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Information on EC 3.4.21.91 - Flavivirin and Organism(s) Zika virus and UniProt Accession A0A024B7W1
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3.4.21.91 FlavivirinSpecify your search results
Word Map
- 3.4.21.91
- polyprotein
- viruses
- ns4a
-
replicons
-
west
- ribavirin
-
anti-hcv
-
direct-acting
-
virologic
-
cirrhosis
-
flaviviral
- boceprevir
- flaviviruses
- encephalitis
-
pegylated
- flaviviridae
- telaprevir
-
subgenomic
-
rna-dependent
-
unwind
-
mosquito-borne
- asunaprevir
-
resistance-associated
-
daclatasvir
- simeprevir
-
hcv-infected
- coronaviruses
-
anti-dengue
- ntpase
-
peptidomimetic
-
nonnucleoside
-
hcv-specific
-
macrodomains
- drug development
-
treatment-naive
-
peginterferon
-
replicase
-
quasispecies
-
interferon-free
-
trans-cleavage
- alphavirus
-
positive-stranded
- sofosbuvir
- analysis
- molecular biology
-
rna-stimulated
- medicine
-
hcv-1b
- synthesis
- gt1
The taxonomic range for the selected organisms is: Zika virus
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Selective hydrolysis of -Xaa-Xaa-/-Yaa- bonds in which each of the Xaa can be either Arg or Lys and Yaa can be either Ser or Ala
Synonyms
ns3 protease, nonstructural protein 3, ns2b-ns3 protease, ns3 serine protease, ns2b-ns3pro, ns2b/ns3 protease, ns3 proteinase, ns3pro, ns2b/ns3, zikv protease, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
NS2B-3 proteinase
-
-
-
-
NS2B-NS3 protease
Yellow fever virus (flavivirus) protease
-
-
-
-
CAS REGISTRY NUMBER
COMMENTARY
154215-26-6
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
Ac-D-Arg-Lys-Orn-Arg(Me)-7-amido-4-carbamoylmethylcoumarin + H2O
?
-
-
-
?
Ac-D-Arg-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin + H2O
?
substrate profiling of the P1 position using Ac-Ala-Arg-Leu-X-7-amido-4-carbamoylmethylcoumarin (X is natural or unnatural amino acid)
-
-
?
human neural transcription factor Sox2 + H2O
?
the substrate protein exhibits likely cleavage sites for NS2B-NS3pro: K95RLR-A99 and R110PRRK-T115
-
-
?
Pyr-Arg-Thr-Lys-Arg-7-amido-4-methylcoumarin + H2O
Pyr-Arg-Thr-Lys-Arg + 7-amino-4-methylcoumarin
-
-
-
?
benzoyl-Nle-Lys-Arg-Arg-7-amido-4-methylcoumarin + H2O
benzoyl-Nle-Lys-Arg-Arg + 7-amino-4-methylcoumarin
benzoyl-norleucyl-lysyl-lysyl-argininyl-7-amido-4-methylcoumarin + H2O
benzoyl-norleucyl-lysyl-lysyl-arginine + 7-amino-4-methylcoumarin
-
-
-
?
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide + H2O
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-L-arginine + 5-amino-2-nitrobenzamide
substrate binding structure in the enzyme's active site (PDB ID 5LC0), modeling. The peptidic substrate with the sequence ABZ-Val-Lys-Lys-Arg-ANB-NH2 is selected as the most efficient peptidic substrate with the nonprimed binding sites of ZIKV NS3 protease
-
-
ir
N-alpha-benzoyl-L-arginine-p-nitroanilide + H2O
N-alpha-benzoyl-L-arginine + p-nitroaniline
-
-
-
-
?
Pyr-Arg-Thr-Lys-Arg-7-amido-4-methylcoumarin + H2O
Pyr-Arg-Thr-Lys-Arg + 7-amino-4-methylcoumarin
-
-
-
?
t-butyloxycarbonyl-glycyl-L-arginyl-L-arginine-4-methylcoumaryl-7-amide + H2O
t-butyloxycarbonyl-glycyl-L-arginyl-L-arginine + 7-amino-4-methylcoumarin
-
-
-
-
?
t-butyloxycarbonyl-glycyl-L-lysyl-L-arginine-4-methylcoumaryl-7-amide + H2O
t-butyloxycarbonyl-glycyl-L-lysyl-L-arginine + 7-amino-4-methylcoumarin
-
-
-
-
?
benzoyl-Nle-Lys-Arg-Arg-7-amido-4-methylcoumarin + H2O
benzoyl-Nle-Lys-Arg-Arg + 7-amino-4-methylcoumarin
-
-
-
-
?
benzoyl-Nle-Lys-Arg-Arg-7-amido-4-methylcoumarin + H2O
benzoyl-Nle-Lys-Arg-Arg + 7-amino-4-methylcoumarin
-
-
-
?
eIF4G1 protein + H2O
?
-
human eukaryotic translation initiation factor 4 gamma 1, high activity
-
-
?
eIF4G1 protein + H2O
?
-
human eukaryotic translation initiation factor 4 gamma 1
-
-
?
additional information
?
-
analysis of substrate specificity at substrate positions P1, P2, P3, and P4, and cleavage site parameters, detailed overview. Enzyme cleavage sequence is Ac-Ala-Arg-Leu-X-7-amido-4-carbamoylmethylcoumarin (X is a natural or nonnatural amino acid). The P4-P1 optimal sequence is D-Arg-Lys-Orn-Arg. Most preferred amino acid residues at P1 are L-Arg(Me) and L-Arg. At P2, the NS2B-NS3 protease displays also a very high level of specificity. The most preferred amino acids are the non-proteinogenic L-Orn (100%) and L-Arg(Z)2 (40%). Natural amino acids interact significantly weaker with the S2 pocket, and only L-Lys and L-Arg are recognized (38% and 17%, respectively). Similarly to the S2 and S1 pockets, the S3 site exhibits very narrow specificity. L-Lys is the most preferentially recognized residue at P3 among all tested amino acids. Other amino acids such as L-Orn, L-Arg, L-Arg(Z)2, L-hArg, L-Agp or L-Phe(guan) are recognized with significantly lower affinity. However, all these amino acids have basic side chains. The S4 site exhibits very broad substrate specificity. Almost all amino acids, including unnatural ones, are recognized at the same level at P4 position
-
-
?
additional information
?
-
-
analysis of substrate specificity at substrate positions P1, P2, P3, and P4, and cleavage site parameters, detailed overview. Enzyme cleavage sequence is Ac-Ala-Arg-Leu-X-7-amido-4-carbamoylmethylcoumarin (X is a natural or nonnatural amino acid). The P4-P1 optimal sequence is D-Arg-Lys-Orn-Arg. Most preferred amino acid residues at P1 are L-Arg(Me) and L-Arg. At P2, the NS2B-NS3 protease displays also a very high level of specificity. The most preferred amino acids are the non-proteinogenic L-Orn (100%) and L-Arg(Z)2 (40%). Natural amino acids interact significantly weaker with the S2 pocket, and only L-Lys and L-Arg are recognized (38% and 17%, respectively). Similarly to the S2 and S1 pockets, the S3 site exhibits very narrow specificity. L-Lys is the most preferentially recognized residue at P3 among all tested amino acids. Other amino acids such as L-Orn, L-Arg, L-Arg(Z)2, L-hArg, L-Agp or L-Phe(guan) are recognized with significantly lower affinity. However, all these amino acids have basic side chains. The S4 site exhibits very broad substrate specificity. Almost all amino acids, including unnatural ones, are recognized at the same level at P4 position
-
-
?
additional information
?
-
no activity with human maltose binding protein
-
-
?
additional information
?
-
-
identification of 31 human proteins that are cleaved by the NS2B-NS3 protease. The cofactor NS2B is the principal determinant in ZVP substrate selection
-
-
?
additional information
?
-
screening for potential intracellular substrates of NS3, substrate specificity analysis of recombinant enzyme NS2BLNNS3pr using synthesized FRET-type substrate libraries, peptide library preparation. Profiling of the P1-P4 substrate specificity of the NS2B-NS3 protease using a combinatorial chemistry approach and the split-and-mix method. Arg is the only preferred P1 residue of the enzyme, preference for Val at P4, and Lys at P3 and P2, no activity for Trp, Tyr, Asp, and Pro at P3 position, structure-function analysis, overview
-
-
?
additional information
?
-
-
screening for potential intracellular substrates of NS3, substrate specificity analysis of recombinant enzyme NS2BLNNS3pr using synthesized FRET-type substrate libraries, peptide library preparation. Profiling of the P1-P4 substrate specificity of the NS2B-NS3 protease using a combinatorial chemistry approach and the split-and-mix method. Arg is the only preferred P1 residue of the enzyme, preference for Val at P4, and Lys at P3 and P2, no activity for Trp, Tyr, Asp, and Pro at P3 position, structure-function analysis, overview
-
-
?
additional information
?
-
the enzyme performs self-cleavage processes viral polyproteins. Protease activity analysis of gZiPro, eZiPro and bZiPro
-
-
?
additional information
?
-
-
the enzyme performs self-cleavage processes viral polyproteins. Protease activity analysis of gZiPro, eZiPro and bZiPro
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
eIF4G1 protein + H2O
?
-
human eukaryotic translation initiation factor 4 gamma 1, high activity
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
NS2B cofactor
the NS2B cofactor is covalently linked to the NS3 protease domain via a Gly-Gly-Gly-Gly-Ser-Gly-Gly-Gly-Gly (G4SG4) linker
-
NS2B cofactor
-
the unique cofactor region of zika virus NS2B-NS3 protease facilitates cleavage of key host proteins. The NS2B cofactor region from Zika virus protease is essential for recognition of host cell substrates. Replacing the NS2B region in another flavivirus protease enabled recognition of novel Zika-specific substrates by hybrid proteases, suggesting that the cofactor is the principal determinant in ZVP substrate selection
-
NS2B cofactor
-
the viral protease is a two-component serine protease formed by the N-terminal part of NS3 and the cofactor region of NS2B, a membrane protein essential for the membrane location of NS3. The NS2B mutant C143S cannot form the disulfide bond between C143 residues of neighboring NS3
-
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
5,6-dichloro-2H-triazolo[4,5-b]pyrazine
NSC157058, the inhibitor decreases ZIKV infection in mice
6-(4-chlorophenyl)-3-[[3-[[6-(4-chlorophenyl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-3-yl]methoxy]phenoxy]methyl]-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole
NSC716903
Aprotinin
a 60-amino acid bovine pancreatic trypsin inhibitor and an efficient inhibitor of ZIKV NS2B-NS3pro
(S)-2-acetamido-6-amino-N-((S)-5-guanidino-1-oxopentan-2-yl)hexanamide
-
inhibitor-enzyme interaction analysis, crystal structure analysis of the complex, NMR spectrometry, overview
1-(4-[3-[4-(furan-3-yl)phenyl]-5-(piperidin-4-ylmethoxy)pyrazin-2-yl]phenyl)methanamine
-
-
4-(aminomethyl)-Nalpha-benzoyl-N-[(1R)-4-carbamimidamido-1-(dihydroxyboranyl)butyl]-L-phenylalaninamide
cn-716, ZIKV protease in complex with a peptidomimetic boronic acid inhibitor reveals a cyclic diester between the boronic acid and glycerol, enzyme binding structure, binds to the active site, crystal structure analysis, detailed overview
5-amino-1-((4-methoxyphenyl)sulfonyl)-1H-pyrazol-3-yl benzoate
-
the benzoyl moiety of the inhibitor forms a covalent bond with the side chain of S135. Structure and dynamics of bZiPro-inhibitor complex in solution
acetyl-K-agmatine
-
acetyl-K-agmatine is weakly bound by the ZIKV protease and does inhibit the enzyme. The Arg and Lys residues of acyl-KR-COOH occupy the S1 and S2 sites of the protease, respectively
N2-(thiophen-2-ylcarbonyl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide
the compound inhibits both unlinked and linked protease with similar potency
additional information
identification of structural scaffolds for allosteric small-molecule inhibitors of this protease, overview. Molecular modeling of the protease-inhibitor complexes suggests that these compounds bind to the druggable cavity in the NS2B-NS3 protease interface and affect productive interactions of the protease domain with its cofactor. The most potent compound demonstrate efficient inhibition of ZIKV propagation in vitro in human fetal neural progenitor cells and in vivo in SJL mice. Docking study and in silico modeling of ZIKV NS2B-NS3pro complexed with inhibitors
-
additional information
-
identification of structural scaffolds for allosteric small-molecule inhibitors of this protease, overview. Molecular modeling of the protease-inhibitor complexes suggests that these compounds bind to the druggable cavity in the NS2B-NS3 protease interface and affect productive interactions of the protease domain with its cofactor. The most potent compound demonstrate efficient inhibition of ZIKV propagation in vitro in human fetal neural progenitor cells and in vivo in SJL mice. Docking study and in silico modeling of ZIKV NS2B-NS3pro complexed with inhibitors
-
additional information
-
N-alpha-benzoyl-L-arginine-p-nitroanilide causes precipitation of the recombinant protease and is therefore not appropiate for enzymatic analysis
-
additional information
-
discovery, X-ray crystallography and antiviral activity of allosteric inhibitors of flavivirus NS2B-NS3 protease. Compound screening followed by medicinal chemistry yield a series of drug-like, broadly active inhibitors of flavivirus proteases with IC50 as low as 120 nM. The inhibitors exhibit significant antiviral activities in cells (EC68: 300-600 nM) and in a mouse model of Zika virus infection. X-ray studies reveal that the inhibitors bind to an allosteric, mostly hydrophobic pocket of dengue NS3 and hold the protease in an open, catalytically inactive conformation
-
additional information
screening for potential intracellular substrates of NS3 and development of specific inhibitors of ZIKV protease, overview
-
additional information
-
screening for potential intracellular substrates of NS3 and development of specific inhibitors of ZIKV protease, overview
-
additional information
-
structural dynamics of Zika virus NS2B-NS3 protease binding to dipeptide inhibitors, overview. The unlinked protease forms the closed conformation and the aldehyde group from the inhibitor forms a covalent bond with the side chain of S135. Compounds derived from protease substrate bind to the protease active site. Nuclear magnetic resonance studies demonstrate that the protease-inhibitor complex is in the closed conformation in solution. No inhibition by acyl-KR-COOH
-
additional information
-
structural insights into the inhibition of Zika virus NS2B-NS3 protease by a small-molecule inhibitor, structure and dynamics, overview. 5-amino-1-((4-methoxyphenyl)sulfonyl)-1H-pyrazol-3-yl benzoate stabilizes the closed conformation of ZIKV protease. Upon hydrolysis in situ into two fragments, the benzoyl group of the inhibitor forms a covalent bond with the side chain of catalytic residue S135, whereas the second fragment exhibits no obvious molecular interactions with the protease, detailed mechanism of action of a covalent inhibitor. Unique adduct of the benzoyl moiety to residue S135
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.00418
Ac-Arg-Lys-Lys-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
0.006
Ac-D-Arg-Lys-Orn-Arg(Me)-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
0.0042
Ac-D-Arg-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
0.00799
Ac-D-Lys-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
0.01385
Ac-Nle-Lys-Lys-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
0.111 - 0.444
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
0.111
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 7.4, 37°C, phosphate buffer
0.444
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 8.5, 37°C, Tris-CHAPS-glycerol buffer
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
5.43
Ac-D-Arg-Lys-Orn-Arg(Me)-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
4.64
Ac-D-Arg-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
7.99
Ac-D-Lys-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
25.94 - 343.73
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
25.94
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 7.4, 37°C, phosphate buffer
343.73
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 8.5, 37°C, Tris-CHAPS-glycerol buffer
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
258.37
Ac-Arg-Lys-Lys-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
905
Ac-D-Arg-Lys-Orn-Arg(Me)-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
1104.76
Ac-D-Arg-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
1000
Ac-D-Lys-Lys-Orn-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
317.69
Ac-Nle-Lys-Lys-Arg-7-amido-4-carbamoylmethylcoumarin
pH 8.5, 22°C
235 - 7760
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
235
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 7.4, 37°C, phosphate buffer
7760
N-(2-aminobenzoyl)-L-valyl-L-lysyl-L-lysyl-N-(3-carbamoyl-4-nitrophenyl)-L-argininamide
pH 8.5, 37°C, Tris-CHAPS-glycerol buffer
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.000208
(S)-2-acetamido-6-amino-N-((S)-5-guanidino-1-oxopentan-2-yl)hexanamide
Zika virus
-
pH and temperature not specified in the publication
0.0015
5-amino-1-((4-methoxyphenyl)sulfonyl)-1H-pyrazol-3-yl benzoate
Zika virus
-
20xa0mM Tris pH 8.5, 10% glycerol, 0.01% Triton X-100, 37°C
0.00104 - 0.00106
N2-(thiophen-2-ylcarbonyl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide
0.00104
N2-(thiophen-2-ylcarbonyl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide
Zika virus
recombinant linked protease, pH 8.5, 37°C
0.00106
N2-(thiophen-2-ylcarbonyl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide
Zika virus
recombinant unlinked protease, pH 8.5, 37°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
8.5
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 10.5
-
at pH 9.0 the enzyme is 5times more active than at pH 7.0, and more than 3times more active than at pH 10.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
37
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
NS2B-NS3 protease consists of the NS2B cofactor and the NS3 protease domain and is essential for cleavage of the ZIKV polyprotein precursor and generation of fully functional viral proteins. Cell toxicity assays. The enzyme cleaves human Sox2 protein, which plays a critical role in neural stem cells by maintaining the activity of multiple genes involved in self-renewal and by priming the epigenetic landscape for the onset of neuronal differentiation. Sox2 is cleaved by ZIKV NS2B-NS3pro only at a very high enzyme/substrate ratio Sox2 insufficiency results in a plethora of developmental neuronal malformations in the human brain
evolution
physiological function
additional information
evolution
comparative analysis of the substrate specificity reveals that ZIKV NS3 protease follows the typical pattern for flaviviruses
evolution
-
the flavivirus NS2B-NS3 protease is highly conserved in Zika, West Nile, and Dengue viruses
physiological function
-
autophagy-related protein 16-1 (ATG16L1) and eukaryotic translation initiation factor 4 gamma 1 (eIF4G1) are dramatically depleted during Zika virus infection. ATG16L1 and eIF4G1 mediate type-II interferon production and host-cell translation, respectively, likely aiding immune system evasion and driving the Zika life cycle
physiological function
the viral protease that processes viral polyproteins
physiological function
the Zika virus (ZIKV) relies on its NS2B/NS3 protease for polyprotein processing
physiological function
the ZIKV NS2B/NS3 protease is responsible for cleavage of the viral polyprotein at five sites
physiological function
-
viral NS2B-NS3 protease processes viral polyprotein and is essential for the virus replication
physiological function
ZIKV expresses the serine protease NS3 which is responsible for viral protein processing and replication
additional information
structure-function studies of ZIKV NS2B-NS3 protease
additional information
increased activity of unlinked Zika virus NS2B/NS3 protease compared to linked Zika virus protease. Enzyme structure homology modeling and molecular dynamics simulations of linked and unlinked protease, overview
additional information
-
increased activity of unlinked Zika virus NS2B/NS3 protease compared to linked Zika virus protease. Enzyme structure homology modeling and molecular dynamics simulations of linked and unlinked protease, overview
additional information
-
solution NMR spectrum of recombinant gZiPro enzyme without inhibitor, overview. The inhibitor-bound gZiPro-AcKR-H complex adopts the closed conformation
additional information
structure of the NS2B-NS3 protease from Zika virus after self-cleavage. Analysis of the post-proteolysis state structure of the enzyme during viral polyprotein processing providing insights into peptide substrate recognition by the protease, Protease activity analysis of gZiPro, eZiPro and bZiPro
additional information
-
structure of the NS2B-NS3 protease from Zika virus after self-cleavage. Analysis of the post-proteolysis state structure of the enzyme during viral polyprotein processing providing insights into peptide substrate recognition by the protease, Protease activity analysis of gZiPro, eZiPro and bZiPro
additional information
-
the unique cofactor region of zika virus NS2B-NS3 protease facilitates cleavage of key host proteins
additional information
-
the unlinked NS2B-NS3 protease adopts a closed conformation in which NS2B engages NS3 to form an empty substrate-binding site
additional information
-
the viral protease is a two-component serine protease formed by the N-terminal part of NS3 and the cofactor region of NS2B, a membrane protein essential for the membrane location of NS3
additional information
-
the viral protease is a two-component serine protease formed by the N-terminal part of NS3 and the cofactor region of NS2B, a membrane protein essential for the membrane location of NS3
PDB
SCOP
CATH
UNIPROT
ORGANISM
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
-
Zika protease is able to form a dimer, and a disulfide bond between C143 residues of neighboring NS3 in the crystal is observed
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
proteolytic modification
structure of the NS2B-NS3 protease from Zika virus after self-cleavage
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified bZiPro (C143S)-inhibitor 5-amino-1-((4-methoxyphenyl)sulfonyl)-1H-pyrazol-3-yl benzoate complex and bZiPro (C143S)-Ac-Lys-Arg complex, X-ray diffraction structure determination and analysis at 1.90 A and 1.51 A resolution, respectively
-
purified enzyme complex with bound inhibitor (S)-2-acetamido-6-amino-N-((S)-5-guanidino-1-oxopentan-2-yl)hexanamide, X-ray diffraction structure determination and analysis at 2.4 A resolution
-
purified free NS2B-NS3 protease and complexed with a peptide reversely oriented at the active site, X-ray diffraction structure determination and analysis at 1.58 A resolution. A second protease in the same crystal binds to the residues K14K15G16E17 from the neighboring NS3 in reverse orientation, resisting proteolysis
-
purified NS2B-NS3 protease complexed with the last four amino acids of the NS2B cofactor bound at the NS3 active site, sitting drop vapour diffusion method, mixing of 0.001 ml of 40 mg/ml protein solution with 0.001 ml of reservoir solution containing 0.2 M Na-malonate, pH 4.0, 10% PEG 3350, 18°C, two days, X-ray diffraction structure determination and analysis at 1.84 A resolution
purified ZIKV protease in complex with a peptidomimetic boronic acid inhibitor, X-ray diffraction structure determination and analysis at 2.7 A resolution
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C143S
-
site-directed mutagenesis, the NS2B mutant cannot form the disulfide bond between C143 residues of neighboring NS3
R95*A/R29G
construction of a complex containing residues 45 to 95 of the NS2B cofactor linked via a Gly-Gly-Gly-Gly-Ser-Gly-Gly-Gly-Gly (G4SG4) linker to the NS3 protease to avoid auto-proteolysis and create the linked protease, the last arginine residue of the NS2B cofactor before the G4SG4 linker is mutated to alanine (R95*A), and arginine 29 of the NS3 protease domain is mutated to glycine (R29G)
additional information
additional information
construction of eZiPro that contains the NS2B cofactor region (residues 45-96) linked with NS3 protein (residues 1-177) through the last five residues of NS2B (K126TGKR130), and the gZiPro construct, which is similar except that the G4SG4 linker replaced K126TGKR130 of NS2B. Protease activity analysis of gZiPro, eZiPro and bZiPro
additional information
-
construction of eZiPro that contains the NS2B cofactor region (residues 45-96) linked with NS3 protein (residues 1-177) through the last five residues of NS2B (K126TGKR130), and the gZiPro construct, which is similar except that the G4SG4 linker replaced K126TGKR130 of NS2B. Protease activity analysis of gZiPro, eZiPro and bZiPro
additional information
-
construction of the NS2B cofactor region linked to the NS3 protease domain via a glycine-rich flexible linker (Gly4-Ser-Gly4 linker), structural dynamics of this conventional Zika protease (gZiPro) using NMR spectroscopy, overview. Although the glycine-rich linker in gZiPro does not alter the overall folding of the protease in solution, gZiPro is not homogenous in ion exchange chromatography. Compared to the unlinked protease construct, the artificial linker affects the chemical environment of many residues including H51 in the catalytic triad. Direct comparison of ZIKV protease constructs with and without an artificial linker, namely gZiPro (with the Gly4-Ser-Gly4 linker), bZiPro (unlinked NS2B NS3protease), and eZiPro (NS2B-NS3 junction sequence KTGKR as the linker). Solution NMR spectrum of recombinant gZiPro enzyme without inhibitor, overview. The inhibitor-bound gZiPro-AcKR-H complex adopts the closed conformation. Effect of the linker on the chemical environment of residues, dynamics of the linked protease, overview
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, tag cleavage via thrombin, dialysis, and gel filtration
recombinant His6-SUMO-tagged linked and unlinked NS3-NS2B(45-95) complex linked from Escherichia coli strain BL21 Codon Plus (DE3) by nickel affinity chromatography, tag cleavage by yeast SUMO Protease 1, followed by another step of nickel affinity chromatography, and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
codon optimized cDNA encoding NS2B-NS3, recombinant expression of His-tagged wild-type enzyme and constructs in Escherichia coli strain BL21(DE3)
deltaNS2B-NS3pro and deltaNS2B-NS3proHDA expressed as a hexahistidine-tagged protein in Escherichia coli BL21-CodonPlus(DE3)-RIL cells
-
recombinant expression of an active NS3pro domain fused with the NS2B cofactor (NS2BLNNS3pro) in a prokaryotic expression system
recombinant overexpression of His6-SUMO-tagged linked and unlinked NS3-NS2B(45-95) complex linked via G4SG4 linker in Escherichia coli strain BL21 Codon Plus (DE3)
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
drug development
drug development
-
the enzyme is a target for the development of structure-based antiviral drugs against ZIKV and related pathogenic flaviviruses
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Bessaud, M.; Pastorino, B.A.; Peyrefitte, C.N.; Rolland, D.; Grandadam, M.; Tolou, H.J.
Functional characterization of the NS2B/NS3 protease complex from seven viruses belonging to different groups inside the genus Flavivirus
Virus Res.
120
79-90
2006
Yellow fever virus, Zika virus, Saint Louis encephalitis virus, Bussuquara virus, Langat virus (P29837), Dengue virus type 3 (Q6YMS4), dengue virus type I (Q91NH1), Yellow fever virus Asibi, Dengue virus type 3 Sri Lanka/1266/2000 (Q6YMS4), dengue virus type I D1/H/IMTSSA/98/606 (Q91NH1), Langat virus TP21 (P29837), Zika virus Ar-D-41644, Saint Louis encephalitis virus MSI-7
Hill, M.E.; Kumar, A.; Wells, J.A.; Hobman, T.C.; Julien, O.; Hardy, J.A.
The unique cofactor region of zika virus NS2B-NS3 protease facilitates cleavage of key host proteins
ACS Chem. Biol.
13
2398-2405
2018
Zika virus
Rut, W.; Zhang, L.; Kasperkiewicz, P.; Poreba, M.; Hilgenfeld, R.; Drag, M.
Extended substrate specificity and first potent irreversible inhibitor/activity-based probe design for Zika virus NS2B-NS3 protease
Antiviral Res.
139
88-94
2017
Zika virus (A0A024B7W1), Zika virus
Shiryaev, S.A.; Farhy, C.; Pinto, A.; Huang, C.T.; Simonetti, N.; Elong Ngono, A.; Dewing, A.; Shresta, S.; Pinkerton, A.B.; Cieplak, P.; Strongin, A.Y.; Terskikh, A.V.
Characterization of the Zika virus two-component NS2B-NS3 protease and structure-assisted identification of allosteric small-molecule antagonists
Antiviral Res.
143
218-229
2017
Zika virus (A0A024B7W1), Zika virus
Kuiper, B.D.; Slater, K.; Spellmon, N.; Holcomb, J.; Medapureddy, P.; Muzzarelli, K.M.; Yang, Z.; Ovadia, R.; Amblard, F.; Kovari, I.A.; Schinazi, R.F.; Kovari, L.C.
Increased activity of unlinked Zika virus NS2B/NS3 protease compared to linked Zika virus protease
Biochem. Biophys. Res. Commun.
492
668-673
2017
Zika virus (A0A384KRS4), Zika virus, Zika virus Puerto Rico isolate (A0A384KRS4)
Gruba, N.; Rodriguez Martinez, J.I.; Grzywa, R.; Wysocka, M.; Skorenski, M.; Burmistrz, M.; Lecka, M.; Lesner, A.; Sienczyk, M.; Pyrc, K.
Substrate profiling of Zika virus NS2B-NS3 protease
FEBS Lett.
590
3459-3468
2016
Zika virus (Q32ZE1), Zika virus
Li, Y.; Phoo, W.W.; Loh, Y.R.; Zhang, Z.; Ng, E.Y.; Wang, W.; Keller, T.H.; Luo, D.; Kang, C.
Structural characterization of the linked NS2B-NS3 protease of Zika virus
FEBS Lett.
591
2338-2347
2017
Zika virus
Yao, Y.; Huo, T.; Lin, Y.L.; Nie, S.; Wu, F.; Hua, Y.; Wu, J.; Kneubehl, A.R.; Vogt, M.B.; Rico-Hesse, R.; Song, Y.
Discovery, X-ray crystallography and antiviral activity of allosteric inhibitors of flavivirus NS2B-NS3 protease
J. Am. Chem. Soc.
141
6832-6836
2019
Dengue virus, West Nile virus, Zika virus
Phoo, W.W.; Li, Y.; Zhang, Z.; Lee, M.Y.; Loh, Y.R.; Tan, Y.B.; Ng, E.Y.; Lescar, J.; Kang, C.; Luo, D.
Structure of the NS2B-NS3 protease from Zika virus after self-cleavage
Nat. Commun.
7
13410
2016
Zika virus (H8XX12), Zika virus
Lei, J.; Hansen, G.; Nitsche, C.; Klein, C.D.; Zhang, L.; Hilgenfeld, R.
Crystal structure of Zika virus NS2B-NS3 protease in complex with a boronate inhibitor
Science
353
503-505
2016
Zika virus (A0A140DLX4), Zika virus, Zika virus Brazilian isolate BeH823339 (A0A140DLX4)
Zhang, Z.; Li, Y.; Loh, Y.R.; Phoo, W.W.; Hung, A.W.; Kang, C.; Luo, D.
Crystal structure of unlinked NS2B-NS3 protease from Zika virus
Science
354
1597-1600
2016
Zika virus
Li, Y.; Zhang, Z.; Phoo, W.W.; Loh, Y.R.; Wang, W.; Liu, S.; Chen, M.W.; Hung, A.W.; Keller, T.H.; Luo, D.; Kang, C.
Structural dynamics of Zika virus NS2B-NS3 protease binding to dipeptide inhibitors
Structure
25
1242-1250
2017
Zika virus
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