Abstract
BackgroundInteraction of non-structural protein 5A (NS5A) of Hepatitis C virus (HCV) with human kinases namely, casein kinase 1α (ck1α) and protein kinase R (PKR) have different functional implications such as regulation of viral replication and evasion of interferon induced immune response respectively. Understanding the structural and molecular basis of interactions of the viral protein with two different human kinases can be useful in developing strategies for treatment against HCV.ResultsSerine 232 of NS5A is known to be phosphorylated by human ck1α. A structural model of NS5A peptide containing phosphoacceptor residue Serine 232 bound to ck1α has been generated using the known 3-D structures of kinase-peptide complexes. The substrate interacting residues in ck1α has been identified from the model and these are found to be conserved well in the ck1 family. ck1α – substrate peptide complex has also been used to understand the structural basis of association between ck1α and its other viral stress induced substrate, tumour suppressor p53 transactivation domain which has a crystal structure available.Interaction of NS5A with another human kinase PKR is primarily genotype specific. NS5A from genotype 1b has been shown to interact and inhibit PKR whereas NS5A from genotype 2a/3a are unable to bind and inhibit PKR efficiently. This is one of the main reasons for the varied response to interferon therapy in HCV patients across different genotypes. Using PKR crystal structure, sequence alignment and evolutionary trace analysis some of the critical residues responsible for the interaction of NS5A 1b with PKR have been identified.ConclusionsThe substrate interacting residues in ck1α have been identified using the structural model of kinase - substrate peptide. The PKR interacting NS5A 1b residues have also been predicted using PKR crystal structure, NS5A sequence analysis along with known experimental results. Functional significance and nature of interaction of interferon sensitivity determining region and variable region 3 of NS5A in different genotypes with PKR which was experimentally shown are also supported by the findings of evolutionary trace analysis. Designing inhibitors to prevent this interaction could enable the HCV genotype 1 infected patients respond well to interferon therapy.
Highlights
Interaction of non-structural protein 5A (NS5A) of Hepatitis C virus (HCV) with human kinases namely, casein kinase 1α and protein kinase R (PKR) have different functional implications such as regulation of viral replication and evasion of interferon induced immune response respectively
Modelling of NS5A peptide bound to human casein kinase 1α model Protein kinases recognize their substrates usually by means of a substrate recognition motif surrounding the phosphoacceptor residue [35]
Prediction of PKR interacting residues of NS5A 1b by evolutionary trace analysis We have predicted the PKR interacting residues in NS5A of genotype 1b by a computational analysis which is entirely independent of the analysis reported in the previous section
Summary
Interaction of non-structural protein 5A (NS5A) of Hepatitis C virus (HCV) with human kinases namely, casein kinase 1α (ck1α) and protein kinase R (PKR) have different functional implications such as regulation of viral replication and evasion of interferon induced immune response respectively. Experimental work have shown that for the rest of the protein whose structure is currently unknown it is natively unfolded [8], lacks secondary structural elements, is less hydrophobic, has high content of positively charged residues, has low complexity regions and many phosphorylation sites which predominantly occurs in intrinsically disordered regions of the protein [9,10,11]. The natively unfolded nature of NS5A makes it capable of interacting with many human as well other nonstructural proteins of HCV thereby carrying out multiple functions [12]
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