Retinol binding proteins (RBPs) are a family of carriers for fat-soluble retinol (preformed vitamin A). Vitamin A is a significant nutrient that performs a vital role in vision, cell growth and differentiation, and embryonic growth in the Coturnix japonica or japenese quail [1]. Retinol binding protein (RBP7) is one of the cellular retinol binding proteins that play a role in cellular metabolism of retinol. Vitamin A is consumed from dietary sources as a retinyl ester or synthesized from β-carotene and is deposited in the liver as a retinyl ester up to it is mobilized for transmission into numerous target tissues. Retinol is one of the forms of vitamin A found from foods of animal origin. Retinal (retinaldehyde), the aldehyde derived from retinol, is essential for vision, while retinoic acid is essential for skin health and bone growth in Coturnix japonica. These chemical compounds are mutually recognized as retinoids, having the same structural motif means having all-trans double bonds found in retinol. Structurally, all retinoids possess a β-ionone ring and a polyunsaturated side chain containing an alcohol, an aldehyde, a carboxylic acid group or an ester group [1]. Also the RBP7 contains controlling components for adipose-specific expression. The RBP7 has a novel adipose tissue-specific promoter. RBP7 is recognizing as an adipose-specific gene in the Coturnix japonica or japenese quail in adipose tissue under the control of RBP7 promoter expression in adipose tissue. The RBP7 promoter may stimulate expression very strongly in the neck and abdominal adipose tissue of Coturnix japonica or japenese quail [2]. Retinoids mostly bind in two ways such as Plasma retinol binding protein (RBP) and Epididymal retinoic acid binding protein (ERABP) carries retinoid in body fluids, while cellular retinol binding proteins (CRBPs) and cellular retinoic acid- binding proteins (CRABPs) carry retinoids within cells [3]. RBP, ERABP, CRBPs i.e. CRBP I, II, III, and IV and CRABPs i.e. CRABP I and CRABP II belong to the lipocalins superfamily in the Structural Classification of Proteins (SCOP) database. The RBP and ERABP belong to their retinol-binding protein-like (RBP) family. CRBPs and CRABPs belong to the fatty acid-binding protein-like (FABP) family [4]. RBP is the specific carrier for retinol (vitamin A alcohol) in the blood [5]. RBP7 plays a key role in terms of development of growth of body weight depends upon the connective tissues known as adipose tissue. The location of an increasing number of new adipose-specific genes has expressively contributed to understand of adipose tissue biology and the etiology of obesity and its related diseases in Coturnix japonica. Comparison of gene expression profiles among various tissues performed by analysis of chicken microarray data, leading to identification of RBP7 as a novel adipose-specific gene in Japanese quail. Adipose-specific expression of RBP7 in the avian species was further confirmed at the protein and mRNA levels [2]. The amount of adipose tissue describes the total energy of body, responsible for the metabolism of the body. In this study we are going to predict and analyse the structure of Retinol Binding Protein 7 (RBP7) and calculate the energy of RBP7 in Coturnix japonica. The position of residues and the residues radius is predicted in Pymol- v1.8.6.0.tar. The energy calculation and the minimum energy calculation of Retinol Binding Protein 7 (RBP7) calculated in Swiss PDB Viewer4.1.0. The Ramachandran Plot assessment is under the analysis in which the φ, ψ values for the amino acids of RBP7 is study under Molprobity.

Collection of the data using PUBMED

Collecting the data for structure prediction of protein present in Coturnix japonica and retrieving of the sequence from NCBI of retinol binding protein 7 Coturnix japonica (Table 1).

Table 1: Collection of the data using PUBMED.

RaptorX

A web-based method using RaptorX (http://raptorx.uchicago.edu/) for protein secondary structure prediction, template based tertiary structure modelling, alignment quality assessment and probabilistic alignment sampling. Raptor X web server delivers high-quality structural models for many targets with only remote templates. Because of this Computational modelling of the three-dimensional atomic arrangement of the amino acid chain is also possible in determining the role of the protein in biological processes [6].

Swiss PDB Viewer4.1.0

Swiss-PdbViewer is an application method that runs a user friendly interface allowing analysing proteins. Amino acid mutations, H-bonds, angles and distances between atoms are easy to obtain in form of graphic and menu interface. Moreover, Swiss-PdbViewer is tightly linked to Swiss-Model, an automated homology modelling server accessible from ExPASy [7].

Pymol- v1.8.6.0.tar

PyMOL is an open-source tool to visualize molecules. It runs on Windows, Linux and MacOS equally well. PyMOL has competences in creating high-quality images from 3D structure; it has well established purposes for influencing structures and some elementary functions to evaluate their chemical properties. The opportunities to write scripts and plugins as well as to integrate PyMOL in custom software are vast and superior to most other programs. PyMOL has been written mostly in the Python language (www.python.org), while the time-critical parts of the system have been coded in C. This way, Python programs intermingle most effortlessly with the PyMOL GUI [8].

Ramachandran assessment by Mol probity

A Ramachandran plot is a visual graphic demonstration of the main-chain conformational tendencies of an amino acid. Ramachandran plot based on experimental data is deciding whether scant data represent genuine conformations. Measured the pair-wise distances of main-chain conformational tendencies among amino acids, and showed the conformational relationships of amino acids are well preserved in a two-dimensional map, leading to the conclusion that the conformational diversity space of amino acids is largely two dimensional. Amino acids in early and late evolutionary phases are situated in different zones in the two-dimensional map [9]. The first is a Ramachandran plot or Ramachandran map, which is simply a scatter plot of the φ,ψ values for the amino acids in a single protein structure or a set of protein structures. It may be restricted to a single amino acid type and/or a single structural feature type, such as protein loops. The second is a Ramachandran distribution, a statistical representation of Ramachandran data, usually in the form of a probability density function. A probability density function gives the probability of finding an amino acid conformation in a specific range of φ,ψ values. For instance, if the function is given on a 10° × 10° grid from −180° to +180° in φ,ψ (1296 values), then the distribution may give the probability per 10° × 10° region. It could also be expressed per degree squared or per radian squared. Such distributions may be derived for specific amino acid types and/or for specific structural features. There are numerous significant considerations in developing Ramachandran distributions from structural data, depending on the purpose of the derived distribution. First, while glycine and proline are usually treated separately, the other 18 amino acids are often treated as a single type. However, these amino acids are moderately different in their proportions of residues in the α, β, polyproline II, and left-handed helical regions. Second, relatively different distributions are resolute when either all residues are used or only those outside the regular secondary structures of α-helices and β-sheets. The concluding assumed to be “intrinsic” favourites of the backbone, not subjective by forming specific hydrogen bonds present in regular secondary structures. Third, the quality and quantity of the data are crucial in determining distributions meant to act as eminence filters for newly determined structures or for structure prediction. As more structures have become available at higher resolutions, it is now possible to use quite large datasets with resolution cut-offs of 1.8 Å or even better. Other filters have been used including B-factors and steric clashes to remove residues that may be model improperly or at least with considerable uncertainty within the electron density. For instance, by using higher resolution structures, B-factors, steric overlaps able to determine Ramachandran distributions with smaller “allowed” and “generously allowed” regions than previous efforts. Fourth, most previous efforts have involved density estimation using simple histogram methods – the counts or proportion of counts of residues in non-overlapping square bins of the φ,ψ space. However, even when a large number of proteins are used, the distribution in φ,ψ space may be quite uneven (Figure 1) [10-12].

Figure 1: Graphical representation of RBP7 structure prediction.

>AKC91103.1 retinol binding protein 7 [Coturnix japonica]

MPVDFSGTWNLVSNDNFEGYMTALGIDFATRKIAKMLK-PQKVIKQDGDSFSIHTTSTFRDYMLQFKIGEEFEEDNKGLD-NRKCKSLVTWDNDKLICVQAGEKKNRGWTHWLEGDDLHLEL-RCEN QVCKQVFKRA

GDT uSeqID: 92 101 (raptorX)

SeqID ModelName Template(s): 75 6at8A-279444_1 6at8A

Rank P-value Score uGDT: 9.0e-12 158 123 (Figures 2-5).

Figure 2: Retinol Binding Protein 7(RBP7) in Coturnix japonica using RaptorX.

Figure 3: Position of Residues of Retinol binding protein 7 in Coturnix japonica using Pymolv-1.8.6.0.

Figure 4: Value of radius of residues of Retinol Binding Protein 7 in Coturnix japonica using Pymolv-1.8.6.0.

Figure 5: Ramachandran Validation using Mol probily (Vincent B. Chen, W. Bryan Arendall III, Jeffrey J. Headd, Daniel A. Keedy).

The total energy of retinol binding protein 7 (RBP7) residues in Coturnix japonica or japenese quail is 1047.341 KJ/mol by computational calculation. The calculation is based on position of amino acids, bonds, angles, torsion, improper, non-bonded and total energy of amino acids making a retinol binding protein7 (RBP7) in japenese quail (Supplementary Data). The energy minimization of amino acids of RBP7 is also calculated by Swiss Pdb Viewer. In which the minimum energy of Bonds is 936.766 KJ/mol, angles 900.843 KJ/mol, torsion 582.304 KJ/ mol, improper 236.255 KJ/mol, nonbonded -1629.27 KJ/mol, and total minimum energy is 1026.898 KJ/mol in japenese quail. This energy is responsible for the threshold value to express the adipose gene and other significant functions in japenese quail. This RBP7 is responsible for major adipose tissue depositions. This expression is balanced with the help of energy. A scatter plot of the φ,ψ values for the amino acids of RBP7 is also analyse. An statistical representation in the form of a probability density function. A probability density function gives the probability of finding an amino acid conformation in a specific range of φ,ψ values. First glycine and proline are usually treated separately; the other 18 amino acids are often treated as a single type. However, these amino acids are moderately different in their proportions of residues in the α, β, polyproline II, and left-handed helical regions. The “intrinsic” favourites of the backbone, not particular by forming specific hydrogen bonds present in regular secondary structures. 97.0% (128/132) of all residues are in favoured (98%) regions. 99.2% (131/132) of all residues were allowed (>99.8%) regions. One outliers (phi, psi) are 2 PRO (-48.1, -72.3) in validation. The position of amino acids, the Position of residues of amino acids and values of bonds, torsion, improper non bonded and total energy calculation of amino acid of RBP7 in Coturnix japonica or japense quail is given below:

1) ATOM 1-8,162-169,275-282,488-495: MET (M).

2) ATOM 9-15,300-306: PRO (P).

3) ATOM 16-22,89-95,325-331, 694-700,777-783,1020-1026,1051-1057: VAL (V)

4) ATOM 23-30,110-117,202-209,358-365,370-377,468-475,592- 599,629-636,722-729, 738-745,918-933: ASP (D).

5) ATOM 31-41, 126-136,210-220,384-394,446-456,513-523,563- 573,1058-1068: PHE (F)

6) ATOM 42-47, 96-101, 378-383,395-400,433-438,680-685: SER (S).

7) ATOM 48-51,146-149,190-193,366-369,541-544,617-620,798- 801,848-851,914-917: GLY (G)

8) ATOM 52-58,170-176,226-232,419-432,439-445,701-707,866- 872: THR (T)

9) ATOM 59-72, 708-721,852-865,883-896: TRP (W)

10) ATOM 73-80,102-109,118-125,600-607,637-644,730-737,829- 836, 1003-1010: ASN (N).

11) ATOM 81-88,182-189,283-290,496-503,621-628,686-693,755-762,897-904,934-941, 952-959,969-976:LEU (L)

12) ATOM 137-145, 545-562, 574-591,802-810,905-913,960- 968,994-1002:GLU (E)

13) ATOM150-161, 476-487: TYR (Y).

14) ATOM 177-181,221-225,261-265,793-797, 1089-1094:ALA (A)

15) ATOM 194-201,253-260,332-339,401-408,533-540,763-770: ILE (I).

16) ATOM 233-243,257-267,645-655,837-847,977-987, 1078- 1088: ARG (R).

17) ATOM244-252,266-274,291-299,316-324,340-348,524- 532,608-616,656-664,671-679, 746-754,811-828, 1033-1041, 1069-1077: LYS (K).

18) ATOM 307-315, 349-357,504-512,784-792, 1011-1019, 1042- 1050: GLN (Q).

19) ATOM 409-418,873-882,942-951: HIS (H).

20) ATOM 665-670, 771-776,988-993, 1027-1032: CYS (C) (Table 2).

Table 2: Position of Residues of Amino Acids and Values of Bonds, Torsion, Improper, Non Bonded and Total Energy Calculation of Amino Acid of RBP7 in Coturnix Japonica or japense quail.

Authors thanks Ankita Sethia, Swati Srivastava, Khushboo Arya and Dolly Chauhan for their critical inputs in this manuscript.