scholarly journals Irreversible inactivation of ISG15 by a viral leader protease enables alternative infection detection strategies

2018 ◽  
Vol 115 (10) ◽  
pp. 2371-2376 ◽  
Author(s):  
Kirby N. Swatek ◽  
Martina Aumayr ◽  
Jonathan N. Pruneda ◽  
Linda J. Visser ◽  
Stephen Berryman ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Viral Infection ◽  
Posttranslational Modifications ◽  
Molecular Basis ◽  
Foot And Mouth Disease ◽  
Peptide Bond ◽  
Infection Detection ◽  
Mouth Disease Virus ◽  
Host Signaling ◽  
Detection Strategies

In response to viral infection, cells mount a potent inflammatory response that relies on ISG15 and ubiquitin posttranslational modifications. Many viruses use deubiquitinases and deISGylases that reverse these modifications and antagonize host signaling processes. We here reveal that the leader protease, Lbpro, from foot-and-mouth disease virus (FMDV) targets ISG15 and to a lesser extent, ubiquitin in an unprecedented manner. Unlike canonical deISGylases that hydrolyze the isopeptide linkage after the C-terminal GlyGly motif, Lbpro cleaves the peptide bond preceding the GlyGly motif. Consequently, the GlyGly dipeptide remains attached to the substrate Lys, and cleaved ISG15 is rendered incompetent for reconjugation. A crystal structure of Lbpro bound to an engineered ISG15 suicide probe revealed the molecular basis for ISG15 proteolysis. Importantly, anti-GlyGly antibodies, developed for ubiquitin proteomics, are able to detect Lbpro cleavage products during viral infection. This opens avenues for infection detection of FMDV based on an immutable, host-derived epitope.

scholarly journals Susceptibility to viral infection is enhanced by stable expression of 3A or 3AB proteins from foot-and-mouth disease virus

Virology ◽  
2008 ◽  
Vol 380 (1) ◽  
pp. 34-45 ◽  
Author(s):  
María F. Rosas ◽  
Yuri A. Vieira ◽  
Raúl Postigo ◽  
Miguel A. Martín-Acebes ◽  
Rosario Armas-Portela ◽  
...  
Keyword(s):  
Viral Infection ◽  
Disease Virus ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Stable Expression ◽  
Mouth Disease Virus ◽  
Foot And Mouth

scholarly journals Crystal structure of the 3C protease from Southern African Territories type 2 foot-and-mouth disease virus

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e1964 ◽  
Author(s):  
Jingjie Yang ◽  
Eoin N. Leen ◽  
Francois F. Maree ◽  
Stephen Curry
Keyword(s):  
Crystal Structure ◽  
Disease Virus ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Proteolytic Processing ◽  
3C Protease ◽  
Fmdv Serotypes ◽  
Capsid Sequence ◽  
Mouth Disease Virus ◽  
Foot And Mouth

The replication of foot-and-mouth disease virus (FMDV) is dependent on the virus-encoded 3C protease (3Cpro). As in other picornaviruses, 3Cproperforms most of the proteolytic processing of the polyprotein expressed from the large open reading frame in the RNA genome of the virus. Previous work revealed that the 3Cprofrom serotype A—one of the seven serotypes of FMDV—adopts a trypsin-like fold. On the basis of capsid sequence comparisons the FMDV serotypes are grouped into two phylogenetic clusters, with O, A, C, and Asia 1 in one, and the three Southern African Territories serotypes, (SAT-1, SAT-2 and SAT-3) in another, a grouping pattern that is broadly, but not rigidly, reflected in 3Cproamino acid sequences. We report here the cloning, expression and purification of 3C proteases from four SAT serotype viruses (SAT2/GHA/8/91, SAT1/NIG/5/81, SAT1/UGA/1/97, and SAT2/ZIM/7/83) and the crystal structure at 3.2 Å resolution of 3Cprofrom SAT2/GHA/8/91.

scholarly journals Expression of heterologous genes in oncolytic adenoviruses using picornaviral 2A sequences that trigger ribosome skipping

2008 ◽  
Vol 89 (2) ◽  
pp. 389-396 ◽  
Author(s):  
Garth M. Funston ◽  
Susanna E. Kallioinen ◽  
Pablo de Felipe ◽  
Martin D. Ryan ◽  
Richard D. Iggo
Keyword(s):  
Fluorescent Protein ◽  
Burst Size ◽  
Foot And Mouth Disease ◽  
Human Adenovirus ◽  
Peptide Bond ◽  
Fusion Partner ◽  
Reading Frame ◽  
Mouth Disease Virus ◽  
Green Fluorescent ◽  
Protein Ix

Insertion of picornaviral 2A sequences into mRNAs causes ribosomes to skip formation of a peptide bond at the junction of the 2A and downstream sequences, leading to the production of two proteins from a single open reading frame. Adenoviral protein IX is a minor capsid protein that has been used to display foreign peptides on the surface of the capsid. We have used 2A sequences from the foot-and-mouth disease virus (FMDV) and porcine teschovirus 1 (PTV-1) to express protein IX (pIX) and green fluorescent protein (GFP) from pIX–2A–GFP fusion genes in an oncolytic virus derived from human adenovirus 5. GFP was efficiently expressed by constructs containing either 2A sequence. Peptide bond skipping was more efficient with the 58 aa FMDV sequence than with the 22 aa PTV-1 2A sequence, but the virus with the FMDV 2A sequence showed a reduction in plaque size, cytopathic effect, viral burst size and capsid stability. We conclude that ribosome skipping induced by 2A sequences is an effective strategy to express heterologous genes in adenoviruses; however, careful selection or optimization of the 2A sequence may be required if protein IX is used as the fusion partner.

scholarly journals Crystal Structure of Foot-and-Mouth Disease Virus 3C Protease

2005 ◽  
Vol 280 (12) ◽  
pp. 11520-11527 ◽  
Author(s):  
James R. Birtley ◽  
Stephen R. Knox ◽  
Agnès M. Jaulent ◽  
Peter Brick ◽  
Robin J. Leatherbarrow ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Disease Virus ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
3C Protease ◽  
Mouth Disease Virus ◽  
Foot And Mouth

scholarly journals Analysis of the aphthovirus 2A/2B polyprotein ‘cleavage’ mechanism indicates not a proteolytic reaction, but a novel translational effect: a putative ribosomal ‘skip’

2001 ◽  
Vol 82 (5) ◽  
pp. 1013-1025 ◽  
Author(s):  
Michelle L. L. Donnelly ◽  
Garry Luke ◽  
Amit Mehrotra ◽  
Xuejun Li ◽  
Lorraine E. Hughes ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Foot And Mouth Disease ◽  
Peptide Bond ◽  
Open Reading Frame ◽  
Translation Product ◽  
Reading Frame ◽  
C Terminus ◽  
Mouth Disease Virus ◽  
Green Fluorescent

The 2A region of the aphthovirus foot-and-mouth disease virus (FMDV) polyprotein is only 18 aa long. A ‘primary’ intramolecular polyprotein processing event mediated by 2A occurs at its own C terminus. FMDV 2A activity was studied in artificial polyproteins in which sequences encoding reporter proteins flanked the 2A sequence such that a single, long, open reading frame was created. The self-processing properties of these artificial polyproteins were investigated and the co-translational ‘cleavage’ products quantified. The processing products from our artificial polyprotein systems showed a molar excess of ‘cleavage’ product N-terminal of 2A over the product C-terminal of 2A. A series of experiments was performed to characterize our in vitro translation systems. These experiments eliminated the translational or transcriptional properties of the in vitro systems as an explanation for this imbalance. In addition, the processing products derived from a control construct encoding the P1P2 region of the human rhinovirus polyprotein, known to be proteolytically processed, were quantified and found to be equimolar. Translation of a construct encoding green fluorescent protein (GFP), FMDV 2A and β-glucuronidase, also in a single open reading frame, in the presence of puromycin, showed this antibiotic to be preferentially incorporated into the [GFP2A] translation product. We conclude that the discrete translation products from our artificial polyproteins are not produced by proteolysis. We propose that the FMDV 2A sequence, rather than representing a proteolytic element, modifies the activity of the ribosome to promote hydrolysis of the peptidyl(2A)-tRNAGly ester linkage, thereby releasing the polypeptide from the translational complex, in a manner that allows the synthesis of a discrete downstream translation product to proceed. This process produces a ribosomal ‘skip’ from one codon to the next without the formation of a peptide bond.

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