Retrieval of the targeted sequences
A total of 182 Human Metapneumovirus Fusion Protein sequences were obtained from the NCBI database [41] as the FASTA format in August 2018. The accession numbers of the obtained sequences with the area and date of the collection were listed in Table 6.
Table 6
the retrieved Human Metapneumovirus Fusion Protein sequences with their Accession number, Date and area of Collection
Accession number
|
Date of collection
|
country
|
YP_012608
|
2004
|
USA
|
BBB35016
|
2015
|
japan
|
BBB35015
|
2015
|
Japan
|
BBB35014
|
20/3/2015
|
Japan
|
BBB35013
|
2015-03-16"
|
Japan
|
BBB35012
|
2015-03-16"
|
Japan
|
BBB35011
|
2015-02-27
|
Japan
|
BBB35010
|
2015-02-21
|
Japan
|
BBB35009
|
2015-02-21
|
Japan
|
BBB35008
|
2015-02-21"
|
Japan
|
BBB35007.1
|
2014-04-12
|
Japan
|
BBB35006.1
|
2014-04-13"
|
Japan
|
BBB35005.1
|
2014-04-05
|
Japan
|
BBB35004.1
|
2013-07-08"
|
Japan
|
BBB35003.1
|
2013-06-22"
|
Japan
|
BBB35002.1
|
2013-06-24
|
Japan
|
BBB35001.1
|
2013-06-24
|
Japan
|
BBB35000.1
|
2013-04-30
|
Japan
|
BBB34999.1
|
2013-04-16
|
Japan
|
BBB34998.1
|
2013-04-08"
|
Japan
|
BBB34997.1
|
2013-03-25
|
Japan
|
BBB34996.1
|
2013-01-21
|
Japan
|
BBB34995.1
|
2015-03-02
|
Japan
|
BBB34994.1
|
2014-08-04
|
Japan
|
BBB34992.1
|
2013-06-25
|
Japan
|
BBB34991.1
|
2013-06-24
|
Japan
|
BBB34990.1
|
2013-06-22
|
Japan
|
ANW38002.1
|
2011
|
Croatia
|
ANW38000.1
|
2013
|
Croatia
|
ANW37997.1
|
2012
|
Croatia
|
ANW37998.1
|
2012
|
Croatia"
|
ANW37996.1
|
2012
|
Croatia
|
ANW37993.1
|
2011
|
Croatia"
|
ANW37992.1
|
2011
|
Croatia"
|
ANW37991.1
|
2011
|
Croatia"
|
ANW37990.1
|
2011
|
Croatia
|
ANW37989.1
|
2011
|
Croatia
|
AII17595.1
|
10-Aug-2011
|
South Korea
|
Determinant of the conserved regions
The obtained Sequences subjected to the multiple alignment tests via the CLUSTALW algorithm [42] on BioEdit software [43] version 7.0.9.1 to identify the conserved regions.
B-Cells Epitopes Prediction
The linear Epitope Prediction tool BepiPred-test on the Immune Epitope Database (IEDB) [44] (figure ).
The epitopes were predicted at a default threshold value of (0.4) from the conserved region in the Human Metapneumovirus Fusion Protein sequences that obtained from the multiple sequences, alignment. The prediction performed using the Markov model [45]. The results listed in Table 1.
The Surface Accessibility Prediction
The Emini surface accessibility prediction tool [43] on the IEDB [46] used to predict the surface accessibility with a default threshold value for each conserved region (Fig. 9). The results are listed in Table 1.
The Antigenic Sites Prediction
The Kolaskar and Tongaonker antigenicity tool on IEDB [47] used for the prediction of the antigenic sites within the Human Metapneumovirus Fusion Protein sequences at a default threshold value of 1.04. The results listed in Table 1, (Fig. 10). For the whole first alignment, sequences see (Fig. 11).
T-Cell Epitopes Prediction
The prediction of the cytotoxic T-cell epitopes performed by using the Major Histocompatibility Complex class I (MHC I) binding prediction tool on IEDB [48]. The epitopes' length adjusted at 9. The conserved epitopes that bind with various HLA alleles at score equal or less than 1.0 percentile rank and 500 IC50 selected for further analysis. Moreover, the prediction of T-cell helper epitopes performed by using the Major Histocompatibility Complex class II (MHC II) binding prediction tool of IEDB [49]. The results listed in Table 3.
The Population Coverage Prediction
The prediction of epitopes binding with various MHC I and MHC II alleles that cover the world population was performed by using the population coverage tool on the IEDB [50]. The results are listed in Table 4.
The Peptides Allergicity and Toxicity Prediction
The AllergenFP v.1.0 [47] web servers used to predict the Allergenicity. Furthermore,
ToxinPred web server [48] used to predict the Toxicity. The results listed in Table 5.
The 3D Structure Modeling and Visualization
The 3D structure of the Human Metapneumovirus Fusion Protein modeled by using the Phyre2 web portal [49] and 3D structure the predicted peptides we modeled by using PEP-FOLD 3.5 web server [50]. The modeled 3D structures visualized by Chimera 1.8 software [51]. The results presented in Figs. 1, 2, and .
5 The Molecular Docking Study
The predicted epitopes were docked with MHC I and MHC II molecules. The Protein Databank [52] was used to obtain the 3D structures of MHC I and MHC II. The 3D structures (PDBIDs:
6AM5, 5TXS, and 5NI9 for MHC IA, MHC IB, and MHC II respectively) were downloaded in PDB format. The. The structures were prepared, minimized for the docking process, and the molecular docking calculation performed by using Cresset Flare software [53]. The results listed in Table 5 and showed in Figs. 4,5,6, and 7.