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Chikungunya Virus Infection
Published in Venkatesan Jayaprakash, Daniele Castagnolo, Yusuf Özkay, Medicinal Chemistry of Neglected and Tropical Diseases, 2019
D. Velmurugan, K. Manish, D. Gayathri
The nsP2 protease domain belongs to the papain super-family of cysteine proteases (Lulla et al. 2006, Sourisseau et al. 2007). Among the CHIKV-encoded enzymes, nsP2 constitutes an attractive target for the development of antiviral drugs. nsP2 is a multifunctional protein of approximately 90 kDa with a helicase motif in the N-terminal portion of the protein while papain-like protease activity resides in the C-terminal portion. The nsP2 protease is an essential enzyme whose proteolytic activity is critical for virus replication. Biochemical characterization of nsP2 cysteine protease of CHIKV has been reported (Pastorino et al. 2008). As an insight into the protease catalytic mechanism, they have studied the effect of different protease inhibitors on the enzymatic activity of CHIKV-nsP2 protease by using the cost-effective BOC-AGG-MCA peptide as substrate. The enzyme was found to be completely resistant to the inhibitors of serine protease (aprotinin), aspartic proteases (pepstatin) and metallo-proteases (EDTA). It has also proven resistance to the cysteine protease inhibitor leupeptin. Based on these findings, nsP2 can be classified as a thiol protease of the papain super family. Pastorino also confirmed that the viral activity of CHIKV-nsP2 protease is contributed by the amino acids in the range 422 to 799 in the C-terminal region of protease-helicase complex. The amino acids in the N-terminal segment (166 to 630) show NTPase or helicase activity (Karpe and Lole 2010).
Rotavirus
Published in Dongyou Liu, Laboratory Models for Foodborne Infections, 2017
Rotaviruses are recognized as the leading cause of severe gastroenteritis in infants and young children worldwide since the discovery of human rotavirus (HRV) in the early 1970s.1–3 Rotaviruses are transmitted via the fecal oral route, are highly contagious and stable in the environment,4 and post a high risk of foodborne infections. Rotavirus is a nonenveloped, 70-nm icosahedral virus in the family of Reoviridae. It has a genome consisting of 11 double-stranded RNA segments surrounded by a distinctive three-layered protein capsid.5 These 11 segments encode six structural proteins, VP1–4, VP6, and VP7, and six nonstructural proteins, NSP1–6.6 Among the 11 viral genes, the properties of the proteins encoded by genes 3, 4, 5, 9, and 10 are known to be related to rotavirus virulence in the host.7 Gene 3 encodes the capping enzyme that affects the level of viral RNA replication.8,9 Genes 4 and 9 produce the outer capsid proteins VP4 and VP7 required to initiate infection, and they induce virus neutralizing antibodies independently from each other.10,11 Gene 5 codes NSP1 that functions as an interferon antagonist.12–14 Gene 10 codes for the nonstructural protein NSP4, which regulates calcium homeostasis and virus replication and acts as an enterotoxin.7 NSP2 (encoded by gene segment 8) is also involved in virulence, especially in mice.15
Coronavirus Epidemics and the Current COVID-19 Pandemic
Published in Debmalya Barh, Kenneth Lundstrom, COVID-19, 2022
Aparna Bhardwaj, Prateek Kumar, Shivani Krishna Kapuganti, Vladimir N. Uversky, Rajanish Giri
NSP1 is the cleavage product of the action of PLpro from the divergent 5′ end sequence. The physiological function of SARS-CoV-2 NSP1 is not yet fully understood. Figure 1.4A shows the crystal structure of the NSP1 N-terminal region (polypeptide of residues 13–128) of SARS-CoV [56]. It blocks the translation in the host by binding to the ribosomal 40S subunit. Correlated computational and experimental studies showed the C-terminal of NSP1 as disordered, but it gains helical conformation when provided with organic solvents in its surrounding environment [57]. Much less is known about the NSP2 of SARS-CoV and SARS-CoV-2, but NSP2 has been suggested to suppress the host immune system, such as NSP1, and to interact with NSP3 [58]. NSP3 is a viral papain-like protease (PLpro) with cysteine in its active site (Figure 1.4B). PLpro cleaves polyproteins for the formation of mature proteins by identifying a specific motif sequence (LXGG) between proteins. It triggers the host innate immunity by its inhibitory action on the production of cytokines and chemokines [59]. It also interacts with NSP4 (predicted to be a transmembrane protein) along with NSP6 (which is also a transmembrane protein) to form a complex to modify the ER into double-membrane vesicles [58]. NSP5 is the main protease (Mpro), also known as 3C-like protease (Figure 1.4C). It is one of the vital enzymes for viral replication and transcription. It cleaves 11 conserved sites on the proteome after its autolytic cleavage from pp1a and pp1ab [60]. These proteases are centrally placed targets for antiviral drugs. However, it was recently reported that viruses mutate at certain hotspots in NSP5, resulting in impaired vaccine efficacy [61]. NSP11 is a small disordered protein of only 13 amino acids, with disordered-like characteristics even in the presence of some natural osmolytes [62].
Spectrum of candidate molecules against Chikungunya virus - an insight into the antiviral screening platforms
Published in Expert Review of Anti-infective Therapy, 2019
Shree Madhu Bhat, Piya Paul Mudgal, Sudheesh n, Govindakarnavar Arunkumar
CHIKV nsP2 based phenotypic assay has been used as a novel target to screen small molecule inhibitors. The ability of the nsP2 protein of CHIKV to induce a transcriptional shutoff allowing the virus to block cellular antiviral response is demonstrated using various luciferase-based reporter gene assays, including a trans-reporter system, where Gal4 DNA binding domain is fused to Fos transcription factor. This assay has been developed into a high-throughput screening system, and more than three thousand small molecules targeting nsP2-mediated shutoff have been screened using this platform. One compound, ID1452-2, which is a synthetic intermediate, was found to partially block nsP2 activity and inhibit CHIKV replication in vitro. Although the results were preliminary to identify the essential features in ID1452-2 mediating this action, the assay proved to be quite selective and could discriminate between structurally resembling molecules. Based on this proof of concept, similar functional assays can be developed towards identifying therapeutic molecules with innovative mechanisms [41].
Research progress in the development of porcine reproductive and respiratory syndrome virus as a viral vector for foreign gene expression and delivery
Published in Expert Review of Vaccines, 2020
Guo Dai, Mei Huang, To Sing Fung, Ding Xiang Liu
The nsp2-encoding region is highly diverse and variable in the PRRSV genome, where substitutions, deletions and insertions usually occur naturally [47]. In particular, gene deletions in the nsp2 hypervariable region often appeared in the field isolates or in vitro cultures of PRRSV, such as the representative PRRSV isolates NADC30-like, Em2007 and GDHY from China, and MN184 from the United States [4,32,48–50]. These isolates contain an over 30-amino acid (aa)-deletion in the nsp2 region relative to the North American prototype strain VR-2332 [4,32,48–50]. In addition, the attenuated PRRSV strain TJM containing a unique 120 aa deletion in nsp2 was successfully obtained by serial passages of the parental, highly pathogenic (HP) strain TJ on MARC-145 cells, thus becoming a good candidate for MLV and marker vaccines against HP-PRRSV [51]. Previous studies on deletions in the nsp2 region with mutant viruses derived from the infectious clone of type 2 strain VR-2332 indicated that deletion up to 403 aa in the hypervariable region did not affect the rescue of infectious virus in vitro [52]. On the other hand, the predicted nsp2 region of strain SP contains an insertion with 36 or 155 aa, compared to the nsp2 gene of VR-2332 and LV [53], suggesting that the nsp2 region of PRRSV can tolerate the insertion of foreign genes. In addition, nsp2 is an immunodominant protein with strong immunogenicity. Pigs inoculated with PRRSV can induce strong antibody responses against nsp2 within 2 weeks [54]. Deletion of some antigenic epitopes in the non-essential region of nsp2 would be an attractive approach for the development of marker vaccines.
Whole genome amplicon sequencing and phylogenetic analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from lineage B.1.36.27 isolated in Hong Kong
Published in Expert Review of Molecular Diagnostics, 2022
Hin Fung Tsang, Allen Chi Shing Yu, Heong Ting Wong, Wai Ming Stanley Leung, Jiachi Chiou, Yin Kwan Evelyn Wong, Aldrin Kay Yuen Yim, Dominic Ngai Chong Tsang, Alan KL Tsang, Wing Tak Wong, Sze Chuen Cesar Wong, William Chi Shing Cho
Regarding amino acid mutations, the most common amino acid mutations were V561A, V1056L, A1997V, P314L, D614G, Q57H, T223I, and S194L. These amino acids mutations were found in all 9 SARS-CoV-2 lineage B.1.36.27 genome analyzed. Amino acids mutations may alter the protein structure and hence affecting protein–protein interactions. V561A mutation is located in the non-structural protein 2 (nsp2). It is proposed that nsp2 might be involved in the disruption of intracellular host signaling during viral infection [14]. V1056I and A1997V mutations are located in the non-structural protein 3 (nsp3), which is involved in polyprotein processing, de-ADP-ribosylation, de-ubiquitination, and de-ISGylation [15]. P314L mutation is located in ORF1b and encoding the nsp12, which is the viral RNA-dependent RNA polymerase (RdRp) [16,17]. Some studies suggested that D614G mutation is in linkage disequilibrium with P314L mutation [17]. D614G mutation is located in the S protein. The presence of the D614G mutation on the S protein can strengthen the folding stability of the S protein and ORF8 protein [16]. Previous study reported that the presence of D614G mutation in the S protein could increase the viral infectivity by enhancing the binding affinity to host angiotensin-conversing enzyme 2 (ACE2) receptor as well as the viral replication and transmission [18,19]. D614G mutation enhances viral replication in the upper respiratory tract and it might be one of the most significant mutations leading to the enhanced transmission of the virus during the fourth wave of COVID-19 outbreak in Hong Kong. Q57H and T223I mutations are located in ORF3a. A recent study indicated that T223I mutation is a destabilizing mutation on the stability of the ORF3a protein by calculating the difference in free energy [20]. In conjunction with the presence of Q57H mutation on ORF3a protein, these mutations might affect the binding affinity and harm the function of ORF3a in apoptosis, resulting in an increase in the viral load in the host cell [16,18]. S194L mutation is located in N gene. The presence of this mutation might enhance the interaction between N protein and E protein, promoting the release of the virus [18]. However, the binding affinity between this N mutant and M protein is decreased. It might attenuate the assembly of the virus [18]. Of the nine analyzed SARS-CoV-2 lineage B.1.36.27 genome sequences, T4304I was identified in the ORF1a of four genome sequences. A1087S and C1236F were identified in the S gene of three genome sequences and one genome sequence, respectively. So far, no researches have been performed on the effects of these mutations. Further studies might be necessary to analyze the impacts of the above mutations to the virulence and transmissibility of SARS-CoV-2. No vaccine escape mutations were identified from the genome of SARS-CoV-2 from lineage B.1.36.27, suggesting this strain is not likely to compromise existing vaccines and antibody therapies [21].