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At least 55 records · Page 3Linked to original sources
PubMed2001

Internally located signal peptides direct hepatitis C virus polyprotein processing in the ER membrane.

An endoplasmic reticulum (ER) signal peptide is an amino acid sequence motif that directs the translocation of nascent polypeptides to the lumen of ER membrane. Most of known ER signal peptides are either N-terminal cleavable or internally uncleavable. In the structural protein region of hepatitis C virus (HCV) polyprotein, however, four internally located cleavable signal peptides are arranged in a tandem array. The published experimental results indicated that the nascent HCV polyprotein is processed in the ER membrane by host signal peptidase(s) to the respective viral proteins. Here we propose that the four ER signal peptides lead the nascent HCV polyprotein to ER membrane, and the four internally located cleavable signal peptides are the sole determinant for the compartment localization of the matured viral proteins. After cleavage at the C-terminus, the signal peptides retain at the C-terminus of mature proteins, and serve as ER membrane anchors. The signal peptide directed polyprotein processing in the ER membrane preludes the virion assembly and budding from the ER membrane. This unique processing may be a general mechanism adopted by many types of virus for virion assembly and replication. The revelation of signal peptidase involved in HCV polyprotein processing presents a novel drug target to suppress HCV viral replication for the much needed HCV therapy.

Amino Acid Sequence↗
PubMed1993

Polyprotein processing in African swine fever virus: a novel gene expression strategy for a DNA virus.

This report shows that African swine fever virus (ASFV)--a large DNA-containing virus--synthesizes a polyprotein to produce several of its structural proteins. By immunoprecipitation analysis, we have found that ASFV polyprotein is a 220 kDa myristoylated polypeptide (pp220) which, after proteolytic processing, gives rise to four major structural proteins: p150, p37, p34 and p14. Processing of the ASFV polyprotein takes place at the consensus sequence Gly-Gly-X and occurs through an ordered cascade of proteolytic cleavages. So far, polyprotein processing as a mechanism of gene expression had been found only in positive-strand RNA viruses and retroviruses. According to the results presented here, ASFV is the first example of a DNA virus that synthesizes a polyprotein as a strategy of gene expression.

African Swine Fever Virus↗
PubMed1993

Mutagenesis of conserved residues at the yellow fever virus 3/4A and 4B/5 dibasic cleavage sites: effects on cleavage efficiency and polyprotein processing.

Flavivirus proteins are produced by co- and post-translational proteolytic processing of a large polyprotein using both host- and virus-encoded enzymes. The flavivirus serine proteinase, which consists of NS2B and NS3, is responsible for cleavages of at least four dibasic sites in the nonstructural region. In this study, a number of substitutions for the conserved amino acids flanking the 3/4A and 4B/5 dibasic cleavage sites [Arg(P2)-Arg(P1) decreases Gly(P1')] were examined for their effects on yellow fever virus (YF) polyprotein processing. The substrate for these studies was a truncated YF polyprotein, called sig2A-5(356), which consists of a signal sequence fused to NS2A and extending through the first 356 amino acids of NS5. At the P1' position (Gly) of the 4B/5 site, only Ser and Ala were allowed while six other substitutions abolished cleavage. Substitutions of the 4B/5 P1 Arg residue with Lys, Gln, Asn, or His were tolerated while replacement with Glu eliminated cleavage. The 4B/5 P2 position (Arg) was found to be tolerant of substitutions with polar or hydrophobic residues which allowed varying degrees of partial cleavage. Previous studies have shown that cleavage at the 3/4A site is incomplete in YF-infected cells and that the cleavage efficiency at this site is significantly less for the sig2A-5(356) polyprotein. Replacement of the 3/4A P1 Arg residue with noncharged polar or hydrophobic residues reduced the cleavage efficiency, whereas substitutions with Glu or Pro abolished cleavage. Studies with polyproteins containing one or both of the 3/4A and 4B/5 cleavage sites blocked indicate that there is not an obligatory processing order for cleavages generating the N termini of YF NS4A, NS4B, and NS5.

Amino Acid Sequence↗
PubMed1994

Proteinase 3C of hepatitis A virus (HAV) cleaves the HAV polyprotein P2-P3 at all sites including VP1/2A and 2A/2B.

Thus far, the only virus-encoded proteinase of hepatitis A virus (HAV) detected is 3C, which was shown to catalyze proteolysis of most of the suggested cleavage sites within the HAV precursor polyprotein. To elucidate whether or not HAV proteinase 3C and its precursors are involved in processing of the yet unidentified sites in the polyprotein P2-P3, the genomic region of 3C including flanking sequences were expressed in a bacterial system and by cell-free translation. In both systems 2A-reactive proteins of 10 (2A) and 16 kDa (delta VP1-2A) were processing products of a polyprotein representing delta VP1-P2-P3* (delta and * denote N- or C-terminally truncated proteins, respectively), thus providing evidence for cleavage at sites VP1/2A and 2A/2B by proteinase 3C. In the cell-free expression system, processing at the P2/P3 junction was rapid and complete, whereas sites 3A/3B, 3B/3C, and 3C/3D were inefficiently cleaved, as evidenced by the accumulation of the stable precursor polypeptides P3* and 3ABC. In contrast to the eukaryotic system, mature 3C was produced in Escherichia coli. Intermolecular cleavage by recombinant 3C occurred at all putative sites within the proteolytically inactive polyprotein P2-P3* mu. The results of this study indicate that proteinase 3C mediates the primary as well as the secondary cleavages of the HAV polyprotein and thus shows an activity profile broader than that of 3C proteinases of other picornaviruses.

3C Viral Proteases↗
PubMed1995

Characterisation and mutational analysis of an ORF 1a-encoding proteinase domain responsible for proteolytic processing of the infectious bronchitis virus 1a/1b polyprotein.

Coronavirus gene expression involves proteolytic processing of the mRNA 1-encoded polyproteins by viral and cellular proteinases. Recently, we have demonstrated that an ORF 1b-encoded 100-kDa protein is proteolytically cleaved from the 1a/1b fusion polyprotein by a viral-specific proteinase of the picornavirus 3C proteinase group (3C-like proteinase). In this report, the 3C-like proteinase has been further analysed by internal deletion of a 2.3-kb fragment between the 3C-like proteinase-encoding region and ORF 1b and by substitution mutations of its catalytic centre as well as the two predicted cleavage sites flanking the 100-kDa protein. The results show that internal deletion of ORF 1a sequences from nucleotide 9911 to 12227 does not influence the catalytic activity of the proteinase in processing of the 1a/1b polyprotein to the 100-kDa protein species. Site-directed mutagenesis studies have confirmed that the predicted nucleophilic cysteine residue (Cys2922) and a histidine residue encoded by ORF 1a from nucleotide 8985 to 8987 (His2820) are essential for the catalytic activity of the proteinase, and that the QS(G) dipeptide bonds are its target cleavage sites. Substitution mutations of the third component of the putative catalytic triad, the glutamic acid 2843 (Glu2843) residue, however, do not affect the processing to the 100-kDa protein. In addition, cotransfection experiment shows that the 3C-like proteinase is capable of trans-cleavage of the 1a/1b polyprotein. These studies have confirmed the involvement of the 3C-like proteinase domain in processing of the 1a/1b polyprotein, the predicted catalytic centre of the proteinase, and its cleavage sites.

Amino Acid Sequence↗
PubMed1998

Processing of the MHV-A59 gene 1 polyprotein by the 3C-like proteinase.

The 3C-like proteinase of mouse hepatitis virus (MHV-3CLpro) is predicted to cleave at least 10 sites in the gene 1 polyprotein, resulting in processing of proteinase, polymerase and helicase proteins from the polyprotein. We have used E. coli expressed recombinant 3CLpro (r3CLpro) to define cleavage sites in carboxy-terminal region of the ORF 1a polyprotein. Polypeptides containing one or more putative 3CLpro cleavage site were translated in vitro from subcloned regions of gene 1, and the polypeptides were incubated with r3CLpro. Analysis of the cleavage products confirmed several putative cleavage sites, as well as identifying cleavage sites not previously predicted by analysis of the MHV sequence. Antibodies directed against a portion of the ORF 1a polyprotein were used to probe virus infected cells, and detected proteins that correspond to the cleavage sites used by 3CLpro in vitro. These results suggest that MHV 3CLpro cleaves at least 7 sites in the ORF 1a polyprotein, and that the specificity of 3CLpro for cleavage site dipeptides may be broader than previously predicted.

Animals↗
PubMed1989

Computer analysis suggests a role for signal sequences in processing polyproteins of enveloped RNA viruses and as a mechanism of viral fusion.

We have used a computer program to scan the entire sequence of viral polyproteins for eucaryotic signal sequences. The method is based on that of von Heijne (1). The program calculates a score for each residue in a polyprotein. The score indicates the resemblance of each residue to that at the cleavage site of a typical N-terminal eucaryotic signal sequence. The program correctly predicts the known N-terminal signal sequence cleavage sites of several cellular and viral proteins. The analysis demonstrates that the polyproteins of enveloped RNA viruses--including the alphaviruses, flaviviruses, and bunyaviruses--contain several internal signal-sequence-like regions. The predicted cleavage site in these internal sequences are often known cleavage sites for processing of the polyprotein and are amongst the highest scoring residues with this algorithm. These results indicate a role for the cellular enzyme signal peptidase in the processing of several viral polyproteins. Not all high-scoring residues are sites of cleavage, suggesting a difference between N-terminal and internal signal sequences. This may reflect the secondary structure of the latter. Signal sequences were also found at the N-termini of the fusion proteins of the paramyxoviruses and the retroviruses. This suggests a mechanism of viral fusion analogous to that by which proteins are translocated through the membranes of the endoplasmic reticulum at synthesis.

Algorithms↗
PubMedDate not supplied

Organization of HIV-1 pol is critical for Pol polyprotein processing.

The HIV pol sequentially encodes protease (PR), reverse transcriptase (RT), and integrase (IN) from the 5'-3' direction. We explored the significance of this gene arrangement. All six possible gene dispositions were examined. In two situations where PR was removed from the leading place and no two genes were in their original location, viral polyprotein processing was abolished. Processing of the polyprotein did not occur when IN was translocated to the front of PR-RT. However, in the following two arrangements, the polyprotein was processed but only at specific sites. First, PR remained in the leading position while the locations of RT and IN were exchanged; viral polyprotein was processed at a site between the upstream transframe peptide (TF) and PR. Second, PR was placed after RT-IN and located at the distal end of Pol. Processing occurred only at the created junction between TF and RT. These results indicated that cleavage after TF occurred autocatalytically but did not proceed to a second site, which needed an extraneous PR for trans-action. Therefore, arranging Pol in the order of PR-RT-IN warrants the streamline processing of the polyprotein once the autocleavage is initiated.

Amino Acid Sequence↗
PubMed1990

Nucleotide sequence of the genome and complete amino acid sequence of the polyprotein of tick-borne encephalitis virus.

The sequence of the genome of tick-borne encephalitis (TBE) virus (Far Eastern subtype, strain Sofjin) coding for structural proteins and nonstructural protein NS1 has been previously reported (A. G. Pletnev, V. F. Yamshchikov, and V. M. Blinov, 1986, FEBS Lett. 200, 317-321; Yamshchikov and Pletnev, 1988, Nucleic Acids Res. 16, 7750. Now we have cloned and sequenced the genomic RNA that encodes all nonstructural proteins. Together with our earlier sequence analyses, these data show that the TBE genome is 10,477 bases in length with a single open reading frame extending from nucleotides 127 to 10,363, encoding 3412 amino acids. The 5'- and 3'-noncoding regions have stem-loop structures. The polyprotein precursor is proteolytically cleaved, apparently by a mechanism resembling that proposed for the expression of polyproteins of the other flaviviruses, such as yellow fever and Kunjin viruses. The deduced TBE gene order is 5'-C-pre(M)M-E-NS1-NS2A-NS2B-NS3-ns4a-NS4B -NS5-3'. The genome structure and the polyprotein of TBE virus is similar to mosquito-borne flaviviruses, although TBE virus is transmitted by ticks. Comparison of the sequence homology of polyproteins of flaviviruses suggests that TBE virus is more closely related to yellow fever virus than to other serological subgroups of flaviviruses. The hydrophobicity profile of the TBE polyprotein is similar to those of other flaviviruses. Nonstructural proteins NS2A, NS2B, ns4a, and NS4B are extremely hydrophobic, suggesting that these proteins are likely associated with cellular membranes. Proteins E, NS1, NS3, and NS5 are the most conserved and these proteins may be involved in the general activities related to viral reproduction.

Amino Acid Sequence↗
PubMed1990

Production of yellow fever virus proteins in infected cells: identification of discrete polyprotein species and analysis of cleavage kinetics using region-specific polyclonal antisera.

Flavivirus proteins are produced by translation of a single long open reading frame and a complex series of cotranslational and post-translational proteolytic cleavages. To study these processing events in yellow fever virus (YF)-infected cells, polyclonal antisera recognizing C, prM, E, NS1, NS2B, NS3, NS4B, and NS5 were generated using peptide and fusion protein immunogens. Evidence suggests that production of the structural protein precursors involves rapid cotranslational processing consistent with signalase cleavages. The synthesis of the NS1 glycoprotein involves cleavage of polyprotein precursors (t1/2 approximately 10 minutes) which probably contain portions of the NS2A gene product. Endoglycosidase F treatment or labeling in the presence of tunicamycin suggests that YF prM and NS1 each have two N-linked oligosaccharides. NS2B is produced without any identifiable precursors or associated polyprotein species. Processing of the NS3-4-5 region is complex and occurs rapidly. A series of polyproteins can be detected whose molecular weights correlate with the cleavage sites defined by available N-terminal amino acid sequence data. However, convincing precursor-product relationships between these polyproteins and the mature NS3 and NS5 proteins could not be demonstrated. In contrast, NS4B appears to be produced by cleavage of a discrete precursor believed to be NS4AB. N-terminal sequence data for the putative NS4AB product has tentatively defined the NS3-4A cleavage site. A scheme for in vivo processing of the YF polyprotein is presented and discussed.

Amino Acid Sequence↗
PubMed1992

A regulatory role for the 32K protein in proteolytic processing of cowpea mosaic virus polyproteins.

We have studied the regulation of proteolytic processing of the polyproteins encoded by cowpea mosaic virus M-RNA and B-RNA. For that purpose mutations were introduced in full-length cDNA clones of these RNAs. RNA transcripts were translated in rabbit reticulocyte lysate and the effect of mutations on the processing was analysed. These studies revealed that the 32K protein is released from the 200K B-polyprotein by an intramolecular cleavage and remains associated with the 170K protein, probably by interaction with the 58K domain of the 170K protein. In this complex the conformation of the 170K protein is such that further cleavages are very slow. This complex carries out the processing of the Gln/Met site in the M-polyprotein. The 170K protein produced by a B-RNA mutant that lacks the 32K coding region was efficiently processed into 110K, 87K, 84K, 60K, 58K and 24K cleavage products. Thus, the 32K protein regulates the B-polyprotein processing by slowing it down and, on the other hand, enhances trans cleavage of M-polyproteins at a Gln/Met site.

Base Sequence↗
PubMed1990

Expression and characterization of infectious bursal disease virus polyprotein in yeast.

Various expression vectors containing a cDNA fragment encoding all but the first five amino acids (aa) of the large polyprotein (N-VP2-VP4-VP3-C) of infectious bursal disease virus were transformed into yeasts. In both Saccharomyces cerevisiae and Schizosaccharomyces pombe, co- or post-translational processing of the unfused large polyprotein occurred, generating a stable C-terminal product (VP3) or correct size, but without any detectable N-terminal product (VP2). Furthermore, when the processing of the polyprotein was interrupted, because of an engineered in-frame site-specific insertion of 4 aa, even VP3 (as part of the unprocessed polyprotein) was undetected. VP2 was detected in S. cerevisiae only when fused to yeast pre-sequences at the N terminus, suggesting that in yeast, VP2 or the unprocessed polyprotein, in the absence of its native N terminus or proper protection of its N-terminal aa residues is susceptible to proteolytic degradation. The first 8 aa of a modified pre-sequence of the CUP1 gene product and the pre-pro sequence of MF alpha 1 gene product have been used for stable intra- and extra-cellular production of VP2, respectively.

Amino Acid Sequence↗
PubMed1993

Role of protein conformation in the processing of dengue virus type 2 nonstructural polyprotein precursor.

The dengue virus type-2 (DEN-2) genome is a positive-strand RNA encoding a single polyprotein precursor, C-prM(M)-E-NS1-NS2A-NS2B-NS3-NS4A-NS4B- NS5, consisting of 3391 amino acids (aa). The N-terminal region of the polyprotein precursor, C-prM(M)-E, encodes the structural proteins and is processed cotranslationally by the host signal peptidase. The nonstructural region NS1-->NS5 is processed by the viral protease(s), as well as by the signal peptidase. A two-component viral protease consisting of NS2B and the serine protease domain of NS3 has been shown to be required for cleavages having the consensus sequence of dibasic aa (K-R, R-R, R-K, or Q-R). In this study, the region encoding all the nonstructural proteins, NS1-->NS5, was expressed using a recombinant vaccinia virus system. Cleavages at the consensus viral protease recognition sites, 2B-3 at the N terminus and 3-4A at the C terminus, are prerequisites to the release of mature NS3 protease. Although the 2B-3 site was cleaved readily in a variety of polyprotein precursors containing the intact NS2B and the NS3 protease domain, the 3-4A site was most efficiently cleaved, similar to that found in DEN-2-infected cells, only in the polyprotein precursor encoding the entire nonstructural region. Removal of NS1 at the N terminus or of NS5 coding sequences at the C terminus affected the cleavage at the 3-4A site to produce the processing intermediate, NS3-NS4A. These results indicate that the conformation of the nonstructural polyprotein precursor, NS1-->NS5, plays a major role in the efficient cleavage at the 3-4A site.

Animals↗
PubMed1999

Self-processing 2A-polyproteins--a system for co-ordinate expression of multiple proteins in transgenic plants.

Achieving co-ordinate, high-level and stable expression of multiple transgenes in plants is currently difficult. Expression levels are notoriously variable and influenced by factors that act independently on transgenes at different genetic loci. Instability of expression due to loss, re-arrangement or silencing of transgenes may occur, and is exacerbated by increasing numbers of transgenic loci and repeated use of homologous sequences. Even linking two or more genes within a T-DNA does not necessarily result in co-ordinate expression. Linking proteins in a single open reading frame--a polyprotein--is a strategy for co-ordinate expression used by many viruses. After translation, polyproteins are processed into constituent polypeptides, usually by proteinases encoded within the polyprotein itself. However, in foot-and-mouth disease virus (FMDV), a sequence (2A) of just 16-20 amino acids appears to have the unique capability to mediate cleavage at its own C-terminus by an apparently enzyme-independent, novel type of reaction. This sequence can also mediate cleavage in a heterologous protein context in a range of eukaryotic expression systems. We have constructed a plasmid in which the 2A sequence is inserted between the reporter genes chloramphenicol acetyltransferase (CAT) and beta-glucuronidase (GUS), maintaining a single open reading frame. Here we report that expression of this construct in wheatgerm lysate and transgenic plants results in efficient cleavage of the polyprotein and co-ordinate expression of active CAT and GUS. Self-processing polyproteins using the FMDV 2A sequence could therefore provide a system for ensuring co-ordinated, stable expression of multiple introduced proteins in plant cells.

Amino Acid Sequence↗
PubMed2003

A Plasmodium falciparum candidate vaccine based on a six-antigen polyprotein encoded by recombinant poxviruses.

To generate broadly protective T cell responses more similar to those acquired after vaccination with radiation-attenuated Plasmodium falciparum sporozoites, we have constructed candidate subunit malaria vaccines expressing six preerythrocytic antigens linked together to produce a 3240-aa-long polyprotein (L3SEPTL). This polyprotein was expressed by a plasmid DNA vaccine vector (DNA) and by two attenuated poxvirus vectors, modified vaccinia virus Ankara (MVA) and fowlpox virus of the FP9 strain. MVAL3SEPTL boosted anti-thrombospondin-related adhesive protein (anti-TRAP) and anti-liver stage antigen 1 (anti-LSA1) CD8(+) T cell responses when primed by single antigen TRAP- or LSA1-expressing DNAs, respectively, but not by DNA-L3SEPTL. However, prime boost regimes involving two heterologous viral vectors expressing L3SEPTL induced a strong cellular response directed against an LSA1 peptide located in the C-terminal region of the polyprotein. Peptide-specific T cells secreted IFN-gamma and were cytotoxic. IFN-gamma-secreting T cells specific for each of the six antigens were induced after vaccination with L3SEPTL, supporting the use of polyprotein inserts to induce multispecific T cells against P. falciparum. The use of polyprotein constructs in nonreplicating poxviruses should broaden the target antigen range of vaccine-induced immunity and increase the number of potential epitopes available for immunogenetically diverse human populations.

Animals↗
PubMed1981

Cleavage of four avian sarcoma virus polyproteins with virion protease p15 removes gag sequences and yields large fragments that function as tyrosine phosphoacceptors in vitro.

The transformation-specific polyproteins of avian sarcoma viruses PRCII, PRCII-p, Fujinami sarcoma virus (FSV), and Esh sarcoma virus (ESV) consist of two domains, one derived from a partial viral gag gene and the other representing an apparently cell-derived insert in the defective viral genome. These gag-linked proteins were cleaved with retrovirion protease p15. Cleavage of PRCII-p polyprotein P170, P105 of PRCII, and P140 of FSV occurred within the gag domain and generated fragments of Mr 130,000, 70,000, and 115,000, respectively, containing all of the transformation-specific sequences linked to a remnant of the original gag sequences. ESV P80 was cleaved inside the transformation-specific domain, yielding a Mr 35,000--38,000 fragment from the NH2-terminal half of the molecule consisting of the entire gag portion and some no-gag sequences and a Mr 48,000 fragment containing most of the transformation-specific sequences. The tyrosine phosphorylation sites of the polyproteins were found in every case in the transformation-specific fragments. The major serine phosphorylation site of ESV P80 was found to reside in the Mr 35,000--38,000 gag-containing fragment, probably within the transformation-specific sequences of that cleavage product. Removal of all of the gag domain of ESV P80 or most of the gag domain in PRCII-p P170, PRCII P105, and FSV P140 does not affect their ability to be phosphorylated by the polyprotein-associated tyrosine-specific protein kinase activities. This observation suggests that the gag sequences of the polyproteins of classes II (PRCII-p, PRCII, and FSV) and III (ESV) avian sarcoma viruses may not be required for this enzymatic function, which appears to be of importance in transformation.

Alpharetrovirus↗
PubMed1991

Human immunodeficiency virus proteinase dimer as component of the viral polyprotein prevents particle assembly and viral infectivity.

Enzymatically active retroviral proteinases are dimers of identical polypeptide chains with a fold similar to that of other aspartic proteinases. Each polypeptide chain, encoded on one of the viral polyproteins, is less than half the size of cellular aspartic proteinases and contains only one of the two active-site aspartate residues. A plasmid was constructed to generate a genetically linked dimer of the proteinase (PR) of human immunodeficiency virus (HIV) type 1, composed of two copies of the PR sequence linked by a structurally flexible hinge region. The expression product was stable and active against HIV polyprotein substrates. Mutational analysis revealed that the linked dimer, and not multimers thereof, contained the proteolytic activity. Expression of the linked dimer as a component of a HIV polyprotein by in vitro translation gave rapid autocatalytic processing, whereas the wild-type polyprotein was stable on prolonged incubation. Transfection of HIV subviral or proviral constructs, containing the linked dimer of HIV PR, gave premature processing of the viral polyproteins, thus preventing particle formation and infectivity. Premature processing also led to increased cell toxicity.

Amino Acid Sequence↗
PubMed1992

Proteolytic processing of a Murray Valley encephalitis virus non-structural polyprotein segment containing the viral proteinase: accumulation of a NS3-4A precursor which requires mature NS3 for efficient processing.

The proteolytic processing of a non-structural polyprotein segment from the cytoplasmic domain of NS2A to the C terminus of NS5 of Murray Valley encephalitis (MVE) virus was examined, when expressed from cDNA via a vaccinia virus recombinant, in transiently transfected COS cells, or synthesized by cell-free translation. Cleavages mediated by the virus-encoded proteinase domain in NS3 at the junctions of NS2A-2B, NS2B-3 and NS4B-5 were catalysed efficiently. However, the cleavage at the NS3-4A junction, also mediated by the NS3 proteinase, was greatly delayed. Little or no NS3 was found, but an 85K precursor molecule accumulated; this was identified as NS3-4A. Termination codons were introduced by site-directed mutagenesis at the junctions of the NS3-4A, NS4A-4B and NS4B-5 genes to generate C-terminal truncations of the MVE virus polyprotein segment. In expression studies of these constructs the predicted NS3-mediated proteolytic cleavages were catalysed, except for that at the NS3-4A junction. In co-infections and co-transfections with constructs encoding the MVE virus nonstructural polyprotein region truncated at the C termini of NS3 or NS4A, efficient processing at the NS3-4A site was induced. Thus it appears that the MVE virus polyprotein is cleaved inefficiently in cis at the NS3-4A junction, whereas the site is processed efficiently in trans by mature NS3. The NS3-4A precursor is also seen in flavivirus-infected cells. Its function remains to be determined, but it could play a role in the replication of flavivirus, in view of the importance of polyprotein processing in the regulation of gene expression of positive-stranded RNA viruses, the modulation of processing at the NS3-4A site by NS3 or NS3-containing precursors described in the present study and the importance of NS3 as an integral part of the viral polymerase complex.

Animals↗