Prediction of MHC Binding Peptides and Antigenic Peptides from Ascaris lumbricoides

The parasitic disease Acariasis is the major concern because of its morbidity and mortality issue. In this investigation, we predicted the binding peptides of the MHC class I and MHC class II by Position Specific Scoring Matrices (PSSM) and Support Vector Machine (SVM) algorithms. We predicted the binding affinity of Cytochrome c oxidase subunit 2 (mitochondrion) having a 232 amino acids long residue sequence, which shows 224 nonamers. We predicted the peptide binding affinity to MHC I moles are as 8mer_H2_Db(The binding thresholds: 33.04,optimal score:52.494), 9mer_H2_Db(Optimal Score: 50.232, Binding Threshold: 17.96),10mer_H2_Db (The Optimal Score: 58.858, Binding Threshold: 41.32), 11mer_H2_Db(Optimal Score: 79.495, Binding Threshold: 56.96) and MHCII molecules are as I_Ab.p, I_Ad.p, I_Ag7. We also study integrates prediction of peptide MHC class I binding; proteasomal C terminal cleavage and TAP transport efficiency by using sequence and properties of the amino acids. We also found the binding of peptides to different alleles by using Position Specific Scoring Matrix. PSSM based server will predict the peptide binders of Cytochrome c oxidase subunit 2 from Ascaris lumbricoides sequence to, which are found antigenic epitopes region in protein.


Introduction
Ascaris lumbricoides is the causative agent of "Ascariasis" in human and other mammals by intestinal roundworm of the genus 'Ascaris'. This parasite is the one of the largest nematode and, the measures of the adult worm measures upto 15-35 cm long and are white or yellow in color. the incubation period of the parasite is 10-24 months in the jejunum and middle ileum of the intestine( of host). Study suggest that, te infected individual sheds the different percentages of the eggs (fertilized egg: 45%, fertilized and unfertilized eggs shedding: 40% and unfertilized eggs: 20%. The fertilized eggs becomes infectious with period of 5-10 days and remain vible for upto 17 months after gets released in the favorable soil [1]. The spreading of the infection is due to soil contamination of hands or food, ingestion, and the subsequent hatching of eggs in the small intestine. The occurrence of this parasite was observed as a zoonotic infection which usually associated with pigs and use of hog manure [2] but, in most endemic areas, it is most likely transmitted from person to person [3]. The infected individuals, experience the symptoms of pneumonitis including wheezing, dyspnea, nonproductive cough, hemoptysis, and fever, and eosinophilia. The larvae incubation timing in the jejunum expected as 65 days and feeds on digestion of the host. The infected children with a marginal diet found to be susceptible to protein, caloric, or suffer from Vitamin A deficiency, which stood up in growth retardation and increases the susceptibility to malaria infection [4] and sometime in overexposed situation, it causes relative immunodeficiency [5]. In the higher infection load of this worm causes severe complication whereas, at minimal level it remains asymptomatic. The occurrence of this disease is worldwide but predominantly found in that geographical area with warm and moist climates especially common in tropical and subtropical areas where there is poor practice of the sanitation and hygiene. The estimated worldwide preponderance of this disease is 25% (0.8-1.22 billion people) [6]. Children's of tropical and arising countries are more prone to this infection, where they are carried on by contamination of soil by human feces or use of untreated feces as fertilizer [7]. Symptomatic ascariasis may manifest as growth retardation, pneumonitis, intestinal obstruction, or hepatobiliary and pancreatic injury but, in most endemic areas, it is most likely transmitted from person to person [3]. The differential diagnoses are acute pancreatitis , biliary colic and community-acquired pneumonia. The vulnerable group for soil-transmitted helminth (STH) infections are children and as well as the impact of this infections directly on their health, nutritious status and cognitive ability [8]. The current strategy to control the STH infection is preventive chemotherapy with albendazole or mebendazole [9]. In the other study conducted by researcher in Chencha district where mass drug administration (MDA) to the most risky population including school-age children (SAC) is used as the central strategy to control soil-transmitted helminth (STH) infection [10]. In this study cytochrome c oxidase subunit II (mitochondrion) protein has been used to investigate its role in antigenicity. Cytochrome c oxidase subunit 2, also known as cytochrome c oxidase polypeptide II which is an oligomeric enzyme, an important component of the respiratory chain which involves in the transfer of electrons from cytochrome c to oxygen. This enzyme complex is found located in the mitochondrial inner membrane in eukaryotes. Cytochrome c oxidase subunit 2 contains two adjacent transmembrane regions in its Nterminus. The considerable part of the protein is generally exposed to the periplasmic or to the mitochondrial intermembrane space. The N-terminal domain of cytochrome C oxidase contains two transmembrane alpha-helices. Cytochrome oxidase deficiency and abnormality has been seen in the Leigh's disease. Investigation shows that any alterations in the catalytic genes of cytochrome c oxidase subunits I and II (COI and COII) have an adverse impact on prognosis in patients with acute myeloid leukaemia (AML) [11]. A "mitochondrial hypothesis" of late onset Alzheimer's disease (AD) has been proposed. The in depth biochemical studies propose that there is a significant decrease in cytochrome oxidase (CO) activity as well as perturbed CO I and CO III mRNA levels in platelets and brain tissue from Alzheimer's patients. The phenotypic expression study of the CO mutation is the major reason for reduced CO activity and compromised mitochondrial function [12]. Antigen protein prediction from Ascaris l umbricoides is necessary for few paradigms of synthetic vaccine development and target validation. Antigenicity prediction of the protein from Ascaris lumbricoides can play an important role in prototype synthetic vaccine development and as well as for target validation.

MHC binding peptide prediction
The major histocompatibility complex (MHC) peptide binding of Ascarislumbricoidespredicted using neural networks trained on C terminals of known epitopes. Rankpep predicting tool predicts peptide binders to MHC-I ligands using PSSMs, whose C-terminal end is likely to be the result of proteosomal cleavage. The sequence similarity is observed to the peptides that bind to a given MHC molecule. Traditionally, the sequence patterns used for the prediction of peptides binding to MHC molecules. Such sequence patterns, however, have proven to be too simple, as the complexity of the binding motif cannot be precisely represented by the few residues present in the pattern [29]. RANKPEP uses "Position Specific Scoring Matrices (PSSMs) or profiles" from set of aligned peptides known to bind to a given MHC molecule as the predictor of MHC-peptide binding and overcome the complexity of the binding motif limitation. RANKPEP web server is a variability masking feature to focus on the prediction of conserved epitopes, which could thus help to avoid immune evasion resulting from mutation. Support Vector Machine (SVM) based method for prediction of promiscuous MHC class II binding peptides from protein sequence; SVM has been trained on the binary input of single amino acid sequence [30][31][32][33].

Solvent accessible regions
We also analyzed the solvent accessible regions of proteins having highest probability that a given protein region lies on the surface of a protein Surface Accessibility, backbone or chain flexibility by Emini et al [35] and Karplus and Schulz [36]. The different scaleswere used to predict the hydrophobic and hydrophilic characteristics of amino acids which is rich in charged and polar residues. The methods used are Sweet et al., Kyte

The predicted peptide of MHC class I & II
We found the binding of peptides to a number of different alleles using Position Specific Scoring Matrix. cytochrome c oxidase subunit 2 (mitochondrion) of Ascaris lumbricoides antigen, with sequence 232 amino acid residues long, having 224nonamers. MHC molecules are cell surface proteins, which actively participate in host immune reactions and involvement of MHC-I and MHC-II in response to almost all antigens. We have predicted MHC-I peptide binders of cytochrome c oxidase subunit 2 (mitochondrion) from Ascaris lumbricoides. We found predicted MHC-I peptide binders of protein 4 test, we found the MHCI and MHCII binding regions. T cell immune responses are derived by antigenic epitopes hence their identification is important for design synthetic peptide vaccine. T cell epitopes are recognized by MHCI molecules producing a strong defensive immune response against Ascaris lumbricoides antigen cytochrome c oxidase subunit 2. Therefore, the prediction of peptide binding to MHCI molecules by appropriate processing of antigen peptides occurs by their binding to the relevant MHC molecules. Because, the C-terminus of MHCI-restricted epitopes results from cleavage by the proteasome and thus, proteasome specificity is important for determining T-cell epitopes. Consequently, RANKPEP also focus on the prediction of conserved epitopes. C-terminus of MHCI-restricted peptides is generated by the proteasome, and thus RANKPEP also determines whether the C-terminus of the predicted MHCI-peptide binders is the result of proteasomal cleavage. Moreover, these sequences are highlighted in purple in the output results. Proteasomal cleavage predictions are carried out using three optional models obtained applying statistical language models to a set of known epitopes restricted by human MHCI molecules as indicated as I_Ab.p, I_Ad.p,I_Ag7.p,I_Ak.p alleles, which is highlighted in red represent predicted binders. Peptides whose score is above the binding threshold will appear highlighted in red and peptides produced by the cleavage prediction model are highlighted in violet. We also use a cascade SVM based TAPPred method which found 72 High affinity TAP Transporter peptide regions (Table 3) which represents predicted TAP binders residues which occur at N and C termini from Ascaris lumbricoides cytochrome c oxidase subunit 2.

Prediction of solvent accessible regions of protein
We also predict solvent accessible regions in proteins; different measurement was performed for the prediction of antigenic activity, surface region of peptides. Emini et al., (Figure 7) predicts the highest probability in position:54 (Residue: K) i.e., 52-YFKSKK-57 with maximum score:5.745, that a given protein region lies on the surface of a protein and are used to identify antigenic determinants on the surface of proteins. Karplus             8 almost all antigens and it gives effects on specific sites. This knowledge of the immune responses to an antigen protein (cytochrome c oxidase subunit 2 from Ascaris lumbricoides) clear that the whole protein is not necessary for raising the immune response, but a small fragment of antigen can induce immune response against whole antigen. This means the increase in affinity of MHC binding peptides may result in enhancement of immunogenicity of cytochrome c oxidase subunit 2 from Ascaris lumbricoides, hence are helpful in silico to design and develop highly predictive computational tools for the identification of T-cell epitopes. Finally, accurate prediction remains vital for the future to design synthetic peptide vaccine. The Overall conducted study and opted results are encouraging. Both the 'sites of action' and 'physiological functions' can be predicted with very high accuracies which is helping to minimize the number of validation experiments. The future perspectives of this method will be useful in cellular immunology, vaccine design, immunodiagnostics, immunotherapeutic and molecular understanding of autoimmune susceptibility.