Viperin (virus inhibitory protein, endoplasmic reticulum-associated, interferon-inducible) has been shown to bind an iron-sulfur cluster with the canonical Radical SAM Superfamily C[X]3C[X]2C motif. Further, it has been shown to bind the SAM moiety and incubation of reduced viperin with SAM results in reductive cleavage of SAM to produce 5′-deoxyadenosine (5′-dAdo), a reaction characteristic of the radical SAM superfamily. The exact function of this family of proteins has yet to be determined, although recent work has provided insight into a possible mode of action for viperin via the demonstration of its interaction with farnesyl diphosphate synthase (FPPS), an enzyme that is essential for isoprenoid biosynthesis, including squalenes and sterols. Further evidence has been provided that the intracellular interaction of viperin with FPPS decreases the activity of FPPS, ultimately disrupting the formation of lipid rafts and thereby increasing the lateral mobility of the plasma membrane. Finally, it has been shown that the iron suplfur cluster is important in conformation stability of the viperin proteins.
Duschene KS, Broderick JB
The antiviral protein viperin is a radical SAM enzyme
▸ Abstract
Viperin, an interferon-inducible antiviral protein, is shown to bind an iron-sulfur cluster, based on iron analysis as well as UV-Vis and electron paramagnetic resonance spectroscopic data. The reduced protein contains a [4Fe-4S](1+) cluster whose g-values are altered upon addition of S-adenosylmethionine (SAM), consistent with SAM coordination to the cluster. Incubation of reduced viperin with SAM results in reductive cleavage of SAM to produce 5'-deoxyadenosine (5'-dAdo), a reaction characteristic of the radical SAM superfamily. The 5'-dAdo cleavage product was identified by a combination of HPLC and mass spectrometry analysis.
FEBS Lett.
2012;584(6):1263-1267
| PubMed ID:
20176015
Haldar S, Paul S, Joshi N, Dasgupta A, Chattopadhyay K
The presence of the iron-sulfur motif is important for the conformational stability of the antiviral protein, Viperin.
▸ Abstract
Viperin, an antiviral protein, has been shown to contain a CX(3)CX(2)C motif, which is conserved in the radical S-adenosyl-methionine (SAM) enzyme family. A triple mutant which replaces these three cysteines with alanines has been shown to have severe deficiency in antiviral activity. Since the crystal structure of Viperin is not available, we have used a combination of computational methods including multi-template homology modeling and molecular dynamics simulation to develop a low-resolution predicted structure. The results show that Viperin is an α-β protein containing iron-sulfur cluster at the center pocket. The calculations suggest that the removal of iron-sulfur cluster would lead to collapse of the protein tertiary structure. To verify these predictions, we have prepared, expressed and purified four mutant proteins. In three mutants individual cysteine residues were replaced by alanine residues while in the fourth all the cysteines were replaced by alanines. Conformational analyses using circular dichroism and steady state fluorescence spectroscopy indicate that the mutant proteins are partially unfolded, conformationally unstable and aggregation prone. The lack of conformational stability of the mutant proteins may have direct relevance to the absence of their antiviral activity.
PLoS One
2012;7(2):None-None
| PubMed ID:
22363738
Grewal TS, Genever PG, Brabbs AC, Birch M, Skerry TM
Best5: a novel interferon-inducible gene expressed during bone formation
▸ Abstract
Regulation of bone formation is important in the pathogenesis of many conditions such as osteoporosis, fracture healing, and loosening of orthopedic implants. We have recently identified a novel rat cDNA (best5) by differential display PCR that is regulated during osteoblast differentiation and bone formation in vitro and in vivo. Expression of best5 mRNA is induced in cultures of osteoblasts by both interferon-alpha (IFN-alpha) or IFN-gamma. Whereas IFN-alpha induced a rapid, transient induction of best5 expression peaking at 4-6 h poststimulation, IFN-gamma elicited a more prolonged induction of best5 expression, which remained elevated 48 h poststimulation. A polyclonal antibody generated to a peptide derived from the best5 coding region recognized a 27 kDa protein on Western blot analysis of osteoblast lysates. We localized BEST5 protein in osteoblast progenitor cells and mature osteoblasts in sections of rat tibiae and in sections of bones loaded in vivo to induce adaptive bone formation. Best5 may therefore be a fundamental intermediate in the response of osteoblasts to stimuli that modulate proliferation/differentiation, such as interferons or mechanical loading. These findings highlight the close interactions between the immune system and bone cells and may open new therapeutic avenues in modulating bone mass.
FASEB J
2000;14(3):523-531
| PubMed ID:
10698968
Boudinot P, Massin P, Blanco M, Riffault S, Benmansour A
vig-1, a new fish gene induced by the rhabdovirus glycoprotein, has a virus-induced homologue in humans and shares conserved motifs with the MoaA family
▸ Abstract
We used mRNA differential display methodology to analyze the shift of transcription profile induced by the fish rhabdovirus, viral hemorrhagic septicemia virus (VHSV), in rainbow trout leukocytes. We identified and characterized a new gene which is directly induced by VHSV. This VHSV-induced gene (vig-1) encodes a 348-amino-acid protein. vig-1 is highly expressed during the experimental disease in lymphoid organs of the infected fish. Intramuscular injection of a plasmid vector expressing the viral glycoprotein results in vig-1 expression, showing that the external virus protein is sufficient for the induction. vig-1 expression is also obtained by a rainbow trout interferon-like factor, indicating that vig-1 can be induced through different pathways. Moreover, vig-1 is homologous to a recently described human cytomegalovirus-induced gene. Accordingly, vig-1 activation may represent a new virus-induced activation pathway highly conserved in vertebrates. The deduced amino acid sequence of vig-1 is significantly related to sequences required for the biosynthesis of metal cofactors. This suggests that the function of vig-1 may be involved in the nonspecific virus-induced synthesis of enzymatic cofactors of the nitric oxide pathway.
J Virol
1999;73(3):1846-1852
| PubMed ID:
9971762
Helbig KJ, Beard MR
The role of viperin in the innate antiviral response
▸ Abstract
Viral infection of the cell is able to initiate a signaling cascade of events that ultimately attempts to limit viral replication and prevent escalating infection through expression of host antiviral proteins. Recent work has highlighted the importance of the host antiviral protein viperin in this process, with its ability to limit a large variety of viral infections as well as play a role in the production of type I interferon and the modulation of a number of transcription factor binding sites. Viperin appears to have the ability to modulate varying conditions within the cell and to interfere with proviral host proteins in its attempts to create an unfavorable environment for viral replication. The study of the mechanistic actions of viperin has come a long way in recent years, describing important functional domains of the protein for its antiviral and immune modulator actions as well as demonstrating its role as a member of the radical SAM enzyme family. However, despite the rapid expansion of knowledge regarding the functions of this highly conserved and ancient antiviral protein, there still remains large gaps in our understanding of the precise mechanisms at play for viperin to exert such a wide variety of roles within the cell.
J Mol Biol
2014;426(6):1210-1219
| PubMed ID:
24157441
Makins C, Ghosh S, Román-Meléndez GD, Malec PA, Kennedy RT, Marsh EN
Does Viperin Function as a Radical S-adenosyl-L-methionine-dependent Enzyme in Regulating Farnesylpyrophosphate Synthase Expression and Activity?
▸ Abstract
Viperin is an endoplasmic reticulum-associated antiviral responsive protein, which is highly up regulated in eukaryotic cells upon viral infection through both interferon-dependent and independent pathways. Viperin is predicted to be a radical S-adenosyl-L-methionine (SAM) enzyme, but it is unknown if viperin actually exploits radical SAM chemistry to exert its antiviral activity. We have investigated the interaction of viperin with its most firmly established cellular target, farnesyl pyrophosphate synthase (FPPS). Numerous enveloped viruses utilize cholesterol-rich lipid rafts to bud from the host cell membrane and it is thought that by inhibiting FPPS activity (and therefore cholesterol synthesis), viperin retards viral budding from infected cells. We demonstrate that, consistent with this hypothesis, over-expression of viperin in human embryonic kidney cells reduces the intracellular rate of accumulation of FPPS, but does not inhibit or inactivate FPPS. The endoplasmic reticulum-localizing, N-terminal amphipathic helix of viperin is specifically required for viperin to reduce cellular FPPS levels. However, although viperin reductively cleaves SAM to form 5-deoxyadenosine in a slow, uncoupled reaction characteristic of radical SAM enzymes, this cleavage reaction is independent of FPPS. Furthermore, mutation of key cysteinyl residues ligating the catalytic [Fe4S4] cluster in the radical SAM domain, surprisingly, does not abolish the inhibitory activity of viperin against FPPS; indeed some mutations potentiate viperin activity. These observations imply that viperin does not act as a radical SAM enzyme in regulating FPPS.
J Biol Chem
2016;None(None):None-None
| PubMed ID:
27834682
Viperin, an interferon-inducible antiviral protein, is shown to bind an iron-sulfur cluster, based on iron analysis as well as UV-Vis and electron paramagnetic resonance spectroscopic data. The reduced protein contains a [4Fe-4S](1+) cluster whose g-values are altered upon addition of S-adenosylmethionine (SAM), consistent with SAM coordination to the cluster. Incubation of reduced viperin with SAM results in reductive cleavage of SAM to produce 5'-deoxyadenosine (5'-dAdo), a reaction characteristic of the radical SAM superfamily. The 5'-dAdo cleavage product was identified by a combination of HPLC and mass spectrometry analysis.
