ω‐Turn: A novel β‐turn mimic in globular proteins stabilized by main‐chain to side‐chain CH···O interaction

Mimicry of structural motifs is a common feature in proteins. The 10‐membered hydrogen‐bonded ring involving the main‐chain C?O in a β‐turn can be formed using a side‐chain carbonyl group leading to Asx‐turn. We show that the N? H component of hydrogen bond can be replaced by a C^γ‐H group in the side chain, culminating in a nonconventional C? H···O interaction. Because of its shape this β‐turn mimic is designated as ω‐turn, which is found to occur ～three times per 100 residues. Three residues (i to i + 2) constitute the turn with the C? H···O interaction occurring between the terminal residues, constraining the torsion angles ?_{i + 1}, ψ_{i + 1}, ?_{i + 2} and χ′_{1(i + 2)} (using the interacting C^γ atom). Based on these angles there are two types of ω‐turns, each of which can be further divided into two groups. C^β‐branched side‐chains, and Met and Gln have high propensities to occur at i + 2; for the last two residues the carbonyl oxygen may participate in an additional interaction involving the S and amino group, respectively. With Cys occupying the i + 1 position, such turns are found in the metal‐binding sites. N‐linked glycosylation occurs at the consensus pattern Asn‐Xaa‐Ser/Thr; with Thr at i + 2, the sequence can adopt the secondary structure of a ω‐turn, which may be the recognition site for protein modification. Location between two β‐strands is the most common occurrence in protein tertiary structure, and being generally exposed ω‐turn may constitute the antigenic determinant site. It is a stable scaffold and may be used in protein engineering and peptide design. Proteins 2015; 83:203–214. © 2014 Wiley Periodicals, Inc.     

Increasing protein stability by improving beta‐turns

Our goal was to gain a better understanding of how protein stability can be increased by improving β‐turns. We studied 22 β‐turns in nine proteins with 66–370 residues by replacing other residues with proline and glycine and measuring the stability. These two residues are statistically preferred in some β‐turn positions. We studied: Cold shock protein B (CspB), Histidine‐containing phosphocarrier protein, Ubiquitin, Ribonucleases Sa2, Sa3, T1, and HI, Tryptophan synthetase α‐subunit, and Maltose binding protein. Of the 15 single proline mutations, 11 increased stability (Average = 0.8 ± 0.3; Range = 0.3–1.5 kcal/mol), and the stabilizing effect of double proline mutants was additive. On the basis of this and our previous work, we conclude that proteins can generally be stabilized by replacing nonproline residues with proline residues at the i + 1 position of Type I and II β‐turns and at the i position in Type II β‐turns. Other turn positions can sometimes be used if the φ angle is near ?60° for the residue replaced. It is important that the side chain of the residue replaced is less than 50% buried. Identical substitutions in β‐turns in related proteins give similar results. Proline substitutions increase stability mainly by decreasing the entropy of the denatured state. In contrast, the large, diverse group of proteins considered here had almost no residues in β‐turns that could be replaced by Gly to increase protein stability. Improving β‐turns by substituting Pro residues is a generally useful way of increasing protein stability. Proteins 2009. © 2009 Wiley‐Liss, Inc.     

Redesigning the type II' β‐turn in green fluorescent protein to type I': Implications for folding kinetics and stability

Both Type I' and Type II' β‐turns have the same sense of the β‐turn twist that is compatible with the β‐sheet twist. They occur predominantly in two residue β‐hairpins, but the occurrence of Type I' β‐turns is two times higher than Type II' β‐turns. This suggests that Type I' β‐turns may be more stable than Type II' β‐turns, and Type I' β‐turn sequence and structure can be more favorable for protein folding than Type II' β‐turns. Here, we redesigned the native Type II' β‐turn in GFP to Type I' β‐turn, and investigated its effect on protein folding and stability. The Type I' β‐turns were designed based on the statistical analysis of residues in natural Type I' β‐turns. The substitution of the native “GD” sequence of i+1 and i+2 residues with Type I' preferred “(N/D)G” sequence motif increased the folding rate by 50% and slightly improved the thermodynamic stability. Despite the enhancement of in vitro refolding kinetics and stability of the redesigned mutants, they showed poor soluble expression level compared to wild type. To overcome this problem, i and i + 3 residues of the designed Type I' β‐turn were further engineered. The mutation of Thr to Lys at i + 3 could restore the in vivo soluble expression of the Type I' mutant. This study indicates that Type II' β‐turns in natural β‐hairpins can be further optimized by converting the sequence to Type I'. Proteins 2014; 82:2812–2822. © 2014 Wiley Periodicals, Inc.     

In silico platform for predicting and initiating β‐turns in a protein at desired locations

Numerous studies have been performed for analysis and prediction of β‐turns in a protein. This study focuses on analyzing, predicting, and designing of β‐turns to understand the preference of amino acids in β‐turn formation. We analyzed around 20,000 PDB chains to understand the preference of residues or pair of residues at different positions in β‐turns. Based on the results, a propensity‐based method has been developed for predicting β‐turns with an accuracy of 82%. We introduced a new approach entitled “Turn level prediction method,” which predicts the complete β‐turn rather than focusing on the residues in a β‐turn. Finally, we developed BetaTPred3, a Random forest based method for predicting β‐turns by utilizing various features of four residues present in β‐turns. The BetaTPred3 achieved an accuracy of 79% with 0.51 MCC that is comparable or better than existing methods on BT426 dataset. Additionally, models were developed to predict β‐turn types with better performance than other methods available in the literature. In order to improve the quality of prediction of turns, we developed prediction models on a large and latest dataset of 6376 nonredundant protein chains. Based on this study, a web server has been developed for prediction of β‐turns and their types in proteins. This web server also predicts minimum number of mutations required to initiate or break a β‐turn in a protein at specified location of a protein. Proteins 2015; 83:910–921. © 2015 Wiley Periodicals, Inc.     

The crystal structure of Z‐(Aib)10‐OH at 0.65 Å resolution: three complete turns of 310‐helix

The synthetic peptide Z‐(Aib)₁₀‐OH was crystallized from hot methanol by slow evaporation. The crystal used for data collection reflected synchrotron radiation to sub‐atomic resolution, where the bonding electron density becomes visible between the non‐hydrogen atoms. Crystals belong to the centrosymmetric space group P . Both molecules in the asymmetric unit form regular 3₁₀‐helices. All residues in each molecule possess the same handedness, which is in contrast to all other crystal structure determined to date of longer Aib‐homopeptides. These other peptides are C‐terminal protected by OtBu or OMe. In these cases, because of the missing ability of the C‐terminal protection group to form a hydrogen bond to the residue i‐3, the sense of the helix is reversed in the last residue. Here, the C‐terminal OH‐groups form hydrogen bonds to the residues i‐3, in part mediated by water molecules. This makes Z‐(Aib)₁₀‐OH an Aib‐homopeptide with three complete 3₁₀‐helical turns in spite of the shorter length it has compared with Z‐(Aib)₁₁‐OtBu, the only homopeptide to date with three complete turns.     

Stereochemistry of Schellman motifs in peptides: Crystal structure of a hexapeptide with a C‐terminus 6 → 1 hydrogen bond

The Schellman motif is a widely observed helix terminating structural motif in proteins, which is generated when the C‐terminus residue adopts a left‐handed helical (α_L) conformation. The resulting hydrogen‐bonding pattern involves the formation of an intramolecular 6 → 1 interaction. This helix terminating motif is readily mimicked in synthetic helical peptides by placing an achiral residue at the penultimate position of the sequence. Thus far, the Schellman motif has been characterized crystallographically only in peptide helices of length 7 residues or greater. The structure of the hexapeptide Boc–Pro–Aib–Gly–Leu–Aib–Leu–OMe in crystals reveal a short helical stretch terminated by a Schellman motif, with the formation of 6 → 1 C‐terminus hydrogen bond. The crystals are in the space group P2₁2₁2₁ with a = 18.155(3) Å, b = 18.864(8) Å, c = 11.834(4) Å, and Z = 4 . The final R₁ and wR₂ values are 7.68 and 14.6%, respectively , for 1524 observed reflections [F_o ≥ 3ς(F_o)]. A 6 → 1 hydrogen bond between Pro(1)CO · · · Leu(6)NH and a 5 → 2 hydrogen bond between Aib(2)CO · · · Aib(5)NH are observed. An analysis of the available oligopeptides having an achiral Aib residue at the penultimate position suggests that chain length and sequence effects may be the other determining factors in formation of Schellman motifs. © 1999 John Wiley & Sons, Inc. Biopoly 50: 13–22, 1999     

Characterization of a type of β‐bend ribbon spiral generated by the repeating (Xaa–Yaa–Aib–Pro) motif: The solution structure of harzianin HC IX,a 14‐residue peptaibol forming voltage‐dependent ion channels

The three‐dimensional solution structure of harzianin HC IX, a peptaibol antibiotic isolated from the fungus Trichoderma harzianum, was determined using CD, homonuclear, and heteronuclear two‐dimensional nmr spectroscopy combined with molecular modeling. This 14‐residue peptide, Ac Aib¹ Asn² Leu³ Aib⁴ Pro⁵ Ala⁶ Ile⁷ Aib⁸ Pro⁹ Iva¹⁰ Leu¹¹ Aib¹² Pro¹³ Leuol¹⁴ (Aib, α‐aminoisobutyric acid; Iva, isovaline; Leuol, leucinol), is a main representative of a short‐sequence peptaibol class characterized by an acetylated N‐terminus, a C‐terminal amino alcohol, and the presence of three Aib‐L ‐Pro motifs at positions 4–5, 8–9, and 12–13, separated by two dipeptide units. In spite of a lower number of residues, compared to the 18/20‐residue peptaibols such as alamethicin, harzianin HC IX exhibits remarkable membrane‐perturbing properties. It interacts with phospholipid bilayers, increasing their permeability and forming voltage‐gated ion channels through a mechanism slightly differing from that proposed for alamethicin. Sequence‐specific ¹H‐ and ¹³C‐nmr assignments and conformational nmr parameters (³J_NHCαH coupling constants, quantitative nuclear Overhauser enhancement data, temperature coefficients of amide and carbonyl groups, NH–ND exchange rates) were obtained in methanol solution. Sixty structures were calculated based on 98 interproton distance restraints and 6 Φ dihedral angle restraints, using high temperature restrained molecular dynamics and energy minimization. Thirty‐seven out of the sixty generated structures were consistent with the nmr data and were convergent. The peptide backbone consists in a ribbon of overlapping β‐turns twisted into a continuous spiral from Asn² to Leuol¹⁴ and forming a 26 Å long helix‐like structure. This structure is slightly amphipathic, with the three Aib–Pro motifs aligned on the less hydrophobic face of the spiral where the Asn² side chain is also present, while the more hydrophobic bulky side chains of leucines, isoleucine, isovaline, and leucinol are located on the concave side. The repetitive (Xaa–Yaa–Aib–Pro) tetrapeptide subunit, making up the peptide sequence, is characterized by four sets of (Φ,Ψ) torsional angles, with the following mean values: Φ_i = −90°, Ψ_i = −27°; Φ_i+1 = −98°, Ψ_i+1 = −17°; Φ_i+2 = −49°, Ψ_i+2 = −50°; Φ_i+3 = −78°, Ψ_i+3 = +3°. We term this particular structure, specifically occurring in the case of (Xaa–Yaa–Aib–Pro)_n sequences, the (Xaa–Yaa–Aib–Pro)‐β‐bend ribbon spiral. It is stabilized by 4 → 1 intramolecular hydrogen bonds and differs from both the canonical 3₁₀‐helix made of a succession of type III β‐turns and from the β‐bend ribbon spiral that has been described in the case of (Aib–Pro)n peptide segments. © 1999 John Wiley & Sons, Inc. Biopoly 50: 71–85, 1999     

Bridging of anions by hydrogen bonds in nest motifs and its significance for Schellman loops and other larger motifs within proteins

The nest is a protein motif of three consecutive amino acid residues with dihedral angles 1,2‐α_Rα_L (RL nests) or 1,2‐α_Lα_R (LR nests). Many nests form a depression in which an anion or δ‐negative acceptor atom is bound by hydrogen bonds from the main chain NH groups. We have determined the extent and nature of this bridging in a database of protein structures using a computer program written for the purpose. Acceptor anions are bound by a pair of bridging hydrogen bonds in 40% of RL nests and 20% of LR nests. Two thirds of the bridges are between the NH groups at Positions 1 and 3 of the motif (N1N3‐bridging)—which confers a concavity to the nest; one third are of the N2N3 type—which does not. In bridged LR nests N2N3‐bridging predominates (14% N1N3: 75% N2N3), whereas in bridged RL nests the reverse is true (69% N1N3: 25% N2N3). Most bridged nests occur within larger motifs: 45% in (hexapeptide) Schellman loops with an additional 4 → 0 hydrogen bond (N1N3), 11% in Schellman loops with an additional 5 → 1 hydrogen bond (N2N3), 12% in a composite structure including a type 1β‐bulge loop and an asx‐ or ST‐ motif (N1N3)—remarkably homologous to the N1N3‐bridged Schellman loop—and 3% in a composite structure including a type 2β‐bulge loop and an asx‐motif (N2N3). A third hydrogen bond is a previously unrecognized feature of Schellman loops as those lacking bridged nests have an additional 4 → 0 hydrogen bond. Proteins 2014; 82:3023–3031. © 2014 Wiley Periodicals, Inc.     

Disorder and order in unfolded and disordered peptides and proteins: A view derived from tripeptide conformational analysis. II. Tripeptides with short side chains populating asx and β‐type like turn conformations

In the preceding paper, we found that ensembles of tripeptides with long or bulky chains can include up to 20% of various turns. Here, we determine the structural and thermodynamic characteristics of GxG peptides with short polar and/or ionizable central residues (D, N, C), whose conformational distributions exhibit higher than average percentage (>20%) of turn conformations. To probe the side‐chain conformations of these peptides, we determined the ³J(H^α,H^β) coupling constants and derived the population of three rotamers with χ₁‐angles of ?60°, 180° and 60°, which were correlated with residue propensities by DFT‐calculations. For protonated GDG, the rotamer distribution provides additional evidence for asx‐turns. A comparison of vibrational spectra and NMR coupling constants of protonated GDG, ionized GDG, and the protonated aspartic acid dipeptide revealed that side chain protonation increases the pPII content at the expense of turn populations. The charged terminal groups, however, have negligible influence on the conformational properties of the central residue. Like protonated GDG, cationic GCG samples asx‐turns to a significant extent. The temperature dependence of the UVCD spectra and ³J(H^NH^α) constants suggest that the turn populations of GDG and GNG are practically temperature‐independent, indicating enthalpic and entropic stabilization. The temperature‐independent J‐coupling and UVCD spectra of GNG require a three‐state model. Our results indicate that short side chains with hydrogen bonding capability in GxG segments of proteins may serve as hinge regions for establishing compact structures of unfolded proteins and peptides. Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Protein prosthesis: β‐peptides as reverse‐turn surrogates

The introduction of non‐natural modules could provide unprecedented control over folding/unfolding behavior, conformational stability, and biological function of proteins. Success requires the interrogation of candidate modules in natural contexts. Here, expressed protein ligation is used to replace a reverse turn in bovine pancreatic ribonuclease (RNase A) with a synthetic β‐dipeptide: β²‐homoalanine–β³‐homoalanine. This segment is known to adopt an unnatural reverse‐turn conformation that contains a 10‐membered ring hydrogen bond, but one with a donor–acceptor pattern opposite to that in the 10‐membered rings of natural reverse turns. The RNase A variant has intact enzymatic activity, but unfolds more quickly and has diminished conformational stability relative to native RNase A. These data indicate that hydrogen‐bonding pattern merits careful consideration in the selection of beneficial reverse‐turn surrogates.     

Expanded turn conformations: characterization and sequence-structure correspondence in alpha-turns with implications in helix folding

Like the beta-turns, which are characterized by a limiting distance between residues two positions apart (i, i+3), a distance criterion (involving residues at positions i and i+4) is used here to identify alpha-turns from a database of known protein structures. At least 15 classes of alpha-turns have been enumerated based on the location in the phi,psi space of the three central residues (i+1 to i+3)-one of the major being the class AAA, where the residues occupy the conventional helical backbone torsion angles. However, moving towards the C-terminal end of the turn, there is a shift in the phi,psi angles towards more negative phi, such that the electrostatic repulsion between two consecutive carbonyl oxygen atoms is reduced. Except for the last position (i+4), there is not much similarity in residue composition at different positions of hydrogen and non-hydrogen bonded AAA turns. The presence or absence of Pro at i+1 position of alpha- and beta-turns has a bearing on whether the turn is hydrogen-bonded or without a hydrogen bond. In the tertiary structure, alpha-turns are more likely to be found in beta-hairpin loops. The residue composition at the beginning of the hydrogen bonded AAA alpha-turn has similarity with type I beta-turn and N-terminal positions of helices, but the last position matches with the C-terminal capping position of helices, suggesting that the existence of a "helix cap signal" at i+4 position prevents alpha-turns from growing into helices. Our results also provide new insights into alpha-helix nucleation and folding.     

Structural and sequence features of two residue turns in beta‐hairpins

Beta‐turns in beta‐hairpins have been implicated as important sites in protein folding. In particular, two residue β‐turns, the most abundant connecting elements in beta‐hairpins, have been a major target for engineering protein stability and folding. In this study, we attempted to investigate and update the structural and sequence properties of two residue turns in beta‐hairpins with a large data set. For this, 3977 beta‐turns were extracted from 2394 nonhomologous protein chains and analyzed. First, the distribution, dihedral angles and twists of two residue turn types were determined, and compared with previous data. The trend of turn type occurrence and most structural features of the turn types were similar to previous results, but for the first time Type II turns in beta‐hairpins were identified. Second, sequence motifs for the turn types were devised based on amino acid positional potentials of two‐residue turns, and their distributions were examined. From this study, we could identify code‐like sequence motifs for the two residue beta‐turn types. Finally, structural and sequence properties of beta‐strands in the beta‐hairpins were analyzed, which revealed that the beta‐strands showed no specific sequence and structural patterns for turn types. The analytical results in this study are expected to be a reference in the engineering or design of beta‐hairpin turn structures and sequences. Proteins 2014; 82:1721–1733. © 2014 Wiley Periodicals, Inc.     

Computationally designed β‐turn foldamers of γ‐peptides based on 2‐(aminomethyl)cyclohexanecarboxylic acid

The γ‐peptide β‐turn structures have been designed computationally by the combination of chirospecific γ 2 , 3 ‐residues of 2‐(aminomethyl)cyclohexanecarboxylic acid (γAmc₆) with a cyclohexyl constraint on the C^α?C^β bond using density functional methods in water. The chirospecific γAmc₆ dipeptide with the (2S,3S)‐(2R,3R) configurations forms a stable turn structure in water, resembling a type II′ turn of α‐peptides, which can be used as a β‐turn motif in β‐hairpins of Ala‐based α‐peptides. The γAmc₆ dipeptide with homochiral (2S,3S)‐(2S,3S) configurations but different cyclohexyl puckerings shows the capability to be incorporated into one of two β‐turn motifs of gramicidin S. The overall structure of this gramicidin S analogue is quite similar to the native gramicidin S with the same patterns and geometries of hydrogen bonds. Our calculated results and the recently observed results may imply the wider applicability of chirospecific γ‐peptides with a cyclohexyl constraint on the backbone to form various peptide foldamers. © 2012 Wiley Periodicals, Inc. Biopolymers 97:1018–1025, 2012.     

Sparsely populated residue conformations in protein structures: Revisiting “experimental” Ramachandran maps

The Ramachandran map clearly delineates the regions of accessible conformational (φ–ψ) space for amino acid residues in proteins. Experimental distributions of φ, ψ values in high‐resolution protein structures, reveal sparsely populated zones within fully allowed regions and distinct clusters in apparently disallowed regions. Conformational space has been divided into 14 distinct bins. Residues adopting these relatively rare conformations are presented and amino acid propensities for these regions are estimated. Inspection of specific examples in a completely “arid”, fully allowed region in the top left quadrant establishes that side‐chain and backbone interactions may provide the energetic compensation necessary for populating this region of φ–ψ space. Asn, Asp, and His residues showed the highest propensities in this region. The two distinct clusters in the bottom right quadrant which are formally disallowed on strict steric considerations correspond to the gamma turn (C7 axial) conformation (Bin 12 ) and the i + 1 position of Type II′ β turns (Bin 13) . Of the 516 non‐Gly residues in Bin 13 , 384 occupied the i + 1 position of Type II′ β turns. Further examination of these turn segments revealed a high propensity to occur at the N‐terminus of helices and as a tight turn in β hairpins. The β strand–helix motif with the Type II′ β turn as a connecting element was also found in as many as 57 examples. Proteins 2014; 82:1101–1112. © 2013 Wiley Periodicals, Inc.     

An amino acid code for β‐sheet packing structure

To understand the relationship between protein sequence and structure, this work extends the knob‐socket model in an investigation of β‐sheet packing. Over a comprehensive set of β‐sheet folds, the contacts between residues were used to identify packing cliques: sets of residues that all contact each other. These packing cliques were then classified based on size and contact order. From this analysis, the two types of four‐residue packing cliques necessary to describe β‐sheet packing were characterized. Both occur between two adjacent hydrogen bonded β‐strands. First, defining the secondary structure packing within β‐sheets, the combined socket or XY:HG pocket consists of four residues i, i+2 on one strand and j, j+2 on the other. Second, characterizing the tertiary packing between β‐sheets, the knob‐socket XY:H+B consists of a three‐residue XY:H socket (i, i+2 on one strand and j on the other) packed against a knob B residue (residue k distant in sequence). Depending on the packing depth of the knob B residue, two types of knob‐sockets are found: side‐chain and main‐chain sockets. The amino acid composition of the pockets and knob‐sockets reveal the sequence specificity of β‐sheet packing. For β‐sheet formation, the XY:HG pocket clearly shows sequence specificity of amino acids. For tertiary packing, the XY:H+B side‐chain and main‐chain sockets exhibit distinct amino acid preferences at each position. These relationships define an amino acid code for β‐sheet structure and provide an intuitive topological mapping of β‐sheet packing. Proteins 2014; 82:2128–2140. © 2014 Wiley Periodicals, Inc.     

Solution structure of the calmodulin‐like C‐terminal domain of Entamoeba α‐actinin2

Cell motility is dependent on a dynamic meshwork of actin filaments that is remodelled continuously. A large number of associated proteins that are severs, cross‐links, or caps the filament ends have been identified and the actin cross‐linker α‐actinin has been implied in several important cellular processes. In Entamoeba histolytica, the etiological agent of human amoebiasis, α‐actinin is believed to be required for infection. To better understand the role of α‐actinin in the infectious process we have determined the solution structure of the C‐terminal calmodulin‐like domain using NMR. The final structure ensemble of the apo form shows two lobes, that both resemble other pairs of calcium‐binding EF‐hand motifs, connected with a mobile linker. Proteins 2016; 84:461–466. © 2016 Wiley Periodicals, Inc.     

Asparagine and glutamine differ in their propensities to form specific side chain-backbone hydrogen bonded motifs in proteins

Short range side chain‐backbone hydrogen bonded motifs involving Asn and Gln residues have been identified from a data set of 1370 protein crystal structures (resolution ≤ 1.5 Å). Hydrogen bonds involving residues i ? 5 to i + 5 have been considered. Out of 12,901 Asn residues, 3403 residues (26.4%) participate in such interactions, while out of 10,934 Gln residues, 1780 Gln residues (16.3%) are involved in these motifs. Hydrogen bonded ring sizes (C_n, where n is the number of atoms involved), directionality and internal torsion angles are used to classify motifs. The occurrence of the various motifs in the contexts of protein structure is illustrated. Distinct differences are established between the nature of motifs formed by Asn and Gln residues. For Asn, the most highly populated motifs are the C₁₀ (CO^δ_i …NH_{i + 2}), C₁₃ (CO^δ_i …NH_{i + 3}) and C₁₇ (N^δH_i …CO_{i ? 4}) structures. In contrast, Gln predominantly forms C₁₆ (CO^ε_i …NH_{i ? 3}), C₁₂ (N^εH_i …CO_{i ? 2}), C₁₅ (N^εH_i …CO_{i ? 3}) and C₁₈ (N^εH_i …CO_{i ? 4}) motifs, with only the C₁₈motif being analogous to the Asn C₁₇structure. Specific conformational types are established for the Asn containing motifs, which mimic backbone β‐turns and α‐turns. Histidine residues are shown to serve as a mimic for Asn residues in side chain‐backbone hydrogen bonded ring motifs. Illustrative examples from protein structures are considered. Proteins 2012; © 2011 Wiley Periodicals, Inc.     

Delineation of a new structural motif involving NHN γ-turn

Macromolecules are characterized by distinctive arrangement of hydrogen bonds. Different patterns of hydrogen bonds give rise to distinct and stable structural motifs. An analysis of 4114 non-redundant protein chains reveals the existence of a three-residue, (i − 1) to (i + 1), structural motif, having two hydrogen-bonded five-membered pseudo rings (the first, an N H···OC involving the first residue, and the second being N H∙∙∙N involving the last two residues), separated by a peptide bond. There could be an additional hydrogen bond between the side-chain at (i-1) and the main-chain NH of (i + 1). The average backbone torsion angles of −76(±21)° and – 12(±17)° at i creates a tight turn in the polypeptide chain, akin to a γ-turn. Indeed, a search of three-residue fragments with restriction on the terminal C^α···C^α distance and the existence of the two pseudo rings on either side revealed the presence 14 846 cases of a variant, termed NHN γ-turn, distinct from the NHO γ-turn (2032 cases) that has traditionally been characterized by the presence of NHO hydrogen bond linking the terminal main-chain atoms. As in the latter, the newly identified γ-turns are also of two types—classical and inverse, occurring in the ratio of 1:6. The propensities of residues to occur in these turns and their secondary structural features have been enumerated. An understanding of these turns would be useful for structure prediction and loop modeling, and may serve as models to represent some of the unfolded state or disordered region in proteins.     

An automated method for consistent helix assignment using turn information

A new automated helix assignment method is presented that leads to a more consistent definition of the helix termini, especially of the helix C-terminus. The method assigns a helix to segments of protein chain where adjacent helical turn structures are observed, capped by specific distorted turn types (e.g., open helical turns without a hydrogen bond) or capping motifs (e.g., the Schellman motif). Helix termini are detected by observing the behavior of the NH group in N-termini and the CO group in C-termini; in each case, the respective group must be free to interact with hydrogen bonding partners outside of the putative helix for a helix terminus to be assigned. The presented assignment method and SHAFT-assigned helices are part of Secbase and are made available with Relibase+ 3.0 and the free web version of Relibase 3.0. The method can also be used for the helix assignments of additional protein structures.     

Isostructural unbranched alkyl‐chains as tools for stabilizing β‐turn structure

To investigate the structural role played by isostructural unbranched alkyl‐chains on the conformational ensemble and stability of β‐turn structures, the conformational properties of a designed model peptide: Plm‐Pro‐Gly‐Pda ( 1 , Plm: H₃C—(CH₂)₁₄—CONH—; Pda: —CONH— (CH₂)₁₄—CH₃) have been examined and compared with the parent peptide: Boc‐Pro‐Gly‐NHMe ( 2 , Boc: tert‐butoxycarbonyl; NHMe: N‐methylamide). The characteristic ¹³C NMR chemical‐shifts of the Pro C^β and C^γ resonances ascertained the incidence of an all‐trans peptide‐bond in low polarity deuterochloroform solution. Using FTIR and ¹H NMR spectroscopy, we establish that apolar alkyl‐chains flanking a β‐turn promoting Pro‐Gly sequence impart definite incremental stability to the well‐defined hydrogen‐bonded structure. The assessment of ¹H NMR derived thermodynamic parameters of the hydrogen‐bonded amide‐NHs via variable temperature indicate that much weaker hydrophobic interactions do contribute to the stability of folded reverse turn structures. The far‐UV CD spectral patterns of 1 and 2 in 2,2,2‐trifluoroethanol are consistent with Pro‐Gly specific type II β‐turn structure, concomitantly substantiate that the flanking alkyl‐chains induce substantial bias in enhanced β‐turn populations. In view of structural as well as functional importance of the Pro‐Gly mediated secondary structures, besides biochemical and biological significance of proteins lipidation via myristoylation or palmytoilation, we highlight potential convenience of the unbranched Plm and Pda moieities not only as main‐chain N‐ and C‐terminal protecting groups but also to mimic and stabilize specific isolated secondary and supersecondary structural components frequently observed in proteins and polypeptides. © 2013 Wiley Periodicals, Inc. Biopolymers 99: 419–426, 2013.