BIBECHANA Vol. 22, No. 3, December 2025, 205-214 ISSN 2091-0762 (Print), 2382-5340 (Online) Journal homepage: http://nepjol.info/index.php/BIBECHANA Publisher:Dept. of Phys., Mahendra Morang A. M. Campus (Tribhuvan University)Biratnagar Hydrogen bonding, spectroscopic (FT-IR and FT-Raman), UV-Vis, and Mulliken charge analysis of Guanosine-5 -Diphosphate: insight from DFT and molecular docking Manoj Kumar Chaudhary1, Tarun Chaudhary2, Bhawani Datt Joshi3 Gulab Singh Verma4, Rajesh Kumar Shukla4, Tirth Raj Paneru5,6,∗ 1Department of Physics, Tribhuvan University, Amrit Campus, Institute of Science and Technology, Kathmandu 44600,Nepal 2Department of Physics, Bageshwari Multiple Campus, Mid-West University, Kohalpur, 21900, Nepal 3Department of Physics, Tribhuvan University, Siddhanath Science Campus, Mahendranagar, 10400, Nepal 4Department of Physics, University of Lucknow, Lucknow-226007, India 5Central Department of General Science, Far Western University, Mahendranagar, 10400, Nepal 6Central Department of Physics, Tribhuvan University, Kathmandu, Nepal ∗Corresponding author. Email: tirthpaneru2015@gmail.com Abstract Hybrid functional B3LYP has been used for quantum chemical calculation of Guanosine-5 -Diphosphate (GDP) from DFT approach. The intra-molecular hydrogen bonding in the title molecule was rendered from Quantum theory of atoms in molecule (QTAIM). The vibrations of different functional groups of the GDP have been examined theoretically from FT-IR and FT-Raman calculation. The electrostatic potential (ESP) surface revealed that the maximum positive potential is attributed to H4, and the highest negative potential corresponds to O31, predicting the sites for the intermolecular hydrogen bonding in the crystal packing of the title compound. The theoretical UV-Vis spectrum is used to calculate the excitation energy as well as excitation state of DGP. The binding affinity of GDP with Ras-related C3 botulinum toxin substrate 1 (RAC1) protein is calculated from molecular docking approach. The inhibition constant of 2P2L is less than that of 2H7V; hence, the title compound is a good inhibitor of 2P2L. Keywords Guanosine-5 -Diphosphate (GDP); AIM; FT-IR; FT-Raman; molecular docking. Article information Manuscript received: January 25, 2025; Revised June 1, 2025; Accepted: July 26, 2025 DOI https://doi.org/10.3126/bibechana.v22i3.74596 This work is licensed under the Creative Commons CC BY-NC License. https://creativecommons. org/licenses/by-nc/4.0/ 205 http://nepjol.info/index.php/BIBECHANA tirthpaneru2015@gmail.com https://doi.org/10.3126/bibechana.v22i3.74596 https://creativecommons.org/licenses/by-nc/4.0/ https://creativecommons.org/licenses/by-nc/4.0/ Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 206 1 Introduction Guanosine is a purine nucleoside with neuro- protective properties, particularly in ischemic stroke, spinal cord injury, and depression. It also reduces neuro-inflammation and oxidative stress [1]. The nucleotides guanosine 5-monophosphate (GMP), guanosine 5-diphosphate (GDP), and guanosine 5-triphosphate (GTP) are the basis of guanine-based purines (GBPs). Ecto-nucleotidases can convert these nucleotides into guanosine [2]. GDP is nucleotides made up of guanine, ribose and two phosphate groups [3]. It has wide applica- tions in biological activity like: cellular signaling and metabolism. GDP is formed due to hydrolysis of guanosine triphosphate (GTP) and there is the evolution of energy which is used in cellular pro- cesses. Moreover, GDP take part in the conversion of succinyl-CoA to succinate and there is the pro- duction of energy [4]. When GDP is changed into GTP due to activation of G-proteins then signals are transmitted into the cell. GDP has crucial role to maintain the cell structure as well as it is in- volved in intracellular transport properties by the formation and deformation of microtubules [5, 6]. The functional groups present in GDP are amine group (-NH2), amide group (-C=O-N), hydroxyl group (-OH) and phosphate group (PO4 2-). These functional groups have crucial role to take part in hydrogen bonding in crystal structure as well as in ligand-protein interaction. This study aims to provide insight into reactive parts of GDP by high- lighting the sites of intra- and intermolecular hy- drogen bonding. The atomic charge on each atom of the title molecule revealed from Mulliken charge analysis. The vibrational spectroscopic techniques (FT-IR and FT-Raman), along with UV-Vis spec- troscopy, conducted through quantum chemical cal- culations, elucidated the vibrations and electronic transitions of significant functional groups in the compound, confirming their role in biological ac- tivity. Moreover, the binding affinity of the title compound with the targeted protein was evaluated through molecular docking to assess its biological performance. 2 Materials and Methods Quantum chemical calculation has been conducted from Gaussian 16 software [7]. The exchange - correlation functional B3LYP was used in this cal- culation [8–10]. The diffused as well as polariza- tion functional basis set 6-311++G(d,p) is taken into consideration to obtain the more accurate re- sult [11]. The density functional theory (DFT) [12]is the wonder full technique which gives the re- sult closer to experimental value and this is imple- mented in Gaussian 16 software. The GaussView 06 software [13] is used to visualize the data which is obtained after calculation. The intra-molecular hydrogen bonding in the investigated molecule has been calculated from quantum theory of atoms in molecule (QTAIM) by using AIMALL software package [14, 15]. The RDG isosurface, scatterplot, and molecular electrostatic potential mapped into the molecular surface produced by the Multiwfn 8.0 and VMD 1.9.1 software packages [16, 17]. The ligand-protein interaction of GDP with predicted target RAC1 is explored from AutoDock and Dis- covery Studio Visualizer 4.5 [18, 19]. The Gauss Sum software has been implemented to explore the excitation state and excitation energy from UV-Vis spectroscopy [20]. 3 Results and Discussion 3.1 Atoms in molecules (AIM) he intra-molecular hydrogen bonding of GDP has been explored by using QTAIM analysis [15]. The judgement of intra-molecular hydrogen bonding in GDP is based on the electron density (ρBCP), Laplacian of electron density (∇2ρBCP), and to- tal electron density (HBCP) at bond critical point (BCP). If the respective value of (ρBCP) and (∇2ρBCP) lies in the range (0.0020.034) a.u and (0.0240.139) a.u, then there is the existence of hy- drogen bonding [21]. The nature and strength of hydrogen bonding is identified in terms of (∇2ρBCP) and HBCP. If their values are greater than zero, then there should be weak hydrogen bond (electro- static). If (∇2ρBCP) > 0 and HBCP < 0; then there should be partially covalent bond. If ((∇2ρBCP)) < 0 and HBCP < 0; there should be strong cova- lent hydrogen bond [22]. The optimized structure of the GDP with atom numbering scheme is de- picted in Figure 1. The molecular graph of GDP highlighting intra-molecular hydrogen bonds is pre- sented in Figure 2. The Table 1 represents the bond length (Å), bond angle (°) and sum of van der Waal radii of interacting atoms (rH + rA). The value of (ρBCP), ∇2(ρBCP), electron kinetic energy density (GBCP), electron potential energy density (VBCP), total electron energy density (HBCP), and interac- tion energy (Eint) at BCP for GDP is presented in Table 2. The distance between interacting atoms is less than the value of rH + rA but (∇2ρBCP) > 0 and HBCP < 0 so all the four intra-molecular hydro- gen bonds are partially covalent in nature. More- over, the interaction energy (Eint) at BCP for O10- H11...O2 is highest (8.311) kcal/mol, and the bond length H11. . . O2 is least (1.813 Å) so, H11. . . O2 is the strongest intra-molecular hydrogen bond out of four. Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 207 Figure 1: 3D structure of GDP obtained from B3LYP/6-311++G(d,p) level of theory Figure 2: Molecular graph of GDP for intra- molecular hydrogen bond. Table 1: Geometrical parameters for intramolecular hydrogen bonds in GDP: bond length (Å), bond angle (°), and sum of van der Waals radii of interacting atoms (rH + rA) in Å. D–H· · ·A D–H (Å) H· · ·A (Å) D–H· · ·A (°) rH + rA (Å) C19–H39· · ·O35 1.078 2.316 134.291 2.72 C15–H41· · ·O7 1.090 2.405 135.688 2.72 O33–H34· · ·O7 0.970 2.016 161.390 2.72 O10–H11· · ·O2 0.987 1.813 148.943 2.72 Table 2: Topological parameters for intramolecular interaction of GDP: electron density at the bond critical point ( ρBCP), Laplacian of (∇2ρBCP), electron kinetic energy density (GBCP), electron potential energy density (VBCP), total electron energy density (HBCP), and interaction energy (Eint) at the BCP. Interaction Bond Length (Å) ρBCP (a.u.) ∇2ρBCP (a.u.) GBCP (a.u.) VBCP (a.u.) HBCP (a.u.) Eint (kcal/mol) H39· · ·O35 2.315 0.0114 0.0427 -0.0019 -0.0069 -0.0088 -2.155 H41· · ·O7 2.405 0.0103 0.0353 -0.0014 -0.0062 -0.0076 -1.937 H34· · ·O7 2.014 0.0170 0.0710 -0.0029 -0.0119 -0.0148 -3.748 H11· · ·O2 1.813 0.0310 0.1093 -0.0005 -0.0265 -0.0270 -8.311 3.2 Non-covalent interaction analysis The non-covalent interaction analysis employed a reduced density gradient (RDG) scatterplot to vi- sualize various interactions in the chemical system using electron density and its derivative. The graph between the sign(λ2)ρ and a dimensionless param- eter RDG insight into the intensity of the interac- tion. The value of RDG can be evaluated by the following equation: [23,24] RDG(r) = 1 2(3π2) 1 3 |∇ρ(r)| ρ(r) 4 3 Different colour codes are used for different types of non-covalent interactions. Weak van der Waals force of attraction is represented by green with the value of sign(λ2)ρ ≈0. Steric repulsion between interacting species is represented by red with the value of sign(λ2)ρ > 0 and strong hy- drogen bonding interaction is represented by blue, with the value of sign(λ2)ρ < 0 [25]. The 2D-NCI RDG graph and its 3D isosurface for the visualisa- tion of non-covalent interaction are shown in Figs, 3 (a) and (b). The RDG isosurface illustrates that the green colour flaky patches appeared between oxygen and hydrogen and nitrogen and hydrogen, which represent the van der Waals interactions be- tween them that are seen in the RDG scatter graph in the region of 0.01 to 0.005 a.u. The red spikes seen in the RDG scatter graph cover a wide range from 0.01 to 0.05 a.u., highlighting the repulsion between the carbon atoms of rings R1, R2, and R3, which is dominant over hydrogen bonding and van der Waals force of attraction. The strong intra- molecular hydrogen bond between H11 and O2 is seen in the isosurface by the blue colour, which is observed in the RDG scatter graph with blue spikes in the range of 0.03 to 0.05 a.u. NCI analysis reveals that the intra-molecular hydrogen bonds H39...O35, H41...O7, and H34...O7 exhibit weak van der Waals forces of attraction, while the intra-molecular hy- drogen bond O10-H11...O2 is a strong hydrogen bond, which was also justified by QTAIM analysis. Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 208 (a) (b) Figure 3: (a) RDG scatter plot of GDP and (b) 3D isosurface showing non-covalent interactions in GDP. 3.3 Spectroscopic studies he number of mode of vibration of the molecular system is explained by the relation 3N-6, where N is the number of atoms in the molecule. The GDP molecule has 43 atoms so there are 123 modes of vibration. The FT-IR and the FT-Raman of the in- vestigated molecule has been examined from same level of theory. The calculated spectrum of FT- IR is presented in Figure 4 and the calculated FT- Raman is presented in Figure 5. The DFT calcula- tion is used to obtain the Raman intensities but it depends on the Raman scattering amplitudes and the Raman scattering cross-section∂σj ∂Ω . The Ra- man scattering amplitude gives Raman intensity for each normal mode of vibration and is given by the relation [26,27]. ∂σj ∂Ω = ( 24π4 45 ) (ν0 − νj) 4 1− exp [ −hcνj kT ] ( h 8π2cνj ) Sj where Sj= scattering activities, νj = predicted wave numbers for jth normal mode, ν0= wavenum- ber of Raman excited state and h, c and k are uni- versal constants. The calculated wavenumber is more than the ex- perimental one due to anharmonicity. So, to reduce the calculated wave number the linear wavenumber linear scaling (WLS) factor is used [28, 29]. The functional groups are the active parts of GDP and can play a vital role in biological action through different intermolecular interactions. These active parts also participate in intra-molecular hydrogen bonding. The signature of functional groups are seen in higher wavenumber in pure form but the signature at lower wave number is determined the fingerprint regions of GDP and they are in mixed mode. In this study, we only discussed the modes of vibrations related to the signature of functional groups that are seen in higher wavenumbers and are important in biological action. The vibration modes of active parts belong to a functional group and are assigned to higher wavenumber regions along with unscaled and scaled wavenumbers, which are presented in Table 3. The vibrational modes associated with the active groups of GDP are dis- cussed in detail in the following sections. 3.3.1 C=O and P=O vibrations The experimental value of N–H stretching vibra- tions is found in the order of (3500-3300) cm-1 [30]. In the GDP compound, asymmetric N26H2 stretch- ing is calculated at 3493 cm1, but its symmetric stretching is calculated at 3396 cm1. The stretch- ing of N23H vibration is computed at 3412 cm¹ with a significant peak in the FT-IR spectra. The in-plane bending of the amine group N26H2 is cal- culated at 1630 cm-1and 1585 cm-1with significant IR absorbance. In the title compound, the signa- tures of CN vibration in rings R1 and R2 are exam- ined at 1556, 1528, and 1487 cm-1with sharp peaks. Experimental OH stretching is generally found in the range (35003200) cm–1 [31]. In the title com- pound, the OH stretching is calculated from DFT at 3633, 3621, 3619, 3517, and 3236 cm¹, corre- sponding to O35H, O8H, O3H, O33H, and O10H, respectively. The amine group and hydroxyl group has significant peaks in IR and Raman spectroscopy and these functional groups interact with residue of amino acid which is justified in the molecular dock- ing section. Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 209 Table 3: The scaled and unscaled wavenumber along with the mode of vibration of different functional groups in GDP. Unscaled Wavenumber (cm−1) Scaled Wavenumber (cm−1) Types of Vibration 3841 3631 ν(OH) stretching vibration of O35H 3827 3621 ν(OH) stretching vibration of O8H 3824 3619 ν(OH) stretching vibration of O3H 3709 3517 ν(OH) stretching vibration of O33H 3681 3493 νa(NH2) asymmetric stretching of N26H2 3591 3412 ν(NH) stretching vibration of N23H 3574 3397 νs(NH2) symmetric stretching of N26H2 3394 3236 ν(OH) stretching vibration of O10H 3263 3118 R2[ν(CH)] stretching vibration of C19H 3165 3029 νa(CH2) asymmetric stretching of C13H2 3110 2979 R3[ν(CH)] stretching vibration of C17H 3104 2974 R3[ν(CH)] stretching vibration of C15H 3098 2969 νs(CH2) symmetric stretching of C13H2 3059 2933 R3[ν(CH)] stretching vibration of C16H 2994 2874 R3[ν(CH)] stretching vibration of C14H 1787 1750 R3[ν(C=O)] stretching vibration of C22=O 1661 1630 δin(NH2) in-plane bending of N26H2 1614 1585 δin(NH2) in-plane bending of N26H2 1583 1556 ν(CN) vibration of CN in Ring (R1 and R2) 1554 1528 ν(CN) vibration of CN in Ring (R1 and R2) 1511 1487 ν(CN) vibration of CN in Ring (R1 and R2) 1505 1481 δin(CH2) in-plane bending of C13H2 1452 1431 R3[δin(CH)] in-plane bending of C17H 1439 1418 R2[δring] ring deformation of R2 1437 1416 γ(CH2) twisting of C15H2 1415 1395 R3[δin(CH)] in-plane bending of CH 1407 1387 δin(OH) in-plane bending of O33H 1384 1365 ω(CH2) wagging of C13H2 1291 1275 ν(P=O) stretching vibration of P6O7 1247 1233 ν(P=O) stretching vibration of P1O2 1166 1154 δin(OH) in-plane bending of O10H 953 946 δin(OH) in-plane bending of O8H and P6O10 stretching 871 866 δin(P6-O10-O8) in-plane bending 865 860 R2[δin(CH)] in-plane bending of C19H 863 859 R2[δin(CH)] in-plane bending of C19H 3.3.2 CH vibration The experimental C-H stretching vibration is ob- tained in the range (3000-3100) cm–1 [31]. In the title compound, the stretching of C19H, C17H, C15H, C16H, and C14H in the rings R2 and R3 are calculated at 3118, 2979, 2974, 2933, and 2874 cm1 , respectively being very sharp peak for C14H with higher IR absorbance. The in-plane bending of CH vibration is computed at 1431 and 1395 cm–1 in the ring R2. For ring R3, the in-plane bending of CH found to be at 860 and 859 cm–1. The asym- metric stretching vibration of C13H2 is calculated at 3029 cm–1 but symmetric stretching of C13H2 is calculated at 2969 cm–1. 3.4 Electrostatic potential surface (ESP) analysis The visual representation of the charge distribution on the molecular surface can be performed by the electrostatic potential surface analysis. This tool aids in the prediction of sites for the intra- and intermolecular interactions by identifying the loca- tions of electrophiles and nucleophiles [32]. The region of positive potential shown by the blue colour region corresponds to the position of the electrophile, which should be favourable for nucle- ophilic attack, and the negative potential region represented by the red colour is the site of elec- trophilic attack [33]. The molecular surface of the title compound mapped with the electrostatic po- tential on its van der Waals surface is shown in Fig- ure 6. The blue and orange dots of the molecular electrostatic surface represent the points of mini- mum and maximum potential. The global mini- mum potential of -95.53 kcal/mol attributed to the O31 atom in the C=O group of ring R1 of GDP is the nucleophile. The maximum positive poten- tial 113.71 kcal/mol corresponds to the H4 atom attached to the OH group in phosphate, making it the best site for nucleophilic attack. From this re- sult, we conclude that H4 and O31 are ideal sites for intermolecular hydrogen bonding, and the crystal packing of the title compound may be contributed Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 210 by the O3-H4. . . O31 hydrogen bonding. Figure 4: Calculated FT-IR spectra of Guanosine-5 - Diphosphate in the range (100-3800) cm-1 calcu- lated from B3LYP/6-311++G(d,p) level of theory. Figure 5: Calculated FT-Raman spectra of Guanosine-5 - Diphosphate in the range (100-3800) cm-1 calculated from B3LYP/6-311++G(d,p) level of theory. Figure 6: Electrostatic potential mapped into molecular vdW surface of Guanosine-5 - Diphos- phate. 3.5 Mulliken Charge Analysis The electronic property of atom in molecular sys- tem has been identified from Mulliken charge anal- ysis. This is calculated on the basis of distribution of electrons in the atoms as well as the formation of polarity in the molecular system [34]. This analysis helps to identify the electrophilic and nucleophilic atoms in the compound that can take part in chem- ical reaction with surrounding species in hydrogen bonding in crystal packing and ligand-protein in- teraction [35]. The distribution of charge on the heavy atoms of GDP is presented in Figure 7. From this analysis it is clear that the concentration of negative charge is mainly on O2, O5, O7, C17, N29, C30, and O31. Out of these atoms, the highest value of negative charge is seen on C17. This is due to shearing of charge among C16, N18 and O32 atoms. Similarly, the concentration of positive charge is more across P6, N18 and C19. But the highest value of posi- tive charge is calculated across N18. These atoms have prominent role to take part in ligand-protein reaction. Figure 7: Mulliken charge with hydrogen summed into heavy atoms on GDP from B3LYP/6- 311++G(d,p). 3.6 UV-Visible Spectra Analysis The essential and useful technique to study the bio- logically active compounds is UV-Vis spectroscopy. The spectrum helps in the characterization of sub- stances and gives an idea regarding their electronic structure. The spectrum is used to check biologi- cal activity of compounds when changes are made to functional groups that insure the change in (in- crease or decrease) drug activity of molecules when derivatives are synthesized [35]. Besides these ac- tivities, it also helps with compound stability, activ- ity quantification, and interaction monitoring which are critical for assessing and maximizing biological activity. UV-Vis spectroscopy can monitor enzyme activity in real time; it helps to study the photo stability of biologically active compounds [36]. It helps to quantify the changes in absorbance upon binding, indicating interaction mechanisms, bind- ing constants, and binding sites. Moreover, it helps to examine the electronic structure, aromatic rings, and functional group present in the compound and the band gap energy of compound to check the na- ture of compound whether it is semiconductor or other type of materials [37, 38]. Besides that it has wide applications to measure the concentration and interaction of proteins and nucleic acids as well as it is used to study the enzyme activity. The UV- Vis spectra of GDP is calculated in gaseous state by employing the TD-B3LYP/6-311++G(d,p) the- Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 211 ory. The UV-Vis spectra is plotted which is pre- sented in Figure 8. The absorption peak is ob- tained at 271 nm which infer that there is delocal- ization of -electron due to guanine base which con- tains conjugated double bonds and aromatic ring in GDP. In guanine base structure there is also ab- sorption of lone pair electrons η due to presence of nitrogen (N) and Oxygen (O) atoms in GDP. Thus, there is transition of π → π∗ and η → π∗ electrons and in such case the order absorption wavelength is (250-280) nm. The percentage con- tribution of molecular orbitals along with oscilla- tor strength, energy gap and maximum absorption wavelength is presented in Table 4. The HOMO or- bital is 114 and LUMO orbital is 115. The first ex- cited state (HOMO→LUMO (100%)) is calculated as 4.06 eV corresponding to λmax=305.17 nm hav- ing minimum oscillator strength=0.0001. This is due to the transition of π → π∗ electrons. Similarly, the second excited state is calculated at 275.06 nm (HOMO→L+2 (94%)) with respective excitation energy and oscillator strength 4.51eV and 0.0237. This is due to transition of π → π∗ and eta → π∗ electrons. Figure 8: UV-Vis absorbance of GDP in gas phase calculated with TD-B3LYP/6311++G(d,p) 3.7 Molecular Docking Molecular docking is crucial technique to explore the ligand-protein interaction. The binding sites as well as binding energy, inhibition constant and lig- and efficiency are the important parameters which explain the docking analysis of ligand with pre- dicted target (protein) [39, 40]. The target protein Ras-related C3 botulinum toxin substrate 1(RAC1) has been predicted from free online SwissTarget- Prediction and two protein 2P2L and 2H7V has been downloaded from protein data bank [41, 42]. The protein has been cleaned by removing the wa- ter molecule and co-crystalized ligand. The lig- and GDP and proteins 2P2L and 2H7V are pre- pared for docking by converting them into PDB file. After that the docking has been performed from AutoDock Vina [18]. The binding sites have been examined from Discovery Studio Visualizer 4.5 [19]. Out of the many docked conformers the best docked conformers have been presented in Figure 9. The conventional hydrogen bonds, residue of amino acid, ligand efficiency, inhibition constant and bind- ing energy of ligand-protein interaction is presented in Table 5. From molecular docking analysis it is confirmed that the binding sites in GDP are O2, O3, O5, O7, O10, O31, N29, and H34. During the molecular docking analysis of GDP with Cell divi- sion control protein 42 homolog (Cdc42) with the PDB codes 1ANO, 1A4R and 1DOA; the binding sites was found to be almost same [43]. The bind- ing energy for 2P2L (-8.0 kcal/mol) is more than the binding energy of 2H7V (-7.4 kcal/mol). More- over, the inhibition constant of former (1.35 µM) is less (3.73 µM) than the later one. The num- ber of conventional hydrogen bond in 2P2L is 10 and the number of hydrogen bond in 2H7V is 7. Besides that, the bond length of conventional hy- drogen bond in 2P2L is less than that of 2H7V. So, former is better inhibited by title compound. Table 4: The conventional hydrogen bonds, residue of amino acid, ligand efficiency, inhibition constant, and binding energy of GDP. Ligand Protein PDB code Bond length (Å) Binding Atoms Amino Acid Binding Energy (kcal/mol) Inhibition Constant (µM) Ligand Efficiency GDP RAC1 2H7V 2.54 O31 LYS116 –7.4 3.73 0.26 2.68 O2 LYS16 2.28 O7 LYS16 2.05 O7 GLY15 2.33 O7 VAL14 2.00 O7 LYS16 2.01 O3 THR17 GDP RAC1 2P2L 2.16 N29 SER41 –8.0 1.35 0.29 2.75 H34 SER41 2.80 H34 SER41 2.22 O2 LYS16 2.45 O2 VAL14 2.32 O2 LYS16 1.96 O2 GLY15 1.75 O3 THR17 2.38 O5 LYS16 2.31 O10 ALA13 Manoj Kumar Chaudhary et al./ BIBECHANA 22 (2025) 205-214 212 Figure 9: 2-D ligand-protein interaction of GDP with 2H7V and 2P2L PDB code of protein RAC1. 4 Conclusion The intra-molecular hydrogen bonding of GDP has been scrutinized from QTAIM analysis and RDG scatter plot and its 3D-isosurface. Four intra- molecular hydrogen bonds have been identified and all are partial covalent in nature. Out of them the intra-molecular hydrogen bond H11...O2 is the strongest as it has the smallest bond distance 1.8134 Å and the highest interaction energy (8.3114 kcal/mol). The maximum positive electrostatic po- tential 113.71 kcal/mol, was observed in H4, and the maximum negative potential associated with O31 of GDP, as predicted by ESP analysis. This analysis confirms that these sites are favourable for intermolecular hydrogen bonding in the solid-state structure of the title compound as well as in ligand- protein interaction analysis. The GDP molecule has 43 atoms and 123 modes of vibrations. All the vi- brations are IR and Raman active. The fingerprint and functional group present in the molecule have been explored. There are significant peaks for car- bonyl group (C=O), amine group (-NH2), and hy- droxyl group (-OH) and phosphate group (PO4 2-). It is confirmed that these functional groups have prominent role for intermolecular hydrogen bond- ing with residue of amino acid which is verified by the molecular docking as well as Mulliken charge and ESP analysis. UV-Vis spectra analysed that there are single significant absorbance peak at 272 nm wavelength which is correspond to the second excited state (HOMO→L+2 (94%)) with respective excitation energy and oscillator strength 4.51eV and 0.0237. This arises due to transition of π → π∗ and η → π∗ electrons. Mulliken charge analysis con- firmed that the atoms O2, O5, O7, C17, N29, C30, and O31 have negative charge and the atoms P6, N18 and C19 have positive charge. These charges take part in ligand-protein interaction which is ex- plored from molecular docking. The protein 2P2L has more binding affinity than 2H7V protein of RAC1. Author contribution M.K. Chaudhary: Conceptualization of research ac- tivity, data analysis and manuscript writing; T. 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