Crystal Conformation Research Articles

Effect of deformation ratio of flow-induced crystallization on polyethylene in crystalline behavior, thermal stability, and tensile loading: A molecular dynamics simulation

Since the physical and mechanical properties of semicrystalline polymers strongly depend on crystalline morphology, understanding the correlation between the crystallization kinetics and crystalline structure of polythene is beneficial for their rational processing and applications. In this study, united-atom (UA) molecular dynamics (MD) simulations were employed to elucidate the variations in crystalline structure, phase transition temperatures, and uniaxial tensile deformation among polyethylene (PE) with randomly oriented and flow-induced crystallization (FIC). The results showed that crystallization occurs in areas with low potential energy. For the FIC process, the entanglement parameter and interplanar spacing was less than for randomly oriented crystallization, and the crystallinity was higher. The relationship between the deformation and the crystallinity of PE with randomly oriented crystallization was expressed, which indicates that the crystalline structure promoted elongation at break of PE in tensile deformation. As PE was oriented to crystallize during the FIC process, the polymer stiffness is positively correlated with the deformation ratio of the PE model, so crystal conformation systems with higher deformation ratios produce higher ultimate stresses in tensile loading. From the visualization results, the crystalline phase was stable and does not easily transform into an amorphous phase during tensile loading. Additional results show that the fracture region of PE at tensile fracture tends to develop in the amorphous phase under tensile loading. Energy analysis showed that the elastic and yield regions were mainly dominated by intra-chain bonds, angles, and dihedral motion of PE, whereas interchain nonbonded interactions mainly dominated strain-hardening regions.

Exploring Macroscopic Dipoles of Designed Cyclic Peptide Ordered Assemblies to Harvest Piezoelectric Properties

AbstractCrystalline materials exhibiting non‐centrosymmetry and possessing substantial surface dipole moments play a critical role in piezoelectricity. Designing biocompatible self‐assembled materials with these attributes is particularly challenging when compared to inorganic materials and ceramics. In this study, we elucidate the crystal conformations of novel cyclic peptides that exhibit self‐assembly into tubular structures characterized by unidirectional hydrogen bonding and piezoelectric properties. Unlike cyclic peptides derived from alternating L‐ and D‐amino acids, those derived from new δ‐amino acids demonstrate the formation of self‐assembled tubes with unidirectional hydrogen bonds. Further, the tightly packed tubular assemblies and higher macrodipole moments result in superior piezoelectric coefficients compared to peptides with lower macrodipole moments. Our findings underscore the potential for designing cyclic peptides with unidirectional hydrogen bonds, thereby paving the way for their application in design of biocompatible piezo‐ and ferroelectric materials.

Exploring Macroscopic Dipoles of Designed Cyclic Peptide Ordered Assemblies to Harvest Piezoelectric Properties.

Crystalline materials exhibiting non-centrosymmetry and possessing substantial surface dipole moments play a critical role in piezoelectricity. Designing biocompatible self-assembled materials with these attributes is particularly challenging when compared to inorganic materials and ceramics. In this study, we elucidate the crystal conformations of novel cyclic peptides that exhibit self-assembly into tubular structures characterized by unidirectional hydrogen bonding and piezoelectric properties. Unlike cyclic peptides derived from alternating L- and D-amino acids, those derived from new δ-amino acids demonstrate the formation of self-assembled tubes with unidirectional hydrogen bonds. Further, the tightly packed tubular assemblies and higher macrodipole moments result in superior piezoelectric coefficients compared to peptides with lower macrodipole moments. Our findings underscore the potential for designing cyclic peptides with unidirectional hydrogen bonds, thereby paving the way for their application in design of biocompatible piezo- and ferroelectric materials.

Stabilizing Bifurcated Hydrogen Bond in 8-Aminoquinoline Appended Peptides.

The hydrogen bonding interaction between an amide N-H and the amide N of the preceding residue is prevalent in proline-containing proteins and peptides. However, the N-H⋅⋅⋅N hydrogen bonding interaction is rare in non-prolyl natural peptides due to restricted dihedral angles. Herein, we stabilize this type of interaction in 8-aminoquinoline appended non-prolyl peptides through bifurcated N⋅⋅⋅H⋅⋅⋅N hydrogen bond. The 8-aminoquinoline-incorporated model peptides 2 a-i were designed, synthesized, and the crystal structures of 2 a-c and 2 i were solved. Analysis of crystal data reveals that the amide N-H of aminoquinoline is involved in bifurcated hydrogen bonding interaction with the nitrogen of the preceding amino acid residue and the nitrogen in quinoline. Analysis of crystal packing, Hirshfeld surface and fingerprint plots confirms that the intermolecular O⋅⋅⋅H contacts significantly contribute to stabilizing bifurcated N⋅⋅⋅H⋅⋅⋅N hydrogen bonding interaction. Furthermore, NMR experiments and CD spectroscopy were conducted to examine the preferred conformation in solution, and the data corroborate with the crystal structure conformation.

Green synthesis of bimetallic CuO@NiO nanocomposite for the removal of glyphosate

Green synthesis of photocatalyst bimetallic CuO@NiO nanocomposite for eliminating organic hazardous glyphosate (Gly) solution has been introduced. The nanocomposite has been successfully developed from mango (Mangifera indica L.) peel extracted solution by a simultaneous reduction process. HRTEM, XRD, and EDX have also been used to explore the nanostructure, crystal conformation, and chemical compositions of CuO@NiO. Using UV-vis spectrometer, we have observed the photo-catalytic activity and kinetic removal rate constant of CuO@NiO in terms of glyphosate elimination under UV light illumination. Compared with pure CuO and NiO nanoparticles, CuO@NiO displayed improved and enhanced photocatalytic activity. This work demonstrates an eco-friendly, low-cost material with high efficiency for removing Gly, which has applications in environmental protection.

Comprehensive Evaluation of 10 Docking Programs on a Diverse Set of Protein-Cyclic Peptide Complexes.

Cyclic peptides have emerged as a highly promising class of therapeutic molecules owing to their favorable pharmacokinetic properties, including stability and permeability. Currently, many clinically approved cyclic peptides are derived from natural products or their derivatives, and the development of molecular docking techniques for cyclic peptide discovery holds great promise for expanding the applications and potential of this class of molecules. Given the availability of numerous docking programs, there is a pressing need for a systematic evaluation of their performance, specifically on protein-cyclic peptide systems. In this study, we constructed an extensive benchmark data set called CPSet, consisting of 493 protein-cyclic peptide complexes. Based on this data set, we conducted a comprehensive evaluation of 10 docking programs, including Rosetta, AutoDock CrankPep, and eight protein-small molecule docking programs (i.e., AutoDock, AudoDock Vina, Glide, GOLD, LeDock, rDock, MOE, and Surflex). The evaluation encompassed the assessment of the sampling power, docking power, and scoring power of these programs. The results revealed that all of the tested protein-small molecule docking programs successfully sampled the binding conformations when using the crystal conformations as the initial structures. Among them, rDock exhibited outstanding performance, achieving a remarkable 94.3% top-100 sampling success rate. However, few programs achieved successful predictions of the binding conformations using tLEaP-generated conformations as the initial structures. Within this scheme, AutoDock CrankPep yielded the highest top-100 sampling success rate of 29.6%. Rosetta's scoring function outperformed the others in selecting optimal conformations, resulting in an impressive top-1 docking success rate of 87.6%. Nevertheless, all the tested scoring functions displayed limited performance in predicting binding affinity, with MOE@Affinity dG exhibiting the highest Pearson's correlation coefficient of 0.378. It is therefore suggested to use an appropriate combination of different docking programs for given tasks in real applications. We expect that this work will offer valuable insights into selecting the appropriate docking programs for protein-cyclic peptide complexes.

The N-terminal domain of Type IV-A1 CRISPR-associated DinG is vulnerable to proteolysis.

CasDinG is an ATP-dependent 5'-3' DNA helicase essential for bacterial Type IV-A1 CRISPR associated immunity. CasDinG contains an essential N-terminal domain predicted to bind DNA. To better understand the role of the N-terminal domain, we attempted to co-crystallize CasDinG with DNA substrates. We successfully crystallized CasDinG in a tightly packed, crystal conformation with previously unobserved unit cell dimensions. However, the structure lacked electron density for a bound DNA substrate and the CasDinG N-terminal domain. Additionally, the tight crystal packing disallowed space for the N-terminal domain, indicating that the N-terminal domain was proteolyzed before crystallization. Follow up experiments revealed that the N-terminal domain of CasDinG is proteolyzed after a few days at room temperature, but is protected from proteolysis at 4°C. These data provide a distinct x-ray crystal structure of CasDinG and indicate the essential N-terminal domain of CasDinG is prone to proteolysis.

Uncovering New Conformational States of the Substrate Binding Pocket of LSD1 Potential for Inhibitor Design via Funnel Metadynamics.

Lysine-specific demethylase 1 (LSD1) is a promising therapeutic target for cancer therapy. So far, over 80 crystal structures of LSD1 in different complex states have been deposited in the Protein Data Bank, which are valuable resources for performing structure-based drug design. However, among all of the crystal structures of LSD1, the substrate binding pocket, which is the most efficient druggable site for designing LSD1 inhibitors at present, is very similar no matter whether LSD1 is in the apo or any holo forms, which is inconsistent with its versatile demethylase functions. To investigate whether the substrate binding pocket is rigid or exhibits other representative conformations different from the crystal conformations that are feasible for designing new LSD1 inhibitors, we performed funnel metadynamics simulations to study the conformation dynamics of LSD1 in the binding process of two effective LSD1 inhibitors (CC-90011 and 6X0, CC-90011 undergoing clinical trials). Our results showed that the entrance of the substrate binding pocket is very flexible. Two representative entrance conformations of LSD1 counting against binding with the substrate of histone H3 were detected, which may be used for structure-based LSD1 inhibitor design. Besides, alternative optimal binding modes and prebinding modes for both inhibitors were also detected, which depicted that the key interactions changed along with the binding process. Our results should provide great help for LSD1 inhibitor design.

Incoherent Energy Transfer between the Baseplate and FMO Protein Explored at Ideal Geometries.

The Förster resonance energy transfer (FRET) between the Fenna-Matthews-Olson (FMO) protein complex and the chlorosomal baseplate (CBP) is investigated by using an idealized model. This simplified model is based on crystal structure and molecular dynamics conformations. Some of the further input, such as the transition dipole moments, was extracted from earlier molecular-level simulations. The resulting model mimics the effects of the relative position between the CBP and the FMO complex on the corresponding FRET efficiency under ideal conditions, involving about 1.3 billion FRET calculations per investigated model. In this idealized model and employing some approximations, it is found that FRET efficiency is almost completely independent of the FMO trimer orientation (displacement, distance, and rotation), despite FMO and CBP being highly structured complexes. Even removing individual FMO BChl triples will only reduce the FRET efficiency by up to 8.6%. An FMO containing only the least efficient BChl triple will retain about 25% of the FRET efficiency of a full FMO complex. In addition to its proposed function as an energetic funnel, FMO is thus identified to act as a highly robust spatial funnel for CBP excitation harvesting, independent of the mutual CBP-FMO orientation.

Probing the Activation Mechanisms of Agonist DPI-287 to Delta-Opioid Receptor and Novel Agonists Using Ensemble-Based Virtual Screening with Molecular Dynamics Simulations.

Pain drugs targeting mu-opioid receptors face major addiction problems that have caused an epidemic. The delta-opioid receptor (DOR) has shown to not cause addictive effects when bound to an agonist. While the active conformation of the DOR in complex with agonist DPI-287 has been recently solved, there are still no FDA-approved agonists targeting it, providing the opportunity for structure-based virtual screening. In this study, the conformational plasticity of the DOR was probed using molecular dynamics (MD) simulations, identifying two representative conformations from clustering analysis. The two MD conformations as well as the crystal conformation of DOR were used to screen novel compounds from the ZINC database (17 million compounds), in which 69 drugs were picked as potential compounds based on their docking scores. Notably, 37 out of the 69 compounds were obtained from the simulated conformations. The binding stability of the 69 compounds was further investigated using MD simulations. Based on the MM-GBSA binding energy and the predicted drug properties, eight compounds were chosen as the most favorable, six of which were from the simulated conformations. Using a dynamic network model, the communication between the crystal agonist and the top eight molecules with the receptor was analyzed to confirm if these novel compounds share a similar activation mechanism to the crystal ligand. Encouragingly, docking of these eight compounds to the other two opioid receptors (kappa and mu) suggests their good selectivity toward DOR.

Crystal polymorphs and molecular conformations of fluorinated ionic liquid: 1-Ethyl-3-methylimidazolium perfluorobutanesulfonate

The phase transitions of a fluorinated ionic liquid (fIL) were examined at low temperature (LT) by X-ray diffraction and Raman spectroscopy. The fIL was 1-ethyl-3-methylimidazolium perfluorobutanesulfonate ([C2mim][PFBS]). The cation and anion possessed conformational degrees of freedom. The trans and gauche conformers of [PFBS]– coexisted in the liquid state. The LT crystal polymorph of the [C2mim][PFBS] was observed, and [C2mim][PFBS] possessed the large unit cells. The gauche’ conformer of [PFBS]– appeared, accompanying with the phase transition at 220 K.

A new avibactam Gemini quaternary ammonium salt: synthesis, self-assembly, vibrational spectra, crystal structures and DFT calculations

Avibactam ([AV]), a non-β-lactam β-lactamase inhibitor, contains a sulfonic acid group, which making separation and purification extremely difficult. The ability of the gemini quaternary ammonium salt N,N,N,N’,N’,N’-hexabutylbutanediammonium dibromide [444][Br]2 to precipitate [AV] from aqueous solutions resulted in the successful synthesis of a new avibactam gemini quaternary ammonium salt [444][AV]2. To understand the self-assembly, the transmission electron microscope (TEM) and spectra of [444] cations and [AV] anions in aqueous solution were investigated. By using a slow evaporation technique, the [444][AV]2 complex single crystal was produced from a water solvent. The title crystal structure conformation was done by single crystal X-ray diffraction (SCXRD) technique, and its belongs to the P-1 space group with the triclinic system. The thermal stability of [444][AV]2 has been investigated by TG/DSC studies, and its experimental vibrational bands (FT-IR and Raman) have been studied and assigned. The stability test indicated that [444][AV]2 exhibits outstanding physical stability under long-term and accelerated conditions. According to DFT calculations, electrostatic potential and Mulliken charges showed that [444] cations were readily adsorbed on the negatively charged [AV] anions surface sites through electrostatic attraction, and the existence of inter-molecular interactions was assessed by Hirshfeld surface of [444][AV]2 single crystal.

Helices in Peptides Containing Differentially Geminally Di‐substituted γ Amino Acid Residues

AbstractWe have explored the relative propensity of the three differentially di‐substituted γ amino acid residues, to be accommodated in all α helical conformations. We have established that small chiral hybrid peptides Boc‐Aib‐γx,x‐Aib‐Leu‐Aib‐Val‐OMe (x,x=2,2/3,3/4,4) containing single geminally di‐substituted γ amino acid residue, can adopt both handedness of helical conformation in crystals and solution. The relative ease of being accommodated in helical structures decreased in the order γ4,4>γ3,3>γ2,2. γ4,4 and γ3,3 readily adopted C12 helical conformations unlike γ2,2. γ2,2 and γ3,3 containing peptides, adopted either handedness of the helical conformation in crystals while in solution, they adopted right‐handed helical conformation. Peptide, containing γ4,4 only adopted right‐handed helical conformation both in the solid and solution states. Our studies established that position of di‐substitution along the backbone of γ amino acid residues determined their conformational preference, and thus might be used to generate diversity in the peptide conformations.

Crystal structures and solution conformations of HtrA from Helicobacter pylori reveal pH-dependent oligomeric conversion and conformational rearrangements

Helicobacter pylori is a Gram-negative microaerophilic bacterium that infects over 50 % of the world's population, making it a major risk factor for chronic gastritis, ulcer diseases of the stomach and duodenum, MALT lymphoma, and gastric cancer. The clinical consequences of H. pylori infection are closely linked with the expression of virulence factors secreted by the bacterium. One such virulence factor is high temperature requirement A (HtrA), which possesses chaperone and serine protease activity. In the host stomach, HtrA secreted from H. pylori (HpHtrA) disrupts intercellular adhesions by cleaving epithelial adhesion proteins including E-cadherin and desmoglein-2. This disruption causes intercellular junctions to open, allowing the bacterium to pass through the epithelial barrier, access the intercellular space, and colonize the gastric mucosa. HtrA proteases are well known for their structural complexity, reflected in their diverse oligomer forms and multi-tasking activities in both prokaryotes and eukaryotes. In this study, we determined crystal structures and solution conformations of HpHtrA monomer and trimer, which revealed large domain rearrangements between them. Notably, this is the first report of a monomeric structure in the HtrA family. We further found a pH-dependent dynamic trimer-to-monomer conversion and concurrent conformational changes that seem closely linked with a pH-sensing ability through the protonation of certain Asp residues. These results advance our understanding of the functional roles and the related mechanisms of this protease in bacterial infection, which may shed light on the development of HtrA-targeted therapies for H. pylori-associated diseases.

Pharmacophore Oriented MP2 Characterization of Charge Distribution for Anti-SARS-CoV-2 Inhibitor Nirmatrelvir

Quantum mechanical second order Møller–Plesset (MP2) perturbation theory and density functional theory (DFT) Becke, 3-parameter, Lee-Yang-Parr (B3LYP) and Minnesota 2006 local functional (M06L) calculations were performed to optimize structure of nirmatrelvir and compute the Merz-Kollman electrostatic potential (MK ESP), natural population analysis (NPA), Hirshfeld, charge model 5 (CM5), and mulliken partial charges. The mulliken partial charge distribution of nirmatrelvir exhibits a poor correlation with the MK ESP charges in MP2, B3LYP, and M06L calculations respectively. The NPA, Hirshfeld, and CM5 partial charge scheme of nirmatrelvir indicate a reasonable correlation with MK ESP charge assignments in B3LYP and M06L calculations. The above correlations were not improved by the inclusion of implicit solvation model. The MK ESP and CM5 partial charges show a strong correlation between the results of MP2 and two DFT methods. The three optimized structures present a certain degree of differences from the crystal bioactive conformation of nirmatrelvir, suggesting the nirmatrelvir-enzyme complex is formed in the induced-fit model. The Reactivity of warhead electrophilic nitrile is justified by the relatively weaker strength of π bonds in the MP2 calculations. The nirmatrelvir hydrogen bond acceptors consistently show strong delocalization of lone pair electrons in three calculations, whereas hydrogen bond donors are found to have high polarization on the heavy nitrogen atoms in MP2 computations. This work helps to parametrize the force field of nirmatrelvir and improve accuracy of molecular docking and rational inhibitor design.

Ultralong organic room-temperature phosphorescence, multiple stimulus responsiveness and high-level anti-counterfeiting based on multifunctional carbazolyl imidazolopyridine

Ultralong organic room-temperature phosphorescence materials induced by host−guest doping and the underlying mechanism have become the current research focus. Furthermore, high-level anti-counterfeiting and information encryption require multifunctional luminescent materials. Here, CzIP and L-CzIP with push−pull electronic system are successfully constructed by imidazopyridine units and commercial/self-made carbazole. The comparison experiment results indicate that trace isomer doping not only shows slight influence on crystal stacking and molecular space conformation but also significantly changes fluorescence and phosphorescence spectra and phosphorescence lifetime by boosting intersystem spin–orbit coupling. Non-radiative relaxation of CzIP and L-CzIP can be effectively inhibited by polymethyl methacrylate (PMMA) doping but not for crystallize. PMMA doping further confirms intrinsic phosphorescent characteristic of L-CzIP, whose room-temperature phosphorescence lifetime (84.83 ms) is close to that of CzIP (100.14 ms). Concentration-dependent fluorescent and phosphorescent emission and dynamic phosphorescent emission are also observed for CzIP in PMMA film. More importantly, a new aminopyridine/CzIP-doped system gives room-temperature phosphorescence lifetimes of 657.56 ms and afterglow lifetimes of 4 s. Notably, CzIP also shows high-contrast mechanochromism, selective response and differentiation to HCl, CF3COOH, and CH3COOH, and excellent detection capability to picric acid in aqueous medium and solid state. Finally, various anti-counterfeiting and encryption patterns are successfully constructed based on multifunction of CzIP. This work not only provides a multifunctional luminous material but also more importantly provides a theoretical basis and experimental guidance for designing novel ultralong organic room-temperature phosphorescence materials and achieving high-level anti-counterfeiting and encryption.

New semiconductor halocadmate [CdnXm](2n–m) crystal structure, molecular conformation and theoretical investigations

The new chemical compound (C6H5ClF3N2)3[Cd2Cl7] has been crystallized as an organic-inorganic hybrid. This structure is one of the rarely known examples, with exhibiting both [CdCl3]1- and [CdCl4]2- anionic environments. Two layers of mono-protoned fluoro organic cations orientated into preferred intermolecular and intramolecular A-H ….X (A ​= ​N,C and X ​= ​Cl,F) hydrogen bonds separate the one-dimensional inorganic zigzag chain [Cd2Cl7]3-.To better understand the cohesion and interactions within the crystal, an examination has been made using the Hirshfeld surface analysis method, as well as another method which is topological analysis using Atom In Molecules (AIM) theory which was carried out to find the Bond Critical Point (BCP).In addition, this compound has been theoretically studied using DFT calculations, in order to examine its compatibility with the experimental results and which may be a means of verifying the homogeneity and purity of the synthesized compound. An experimental studies with infrared (FT-IR) and Raman spectroscopy are used to conduct and discuss the vibration modes of the existing groups in this new salt, this experimental study is compared to the theory. The energy gap, which is Eg ​= ​3.64 ​eV and the UV–visible absorption spectra were used which enabled us to determine the nature of the material in question and is a semiconductor. The emission of this compound covers a significant portion of the visible light spectrum.

A modified bonded model approach for molecular dynamics simulations of New Delhi Metallo-β-lactamase

Modelling metalloproteins using the classical force fields is challenging. Several methods have been devised to model metalloproteins in force fields. Of these methods, the bonded model, combined with Restrained Electrostatic Potential (RESP) charge fitting, proved its superiority. The latter method was facilitated by the development of the python-based Metal Centre Parameter Builder (MCPB.py) AmberTool. However, the standard bonded model method offered by the MCPB.py tool may not be appropriate for validating and refining the binding modes predicted by docking when crystal structures are lacking. That is because the representation of coordination interactions between any bound ligand and metal ions by covalent bonds can hinder the flexibility of the ligand. Therefore, a new modification to the standard bonded model approach is proposed here. Molecular dynamics (MD) simulations based on the new modified bonded model (MBM) approach avoid the bias caused by coordination bonds and, unlike hybrid QM/MM MD, allow for sufficient sampling of the binding mode given the currently available computational power. The MBM MD approach reproduced the studied crystal structure conformations of New Delhi Metallo-β-lactamase 1 (NDM-1). Furthermore, the MBM approach described the binding interactions of intact β-lactams with NDM-1 reasonably, and predicted a non-productive binding mode for the poor NDM-1 substrate aztreonam whilst predicting productive binding modes for known good substrates. This study presents a useful MD method for metallo-β-lactamases and provides better understanding of β-lactam substrates recognition by NDM-1. The proposed MBM approach might also be useful in the investigation of other metal-containing protein targets.

Drug‐Ribosome Interaction Energies at Site‐E Reveal a Reversed Pattern with Respect to Site‐A, While Showing a Mismatch of Crystal vs. Solution Conformations

AbstractAlthough ribosome is a much hunted target in cancer research, scanty success was achieved with small‐molecule modulators, either because of general toxicity or weak interactions with nucleotides and proteins. On the latter ground, the strength of interactions has been quantitatively evaluated here through MD and QM‐MM simulations in explicit water with the only models of yeast ribosome‐modulator complexes at site‐E for which static structural data were available. It turned out that site‐E is shaped per se by powerful electrostatic forces toward a Mg++ ion, which only allow adaptation of modulators to the site, by mainly sticking to the same Mg++ion. This is in contrast with site‐A, which is known from previous studies to be shaped by a network of H‐bonds with the modulator. From these studies lines of guidance begin to emerge as to tailoring ribosome modulators for better performance as antitumor agents.

Influence of Proline Chirality on Neighbouring Azaproline Residue Stereodynamic Nitrogen Preorganization.

We report herein the first systematic crystal structural investigation of azaproline incorporated in homo- and heterochiral diprolyl peptides. The X-ray crystallography data of peptides 1-5 illustrates that stereodynamic nitrogen in azaproline adopted the stereochemistry of neighbouring proline residue without depending on its position in the peptide sequence. Natural bond orbital analysis of crystal structures indicates OazPro -C'Pro of peptides 4 and 5 participating in n→π* interaction with stabilization energy about 1.21-1.33 kcal/mol. Density functional theory calculations suggested that the endo-proline ring puckering favoured over exo-conformation by 6.72-7.64 kcal/mol. NBO and DFT data reveals that the n→π* interactions and proline ring puckering stabilize azaproline chirality with the neighbouring proline stereochemistry. The CD, solvent titration, variable-temperature and 2D NMR experimental results further supported the crystal structures conformation.