Molecular Structure, Vibrational Spectra and Docking Studies of Abacavir by Density Functional Theory

. In this study, optimized geometry, spectroscopic (FT-IR, FT-Raman, UV) analysis, and electronic structure analysis of Abacavir were investigated by utilizing DFT/B3LYP with 6-31G(d,p) as a basis set. Complete vibrational assignments and correlation of the fundamental modes for the title compound were carried out. The calculated molecular geometry has been compared with available X-ray data of Abacavir. The calculated HOMO and LUMO energies show that charge transfer occurs within the molecule. The molecular stability and bond strength have been investigated by applying the Natural Bond Orbital (NBO) analysis. The computational molecular docking studies of title compound have been performed.


Introduction
The managing of human immunodeficiency virus (HIV) includes the use of multiple antiretroviral, since single drug therapy becomes ineffective due to development of HIV resistant strains. According to treatment guidelines antiretroviral regimen should contain at least two nucleoside analogue reverse transcriptase inhibitors (NRTIs) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) in a fixed dose combination. This is very efficient in the treatment of HIV. Abacavir (ABC) is a nucleoside reverse transcriptase inhibitor (NRTI) with activity against Human Immunodeficiency Virus Type 1 (HIV-1). ABC is phosphorylated to active metabolites that compete for incorporation into viral DNA. Intracellularly, ABC is converted by cellular enzymes to the active metabolite carbovir triphosphate, an analogue of deoxyguanosine-50-triphosphate (dGTP). Chemically ABC is [(1S,4R)-4-[2-amino-6-(cyclopropylamino)-9H-purin-9-yl] cyclopent-2-en-1-yl] methanol [1,2]. The introduction of highly active antiretroviral therapy (HAART) has profoundly altered both the course and prognosis of HIV infection. After 1996, the availability of protease inhibitors (PI) transformed HIV-infection from a progressive and almost uniformly fatal condition to a treatable chronic infection. Choosing an initial antiretroviral regimen is one of the most important decisions faced by clinicians managing HIV disease. Several antiretroviral combinations have proven sufficiently potent to achieve viral suppression in most treated patients. However, maintaining efficacy depends on other factors, such as the durability of antiviral suppression, tolerability, risk of long-term toxicity, and patient convenience [3].
The present work mainly deals with detailed structural conformation, experimental FT-IR and FT-Raman spectra, vibrational assignments using total energy distribution (TED) and Molecular docking activity as well as DFT/B3LYP calculations for ABC. Vibrational spectra of Abacavir have been analyzed on the basis of calculated total energy distribution (TED). Theoretically computed vibrational wavenumbers were compared with experimental values. The natural bond orbital (NBO) analysis can be employed to identify and substantiate the possible intra and intermolecular interactions between the units that would form the H-bonded network. The UV-Vis spectroscopic studies along with HOMO-LUMO analysis have been used to explain the charge transfer within the molecule.

Experimental details
The compound Abacavir was purchased from Aldrich chemicals, USA and used as such to record the FT-IR and FT-Raman, UV spectra. The FT-IR spectrum of Abacavir compound was recorded in the range of 4000-400 cm -1 on a BRUKER Optik GmbH FT-IR spectrometer using KBr pellet technique. The spectrum was recorded in the room temperature, with scanning speed of 10 cm -1 , and spectral resolution: 4 cm -1 . FT-Raman spectrum of the title compound was recorded using 1064 nm line of Nd:YAG laser as excitation wavelength in the region 3500-50 cm -1 on a BRUKER RFS 27: FT-Raman Spectrometer equipped with FT-Raman molecule accessory. The spectral resolution was set to 2 cm -1 in back scattering mode. The laser output was kept at 100mW for the solid sample. The ultraviolet absorption spectra of ABC were examined in the range 200-800 nm using Cary 500 UV-VIS-NIR spectrometer. The UV pattern is taken from a 10 to 5 M solution of ABC, dissolved in ethanol solvent. The theoretically predicted IR and Raman spectra at B3LYP/6-31G(d,p) level calculation along with experimental FT-IR and FT-Raman spectra are shown in Fig. 2 and 3.

Computational details
The density functional theory DFT/B3LYP with the 6-31G(d,p) as basis set was adopted to calculate the properties of Abacavir in the present work. All the calculations were performed using Gaussian 03W program package [4] with the default convergence criteria without any constraint on the geometry [5]. The assignments of the calculated wavenumbers are aided by the animation option of Gauss View 5.0 graphical interface for Gaussian programs, which gives a visual presentation of the shape of the vibrational modes along with available related molecules [6]. Furthermore, theoretical vibrational spectra of the title compound were interpreted by means of TED using the VEDA 4 program [7]. The optimized structural parameters were used in the vibrational frequency calculations at DFT levels to characterize all stationary points as minima. As the hybrid B3LYP functional tends to overestimate the fundamental normal modes of vibration, the computed frequencies were scaled with appropriate values to bring harmonization between the theoretical and experimental wavenumbers [8]. Vibrational frequencies were computed at DFT level which had reliable one-to-one correspondence with experimental IR and Raman frequencies [9]. The Natural Bond Orbital (NBO) calculations were performed using NBO 3.1 program [10] as implemented in the Gaussian 03W [4] package at the DFT/B3LYP level; in order to understand various second order interactions between filled orbital of one subsystem and vacant orbital of another subsystem which is a measure of the intermolecular delocalization or hyper conjugation.

Prediction of Raman intensities
The Raman activities (S Ra ) calculated with Gaussian 03W program [4] converted to relative Raman intensities (I Ra ) using the following relationship derived from the intensity theory of Raman scattering [11] ) where, ν 0 is the laser exciting wavenumber in cm -1 (in this work, we have used the excitation wavenumber ν 0 = 9398.5 cm -1 , which corresponds to the wavelength of 1064 nm of an Nd-YAG laser), ν i the vibrational wavenumber of the i th normal mode (cm -1 ) while S i is the Raman scattering activity of the normal mode ν i [12].

Docking Studies
The molecular structure of protein (PDB ID: 3VRI) was taken from RCSB Protein Data Bank, http://www.rcsb.org/pdb [13]. Initial structures of Abacavir were generated by ChemBioOffice 2008. The geometries of Abacavir legand were subsequently optimized at DFT/B3LYP/ 6-31G (d,p) by Gaussian 03 [4]. The molecular modeling docking calculations of Abacavir legand with 3VRI protein were carried out by means of the Autodock tools (ADT) v1.5.4 [14] and Autodock 4.2.3 program from the Scripps Research Institute. In docking study, the search was extended over the whole receptor Abacavir used as blind docking. The grid maps were generated with 0.375 Å spaces using a grid box of 70-70-70 Å. The search was carried out with the

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Lamarckian Genetic Algorithm because it has been pointed out to be most efficient, reliable and successful methods in Autodock [15]. The docking parameters used were as follows: GA population size = 150; maximum number of energy evaluation = 25,00,000 and others used were default parameters. The docking conformation with the lowest binding free energy was used for further analysis by Molegro Molecular Viewer software from http:// www.clcbio.com/products/molegro/ [16].

Structural analysis
The optimized geometric parameters such as bond lengths, bond angles and dihedral angles of the title molecule were given in Table 1 using DFT calculation with 6-31G(d,p) as a basis set. The atom numbering scheme of the title compound adopted in this study is given in Fig. 1. To the best of our knowledge, experimental data on the geometric structure of the title molecule are not available till date in the literature. Our molecule Abacavir is compared with XRD data of closely related molecules Abacavir methanol 2.5-solvate [17]. The C-C bond length of the purine ring found at C11-C16=1.396 Å and C15-C16=1.414 Å calculated from DFT method which is agree well with XRD value at 1.383 Å and 1.412 Å respectively. The C-N bond length of the purine ring varies from 1.338 Å to 1.335 Å by DFT method 1.334 Å to 1.370 Å by XRD respectively. Similarly, in the cyclopentane ring C-C bond length varies in the range 1.334 Å to 1.556 Å by DFT and 1.329 Å to 1.550 Å by XRD respectively also C-H bond length of this ring found from 1.086 Å to 1.100 Å by DFT and 0.990 Å to 1.000 Å by XRD respectively this is correlate well with calculated values as well as literature data [18].
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Vibrational assignments
The detailed vibrational assignments (characterized by PED) of fundamental modes of Abacavir along with the calculated IR and Raman intensities are reported in Table 2. Theoretical and corresponding scaled (SQM) frequencies were calculated for title compound at B3LYP/6-31G(d,p) level have been collected along with IR intensities and Raman scattering activities. Theoretically calculated wavenumbers are usually larger than the experimental values, due to the exclusion of scaled frequencies, incomplete incorporation of electron correlation and the use of finite basis set. The over estimations are mostly systematic and can be corrected by following an empirical scaling procedure over the obtained theoretical frequencies fit to the corresponding experimental frequencies. In this study, selective scaling procedures were applied. For the B3LYP/6-31G(d,p) level, the scale factors of 0.9608 reported in reference [20] respectively.

Purine Ring vibrations
The C-C stretching vibrations of the purine ring observed in the Spectral region over 1650-1000 cm -1 [21]. In our present investigation C-C stretching vibrations observed at 1606 cm -1 in FT-IR and 1607 cm -1 in FT-Raman spectrum and calculated at 1595 cm -1 and 1569 cm -1 by DFT method. The hetero aromatic structure shows the presence of C-H stretching vibrations in the region 3100-3000 cm -1 , which is the characteristic region for the ready identification of CH stretching vibrations [22]. For title compound this vibration observed at 3196 cm -1 in FT-IR spectrum and 3131 cm -1 by DFT method this is a pure mode, the contribution of PED for this mode is 100%. Silverstein et al. [23]

Amino group vibration
According to Socrates [23] the stretching of amino group appeared around 3500-3000 cm -1 in absorption spectra. For title molecule the wavenumbers 3465 cm -1 and 3592 cm -1 calculated by DFT calculations gives the N-H symmetric and antisymmetric stretching vibrations, no bands observed in FT-IR and FT-Raman spectrums for this vibration. The wavenumber at 1569 cm -1 predicted by DFT method has been identified as NH 2 scissoring vibration.

Methylene and OH group vibrations
The asymmetrical stretching (asymCH 2 ) and symmetrical stretching (symCH 2 ) bands of the methylene group occur near 2926 and 2853 cm −1 , respectively [24]. In our present work the predicted wave numbers at 2967 cm -1 and 2872 cm -1 are identified as asymmetrical and symmetrical stretching vibrations respectively. The FT-Raman band 1455, 1369 cm -1 are observed scissoring and wagging modes of the CH 2 group. These modes are computed at 1459 and 1364 cm -1 by DFT method. The O-H stretching vibration is theoretically predicted at 3676 cm −1 (mode no. 1) by B3LYP method. The computed wavenumbers at 1343 cm -1 and 295 cm -1 gives the COH inplane and CCOH out-off plane bending vibrations respectively.

Cyclopropyl ring vibrations
The CH 2 symmetrical and asymmetrical stretching vibrations observed at 3032-3005 cm -1 and 3119-3100 cm -1 by DFT method. The FT-IR band at 1407 cm -1 and computed wavenumbers at 1419, 1415 cm -1 by DFT method are identified as scissoring CH 2 vibrations of the Cyclopropyl ring. The C-C stretching vibrations of this ring has been identified at 1192 cm -1 in FT-IR and 1199, 815, 805 cm -1 by DFT calculation. The C-C-H inplane bending vibrations identified at 1152, 1150, 742 cm -1 by DFT method.

NBO analysis
In the NBO analysis, the electron wave functions are interpreted in terms of a set of occupied Lewis type (bond or lone pair) and a set of unoccupied non-Lewis (anti-bond or Rydberg) localized NBO orbitals. The delocalization of electron density (ED) between these orbitals corresponds to a stabilizing donor acceptor interaction. A useful aspect of the NBO method is that it gives information about interactions in both filled and virtual orbital spaces, which could enhance the analysis of intra-and intermolecular interactions.
The second-order Fock matrix was carried out to evaluate the donor acceptor interactions in the NBO basis. The interactions result in a loss of occupancy from the localized NBO of the idealized Lewis structure into an empty nonLewis orbital. For each donor (i) and acceptor (j), the stabilization energy E (2) associated with the delocalization i / j is estimated as , ( 2 where q i is the donor orbital occupancy, ε i and ε j are diagonal elements and F(i, j) is the offdiagonal NBO Fock matrix element.

UV-Visible spectral studies
The highest occupied molecular orbitals (HOMOs) and the lowest-lying unoccupied molecular orbitals (LUMOs) are named as frontier molecular orbitals (FMOs). The FMOs play an important role in the optical and electric properties, as well as in quantum chemistry and UV-VIS spectra [26]. Absorption maxima (λmax) (nm) for lower-lying singlet states of the molecule of the molecule have been calculated by TD-DFT/B3LYP method. The computed properties such as 20 ILCPA Volume 72 absorption wavelength (λ), excitation energies (E), frontier molecular orbital energies, and oscillator strengths (f) are listed in Table 4. Fig. 4 shows the observed UV-Vis spectra of title compound in ethanol solvent. For TD-DFT calculations, the theoretical absorption band was predicted at 240.18 nm with oscillator strength being 0.0906 in ethanol solvent and at 239.68 nm with oscillating strength 0.0765 in gas phase can easily be seen that this corresponds to the experimental absorption at 240 nm. Fig. 5 (a, b) shows the distributions and energy levels of the HOMOs and LUMOs orbitals computed at the B3LYP/6-31G(d,p) level for Abacavir. The conjugated molecules are described by a small HOMO-LUMO separation which is the consequence of a significant amount of intra molecular charge transfer from electron donor groups to the capable electron acceptor groups through π-conjugated path [27]. The calculated energy values of HOMO and LUMO are -5.1506 eV and -0.1474eV and the frontier orbital energy gap value is -5.0032 eV for Abacavir.    Figure 5 (a, b). The atomic orbital compositions of the frontier molecular orbital for Abacavir.

Molecular electro static potential analysis
MEP is pertained to the electronic density and is a very useful descriptor in understanding sites for nucleophilic reaction and electrophilic attack as well as hydrogen bonding interactions [28]. It also provides visual understanding of relative polarity of the molecule. The MEP surface has been plotted for the molecules Abacavir in Fig. 6. From the Fig.6 it can be seen that region of negative charge is pictured out in red colour and it is found around the electronegative N in the purine ring and O20 atom in the hydroxy groups in the molecule Abacavir. The red region gives the electrophilic attack. The blue colour region represents strong positive region (all hydrogen atoms) and is prone to nucleophilic attack.

Molecular docking studies
Molecular docking studies were performed to investigate the binding affinities of the title compound ABC and the human protein Abacavir, PDB ID: 3VRI [29]. The ligand-protein complex stability was successfully made by some features such as hydrogen bond interactions, vander Waals forces, π-π stacking, hydrophilic and hydrophobic interactions. High resolution crystal structure of Abacavir compound hydrogen and water molecules are reductase was downloaded from the protein data bank website (PDB ID: 3VRI) and all molecular docking calculations were performed on Auto Dock-Vina software [30]. The protein was prepared for docking by removing the co-crystallized ligands, waters and co-factors and the Auto Dock Tools graphical user interface was used to calculate Kollman charges and polar hydrogens. Molecular docking studies were performed to investigate the higher binding affinities and total intermol energy of the newly compound Abacavir is -7.74 kcal/ mol and -5.19 kcal/ mol lower binding affinities and total intermol energy of the title molecules is -9.16 kcal/ mol and -6.62 kcal/ mol respectively. The hydrophobic interactions between ABC and TYR 74, ASP114, Gly-217, SCR116 is found at 2.81 Å, 3.04 Å, and 3.12 Å. On the other hand, a π-π stacking exists between phenyls of ABC and TRP147 is 3.47 Å. The title molecule is given in Fig. 7 and the values are tabulated in Table 5.

Conclusions
This paper presents spectroscopic and molecular modeling studies on the interaction of Abacavir using, FT-IR, FT-Raman, UV-Vis spectrum. The equilibrium geometries, HOMO-LUMO analysis and vibrational frequencies of Abacavir have been calculated at B3LYP levels using 6-31G(d,p) basis set. The title molecular HOMO-LUMO energy gap is -5.0032 eV. The agreement between the optimized and experimental crystal structure is quite good, which shows that the geometry optimization almost exactly reproduces the experimental conformation. The electronic properties are also calculated and compared with the experimental UV-Vis spectrum. The fundamental vibrational modes of the title compound have been precisely assigned and analyzed and the experimental results were compared with the theoretical values. A good agreement between experimental and calculated normal modes of vibrations has been observed. The NBO analysis indicates the intramolecular charge transfer between the bonding and antibonding orbitals. Molecular docking study has been performed by in silico method to analysis their HIVaspects against Abacavir carrier protein. Hydrogen bonds between docked Abacavir and amino acids residues of protein. Docked conformation of ligand in the binding site of Abacavir with 3VRI respectively.
International Letters of Chemistry, Physics and Astronomy Vol. 72