Sulfur takes center stage in a thioxoacetamide crystal and docking study

Chemists have synthesized and fully characterized a new thioxoacetamide compound, N-benzyl-2-((4-chlorobenzyl)amino)-2-thioxoacetamide, known as 1b, and shown that its thiocarbonyl sulfur is not a passive bystander but an active architect of the crystal. The team, led by researchers at The National Institute of Engineering in Mysore and the University of Mysore, combined single-crystal X-ray diffraction, Hirshfeld surface analysis, density functional theory calculations, molecular docking, molecular dynamics simulations, and ADMET predictions to build a complete structure–interaction profile of the molecule. The work appears in Frontiers in Chemical Biology.

The compound was made in two steps from benzylamine, chosen for its high nucleophilicity and commercial availability. Benzylamine was first converted to N-benzyl-2-chloroacetamide with chloroacetyl chloride and triethylamine in dichloromethane at 0 °C, then transformed into the key intermediate methyl 2-(benzylamino)-2-oxoethanedithioate in 87% yield using elemental sulfur in dimethyl formamide. A final condensation of this intermediate with 4-chlorobenzylamine in ethanol at room temperature for 10–15 minutes delivered the target compound in 93% yield as a pale yellow solid. Nuclear magnetic resonance spectroscopy, infrared spectroscopy, and high-resolution mass spectrometry all confirmed the structure, with the protonated molecular ion observed at m/z 319.0681 against a calculated value of 319.0672.

Crystals suitable for diffraction were grown by slow evaporation from acetonitrile over 24–36 hours at room temperature. X-ray data collected on a Rigaku R-AXIS RAPID-S diffractometer with graphite-monochromated Mo-Kα radiation showed that 1b crystallizes in the triclinic system with two molecules per unit cell and a cell volume of 764.53 (19) ų. Refinement on F² converged with an R1 value of 0.0734 for data with I greater than 2σ(I), supporting the reliability of the structural model. The molecule adopts a distinctly non-planar conformation: the central thioxoacetamide fragment forms dihedral angles of 112.72° with the chlorobenzyl ring and 114.64° with the N-benzyl ring, a flexibility the authors suggest could matter both for solid-state assembly and for adapting to protein binding surfaces.

The most striking result concerns how the molecules pack together. Neighboring molecules are linked by N1–H1···O1 hydrogen bonds that generate R2²(10) ring motifs, while a complementary set of N2–H2···S1 interactions forms additional R2²(10) motifs with a different symmetry code. Acting cooperatively, these N–H···O and N–H···S contacts propagate a one-dimensional supramolecular chain along the a-axis. The authors argue that these recurring motifs demonstrate the thioxo group is an active participant in defining the crystal architecture rather than an incidental substituent, providing experimental evidence that C=S-centered hydrogen bonding can enforce specific packing arrangements.

Hirshfeld surface analysis, performed with CrystalExplorer 17.5, put numbers on this picture. The dominant contributions to the crystal packing come from H···H contacts at 42.2%, reflecting extensive van der Waals interactions among the many hydrogen atoms, followed by C···H/H···C contacts at 20.9% and H···Cl/Cl···H contacts at 11.6%. Crucially, the S···H/H···S contribution of 7.8% directly reflects the crystallographically observed N–H···S hydrogen bonding, while O···H/H···O contacts contribute 6.6%, with the shortest combined contact distance for the oxygen interactions at approximately 2.0 Å, consistent with strong directional hydrogen bonding. The globularity value of 0.715 and asphericity of 0.445 quantify a moderately anisotropic, non-spherical molecular shape.

Energy framework analysis added an energetic dimension to the packing story. The calculated pairwise interaction energies include an electrostatic component of −118.91 kJ mol⁻¹, polarization of 19.24 kJ mol⁻¹, dispersion of −222.10 kJ mol⁻¹, and repulsion of 122.61 kJ mol⁻¹, summing to a total interaction energy of −232.81 kJ mol⁻¹. Although the directional hydrogen bonds organize the supramolecular architecture, the dispersion component is nearly twice the electrostatic one, indicating that overall crystal stability is governed primarily by cumulative van der Waals contacts distributed throughout the lattice, a pattern typical of organic molecular crystals.

Density functional theory calculations at the B3LYP/6–311+G(d,p) level closely reproduced the experimental geometry, with an overall RMSD of 0.646 Å and correlation coefficients of 0.9974 for bond lengths, 0.9819 for bond angles, and 0.9780 for torsion angles. The HOMO and LUMO energies were calculated as −6.276 eV and −2.133 eV, giving an energy gap of 4.143 eV that the authors interpret as a balance between chemical stability and reactivity. Global reactivity descriptors include a hardness of 2.072 eV, softness of 0.483 eV⁻¹, electronegativity of 4.204 eV, and electrophilicity index of 4.267 eV, while the low dipole moment of 1.003 D suggests weak molecular polarity. Molecular electrostatic potential mapping showed negative potential localized around the carbonyl oxygen and, to a lesser extent, the thioxo sulfur, matching the hydrogen-bond acceptor roles seen in the crystal.

To probe biological relevance, the team docked 1b into the nucleotide-binding pocket of human thymidine kinase 1, using the crystal structure with PDB ID 1XBT at 2.40 Å resolution. The docking produced a binding energy of −8.00 kcal/mol, with the strongest polar contact a hydrogen bond between the thioxo sulfur and GLN100 at 2.56 Å. Additional stabilizing contacts include a π-donor hydrogen bond between PHE29 and the chlorobenzene ring centroid at 2.65 Å, a π–σ interaction with LEU124 at 3.86 Å, a π–sulfur interaction with MET28 at 4.62 Å, and π–π stacked and T-shaped interactions with TYR181 and PHE133 at centroid distances of 4.33 Å and 5.08 Å respectively. A comparison with the co-crystallized ligand dTTP showed that 1b occupies a similar position in the pocket, and a redocking check yielded an all-atom RMSD of 4.8917 Å between the native and redocked conformations.

Molecular dynamics simulations of the 1b–1XBT complex ran for 100 ns using the Desmond package with the OPLS force field. The protein Cα RMSD stabilized around 1.6–1.8 Å up to roughly 60 ns before rising gradually to about 2.3–2.4 Å, while the ligand RMSD fluctuated between 1.2 and 2.5 Å without abrupt spikes or continuous drift, suggesting the ligand remains bound throughout the simulation. Most residues showed low RMSF values of about 0.4–1.0 Å, with larger fluctuations confined to flexible loop and terminal regions around residues 38–45 and 140–150 that are generally not involved in ligand binding.

The predicted ADMET profile, generated with the pkCSM web server and supplemented by SwissADME projections, indicates high human intestinal absorption of 89.82%, Caco-2 permeability of 1.783 log Papp, a topological polar surface area of 73.22 Ų, a consensus LogP of 3.41, and a molecular weight of 318.82 g mol⁻¹, satisfying Lipinski’s rule of five and several other drug-likeness criteria without violations. Predicted toxicity screens found no AMES mutagenicity, hepatotoxicity, skin sensitization, or hERG I inhibition, though predicted hERG II inhibition suggests cardiotoxicity should be evaluated in future work. The authors are careful to stress the limits of their study: the docking and ADMET results are computational predictions that do not prove TK1 inhibition, biological activity, or safety, and experimental biochemical and cellular studies will be needed to validate the anticipated interactions. Within those bounds, the work delivers a detailed, multi-technique portrait of a thioxoacetamide in which the C=S group emerges as a structure-directing element in the crystal and a plausible interaction point in a protein binding site.

Subject of Research: Crystal structure, electronic properties, and computational docking analysis of a newly synthesized thioxoacetamide derivative

Article Title: Structural and computational insights into a thioxoacetamide: role of C=S interactions in crystal packing and molecular recognition

Article References: Siddegowda, N., Santhosh, C., Ramarathinam, R., Doddakempegowdanakoppalu Chandra, V. K., Iyengar, D. S., Rangappa, M. G., Nagarajappa, L. T., Vishnu Kumar, I. K., & Sadashiva, M. P. (2026). Structural and computational insights into a thioxoacetamide: role of C=S interactions in crystal packing and molecular recognition. Frontiers in Chemical Biology, 5, Article 1972711. https://doi.org/10.3389/fchbi.2026.1972711

Image Credits: AI Generated

DOI: 10.3389/fchbi.2026.1972711

Keywords: thioxoacetamide, crystal structure, X-ray diffraction, Hirshfeld surface analysis, hydrogen bonding, DFT calculations, molecular docking, thymidine kinase 1, molecular dynamics, ADMET, supramolecular chemistry, Structural