protonated terminal carboxylic acid group and the carbonyl oxygen of the penultimate amino acid residue had been suggested to stabilize conformations including trans proline. For the systems described here this hypothesis can be ruled out, since the cis:trans ratio is identical for molecules with methyl ester protected and free protonated terminal carboxylic acid groups of proline. Direct evidence for hydrogen bond formation between the deprotonated terminal carboxylic acid group and the amide proton of the penultimate amino acid residue in the molecular species containing cis proline was obtained from ¹H nmr studies. However, the cis:trans ratio of the Ala-Pro bond was not affected by N-methylation of the penultimate amino acid residue, which prevents formation of this hydrogen bond. Overall the experimental observations lead to the conclusion that the relative energies of the peptide conformations including cis or trans proline are mainly determined by intramolecular electrostatic interactions, whereas in the molecules considered, intramolecular hydrogen bonding is a consequence of specific peptide backbone conformations rather than a cause for the occurrence of energetically favored species. Independent support for this conclusion was obtained from model consideration which indicated that electrostatic interactions between the terminal carboxylic acid group and the carbonyl oxygen of the penultimate amino acid residue could indeed account for the observed relative conformational energies of the species containing cis and trans proline, respectively.     

Conformations of cysteine disulfides of peptide toxins: Advantage of differentiating forward and reverse asymmetric disulfide conformers

Conformations of cysteine disulfides were analyzed in X-ray, nuclear magnetic resonance (NMR), and co-crystal structures of peptide toxins retrieved from Protein Data Bank. The parameters side chain torsional angles, disulfide strain energy, interatomic Cα/Cβ distances, and Ramachandran angles were used as probes to derive conformational features of cysteine disulfides. Schmidt, Ho, and Hogg (2006 Schmidt, B., Ho, L., &; Hogg, P. J. (2006). Allosteric disulfide bonds. Biochemistry, 45, 7429–7433.[Crossref], [PubMed], [Web of Science ®] , [Google Scholar]) Allosteric disulfide bonds. Biochemistry, 45, 7429–7433 scheme was adapted to classify the disulfide conformations of peptide toxins. Anomalies were observed while treating “forward” and “reverse” asymmetric disulfide conformers as same disulfide conformation in peptide toxins. Thus, new scheme was proposed to classify “forward” and “reverse” asymmetric disulfide conformers separately. Total available conformers space for classification of toxins disulfides is 32. Interestingly, all 32 disulfide conformations are observed in peptide toxins. –LHSpiral is predominant disulfide conformation of peptide toxins. Significant variations were observed in population of occurrence of disulfide conformers, disulfide strain energy, and distribution of D_Cα-Cα and D_Cβ-Cβ values between X-ray, NMR, and co-crystal structures of peptide toxins. The observed differences in conformations of disulfides of same peptide toxins between different states were used as platform to demonstrate advantage of differentiating forward and reverse asymmetric disulfide conformers. Newly proposed scheme allows accurate representation of true conformational diversity of disulfides between X-ray and NMR structures of same peptide toxins. Newly proposed scheme also permits to derive additional structural information from nomenclature which was illustrated by comparing conformations of disulfides between unbound and bound form of toxin with channel/receptor. The results will be of interest for growing field of structural venomics and conformational analysis of peptide/protein disulfides.

Communicated by Ramaswamy H. Sarma     

The molecular structure of a curl-shaped retinal isomer

Computational studies of retinal protonated Schiff base (PSB) isomers show that a twisted curl-shaped conformation of the retinyl chain is a new low-lying minimum on the ground-state potential energy surface. The curl-shaped isomer has a twisted structure in the vicinity of the C₁₁=C₁₂ double bond where the 11-cis retinal PSB isomerizes in the rhodopsin photoreaction. The twisted configuration is a trapped structure between the 11-cis and all-trans isomers. Rotation around the C₁₀–C₁₁ single bond towards the 11-cis structure is prevented by steric interactions of the two methyl groups on the retinyl chain and by the torsion barrier of the C₁₀–C₁₁ bond in the other direction. Calculations of spectroscopic properties of the 11-cis, all-trans, and curl-shaped isomers provide useful data for future identification of the new retinal PSB isomer. Circular dichroism (CD) spectroscopy might be used to distinguish between the retinal PSB isomers. The potential energy surface for the orientation of the β-ionone ring of the 11-cis retinal PSB reveals three minima depending on the torsion angle of the β-ionone ring. Two of the minima correspond to 6-s-cis configurations and one has the β-ionone ring in 6-s-trans position. The calculated CD spectra for the two 6-s-cis configurations differ significantly indicating that the sign of the β-ionone ring torsion angle could be determined using CD spectroscopy. Calculations of the CD spectra suggest that a flip of the β-ionone ring might occur during the first 1 ps of the photoreaction. Rhodopsin has a negative torsion angle for the β-ionone ring, whereas the change in the sign of the first peak in the experimental CD spectrum for bathorhodopsin could suggest that it has a positive torsion angle for the β-ionone ring. Calculated nuclear magnetic resonance (NMR) shielding constants and infrared (IR) spectra are also reported for the retinal PSB isomers. Figure The figure shows the optimized molecular structure of the curl-shaped retinal isomer. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.