Proprotein convertase furin is responsible for the processing of a wide variety of precursors consisted of signal peptide, propeptide and mature peptide in mammal. Many precursors processed by furin have important physiological functions and can be recombinantly expressed in Pichia pastoris expression system for research, pharmaceutical and vaccine applications. However, it is not clear whether the furin cleavage sites between the propeptide and mature peptide can be properly processed in P. pastoris, bringing uncertainty for proper expression of the coding DNA sequences of furin precursors containing the propeptides and mature peptides. In this study, we evaluated the ability of P. pastoris to process furin cleavage sites and how to improve the cleavage efficiencies of furin cleavage sites in P. pastoris. The results showed that P. pastoris can process furin cleavage sites but the cleavage efficiencies are not high. Arg residue at position P1 or P4 in furin cleavage sites significantly affect cleavage

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. Success of current therapies is still limited and outcome is particularly poor for metastatic alveolar rhabdomyosarcoma (aRMS). We previously identified the proprotein convertase furin as potential target for specific drug delivery with RMS-homing peptides. Furin is a protease that converts inactive precursor proteins into bioactive proteins and peptides. In this study, we investigate the biological role of furin in aRMS progression in vitro and in vivo. Furin expression was confirmed in over 86% RMS biopsies in a tissue microarray (n=89). Inducible furin silencing in vitro led to significant impairment of cell viability and proliferation in all investigated aRMS cell lines, but not in MRC5 fibroblasts. Furthermore, the aRMS cell lines Rh3 and Rh4 revealed to be very sensitive to furin silencing, undergoing caspase-dependent cell death. Notably, furin silencing in vivo led to complete remission of established Rh4 tumors ...

Toll-like receptor 7 (TLR7) triggers antiviral immune responses by recognizing viral single-stranded RNA in endosomes, but the biosynthetic pathway of human TLR7 (hTLR7) remains unclear. Here, we show that hTLR7 is proteolytically processed and that the C-terminal fragment selectively accumulates in endocytic compartments. hTLR7 processing occurred at neutral pH and was dependent on furin-like proprotein convertases (PCs). Furthermore, TLR7 processing was required for its functional response to TLR7 agonists such as R837 or influenza virus. Notably, proinflammatory and differentiation stimuli increased the expression of furin-like PCs in immune cells, suggesting a positive feedback mechanism for TLR7 processing during infection. Because self-RNA can under certain conditions activate TLR7 and trigger autoimmunity, our results identify furin-like PCs as a possible target to attenuate TLR7-dependent autoimmunity and other immune pathologies.

Matrix metalloproteinases (MMPs), a family of zinc-dependent endopeptidases implicated in tumor invasion and metastasis, must undergo zymogen activation prior to expressing any proteolytic activity. Although the cysteine-switch model predicts the well-established autoactivation process, approximately 40% of the known MMPs possess a conserved RXKR motif between their pro- and catalytic domains and, thus, could be activated directly by members of the proprotein convertase family. To further understand this process, we analyzed the activation of proMT3-MMP as a model system. We demonstrated that the conversion of MT3-MMP zymogen into active form is dependent on both the furin-type convertase activity and the R(116)RKR motif. Consistently, MT3-MMP was colocalized with furin in the trans-Golgi network by confocal microscopy. However, neither furin activity nor its recognition site in MT3-MMP is required for the observed colocalization. In fact, the colocalization pattern remains intact, even in the presence

Glioblastoma is the most common and aggressive intrinsic brain tumor in adults. Self-renewing, highly tumorigenic glioma-initiating cells (GIC) have been linked to glioma invasive properties, immunomodulation, and increased angiogenesis, leading to resistance to therapy. TGF-β signaling has been associated with the tumorigenic activity of GIC. TGF-β is synthesized as a precursor molecule and proteolytically processed to the mature form by members of the family of the proprotein convertases subtilisin/kexin. In this study we report that furin is unique among the proprotein convertases subtilisin/kexin in being highly expressed in human GIC. Furin cleaves and promotes activation of pro-TGF-β1 and pro-TGF-β2, and TGF-β2 in turn increases furin levels. Notably, TGF-β2 controls furin activity in an ALK-5-dependent manner involving the ERK/MAPK pathway. We thus uncover a role of ERK1 in the regulation of furin activity by supporting a self-sustaining loop for high TGF-β activity in GIC. ...

Furin is an evolutionarily conserved proprotein convertase (PC) family enzyme with a broad range of substrates that are essential for developmental, homeostatic, and disease pathways. Classical genetic approaches and in vitro biochemical or cell biological assays identified that precursor forms of most growth factor family proteins are processed by Furin. To quantitatively assess the potential role of Furin in cleaving and modulating intercellular dispersion of a Drosophila signaling protein, we developed a simple assay by combining genetics, ex vivo organ culture, pharmacological treatment, and imaging analyses. The protocol herein describes how to ex vivo culture Drosophila wing imaginal discs expressing a fluorescently tagged Drosophila Fibroblast Growth Factor (FGF, Branchless/Bnl) over a long period of time in the presence of Furin inhibitors and monitor the cleavage and intercellular dispersion of the truncated Bnl parts using microscopy. Although the assay described here is for assessing the

The fusogenic potential of Class I viral envelope glycoproteins is activated by proteloytic cleavage of the precursor glycoprotein to generate the mature receptor-binding and transmembrane fusion subunits. Although the coronavirus (CoV) S glycoproteins share membership in this class of envelope glycoproteins, cleavage to generate the respective S1 and S2 subunits appears absent in a subset of CoV species, including that responsible for the severe acute respiratory syndrome (SARS). To determine whether proteolytic cleavage of the S glycoprotein might be important for the newly emerged SARS-CoV, we introduced a furin recognition site at single basic residues within the putative S1-S2 junctional region. We show that furin cleavage at the modified R667 position generates discrete S1 and S2 subunits and potentiates membrane fusion activity. This effect on the cell-cell fusion activity by the S glycoprotein is not, however, reflected in the infectivity of pseudotyped lentiviruses bearing the cleaved ...

Bone morphogenetic protein 10 (BMP10) is a member of the TGF-β superfamily and plays a critical role in heart development. In the postnatal heart, BMP10 is restricted to the right atrium. The inactive pro-BMP10 (∼60 kDa) is processed into active BMP10 (∼14 kDa) by an unknown protease. Proteolytic cleavage occurs at the RIRR(316)↓ site (human), suggesting the involvement of proprotein convertase(s) (PCs). In vitro digestion of a 12-mer peptide encompassing the predicted cleavage site with furin, PACE4, PC5/6, and PC7, showed that furin cleaves the best, whereas PC7 is inactive on this peptide. Ex vivo studies in COS-1 cells, a cell line lacking PC5/6, revealed efficient processing of pro-BMP10 by endogenous PCs other than PC5/6. The lack of processing of overexpressed pro-BMP10 in the furin- and PACE4-deficient cell line, CHO-FD11, and in furin-deficient LoVo cells, was restored by stable (CHO-FD11/Fur cells) or transient (LoVo cells) expression of furin. Use of cell-permeable and cell surface

Proteolytic processing is required for the activation of numerous viral glycoproteins. Here we show that the envelope glycoprotein from the Zaire strain of Ebola virus (Ebo-GP) is proteolytically processed into two subunits, GP1 and GP2, that are likely covalently associated through a disulfide linkage. Murine leukemia virions pseudotyped with Ebo-GP contain almost exclusively processed glycoprotein, indicating that this is the mature form of Ebo-GP. Mutational analysis identified a dibasic motif, reminiscent of furin-like protease processing sites, as the Ebo-GP cleavage site. However, analysis of Ebo-GP processing in LoVo cells that lack the proprotein convertase furin demonstrated that furin is not required for processing of Ebo-GP. In sharp contrast to other viral systems, we found that an uncleaved mutant of Ebo-GP was able to mediate infection of various cell lines as efficiently as the wild-type, proteolytically cleaved glycoprotein, indicating that cleavage is not required for the ...

SARS-CoV-2 enters cells via its spike glycoprotein which must be cleaved sequentially at the S1/S2, then the S2 cleavage sites (CS) to mediate membrane fusion. SARS-CoV-2 has a unique polybasic insertion at the S1/S2 CS, which we demonstrate can be cleaved by furin.

Furin, a subtilisin/kesin-like proprotein convertase (PC), activates membrane-bound MT1-MMP, which facilitates pro-gelatinase A (MMP2). These activated MT1-MMP and MMP2 are involved in cancer cell invasion and metastasis. In this study, we investigate the contribution of MMP2 activated by furin to cellular invasiveness of cumulatively irradiated cells.. Using previously established AMC-HN3 and AMC-HN8 cell line from laryngeal carcinoma patient, we have generated isogenic model of cumulatively irradiated AMC-HN3R and AMC-HN8R cell line, respectively. AMC-HN3R cells were increased furin expression with upregulation of MT1-MMP/MMP2 and their invasiveness by two fold (p , 0.05) compared to AMC-HN3, while AMC-HN8R cells had no differences compared to AMC-HN8. In case of AMC-HN3R, inhibition of furin activity with the synthetic inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl-keton, CMK, showed a significant decrease of MT1-MMP/MMP2 and in vitro cellular invasiveness. Tumors obtained after subcutaneous ...

SDS-PAGE studies of EBO virion glycoproteins (Fig. 1 and 3A) and N-terminal sequencing have provided definitive evidence that GP is cleaved (from GP0 form) into two molecules, GP1 and GP2. Cleavage of the structural GP gene product of EBO virus was expected for the N-terminal signal sequence and occurs at the predicted site. Cleavage of GP by furin had been predicted from the RRTRR sequence, but identification of the C-terminal fragment in virions was complicated by its comigration with VP24 in SDS-PAGE. However, prior studies detected GP2 in EBO-infected tissue culture fluids (17, 21), and it was suggested that this uncharacterized glycoprotein might be derived from a cleavage event involving GP. Indirect evidence for furin cleavage of EBO virus GP was recently demonstrated through the use of specific inhibitors of furin activity (21).. When GP sequences of all known EBO viruses are aligned, one finds that the furin cleavage site is conserved but differs slightly from one EBO species to ...

Members of the bacterial Shiga toxin family consist of a single A subunit that is non-covalently associated with a pentamer of B subunits. These toxins bind to receptors on susceptible mammalian cells and enter the cells by endocytic uptake. During cell entry, the 32 kDa A subunit is cleaved by the membrane-anchored protease furin to generate a catalytically active, 27·5 kDa A1 fragment and a 4·5 kDa A2 fragment. Previous studies have shown that mutating the furin site to prevent cleavage did not significantly affect toxin potency, suggesting that cleavage is not required for toxin activity. Here it is confirmed that preventing cleavage at the usual processing site does not prevent proteolytic processing of the Escherichia coli Shiga-like toxin-1 A subunit. However, simultaneous mutation of both the primary furin-recognition site and a nearby putative furin cleavage site did prevent intracellular processing of the A subunit. Comparison of the cytotoxicities of purified recombinant toxins to cultured

Proteases (also called Proteolytic Enzymes, Peptidases, or Proteinases) are enzymes that hydrolyze the amide bonds within proteins or peptides. Most proteases act in a specific manner, hydrolyzing bonds at or adjacent to specific residues or a specific sequence of residues contained within the substrate protein or peptide. Proteases play an important role in most diseases and biological processes including prenatal and postnatal development, reproduction, signal transduction, the immune response, various autoimmune and degenerative diseases, and cancer. They are also an important research tool, frequently used in the analysis and production of proteins. Furin is a calcium dependent serine endoprotease that processes numerous proproteins of different secretory pathways into their mature forms by cleaving at the carboxyl side of the recognition sequence, R-Xaa-(K/R)-R, where Xaa can be any amino acid. Recombinant human Furin is a 61.7 kDa protein, corresponding to residues 124 through 715 of the ...

of Sciences of Ukraine, Kyiv. Received: 22 February 2019; Accepted: 17 May 2019. A series of novel triphenylphosphonium derivatives of 1,3-oxazole containing at C2 and C5-positions electron withdrawing or electron-donating groups were synthesized and characterized by 1H, 31P NMR and IR spectroscopy, element analysis and chromato-mass spectrometry. These compounds were found to be a new class of non-peptide inhibitors of furin. Depending on the chemical structure, they inactivated enzyme at micromolar level by mechanism of competitive, non-competitive or mixed inhibition. Evaluation of the synthesized derivatives as furin inhibitors showed that among the triphenylphosphonium salts studied by us, oxazole 12 containing 2,4-dichlorophenyl- in the C2-position and MeS-group at C5 is the most active (Ki = 1.57 μM) competitive inhibitor of furin. Our results provided evidence that chemical modification of 1,3-oxazole-4-yl-triphenylphosphonium salts may be useful for developing new more potent and ...

This tutorial explains the structure of the spike protein, accompanied by interactive animations of structures determined by cryo-electron microscopy. The furin cleavage site and receptor binding sites are marked. Animations ready for Powerpoint slides are provided. The coronavirus pandemic of 2020 is caused by a virus called SARS-CoV-2. In the electron microscope, it looks like a crown because it has protein spikes sticking out. The virus enters and infects cells after its spike protein binds to the ACE2 receptor on cells in the lung or elsewhere. The first step in this binding is priming, when a protein-cutting enzyme such as furin clips the spike protein. This causes it to stick out its receptor-binding domain, making binding to ACE2 more efficient. Relevant research journal publications are cited. ...

The potency of immunotherapies targeting endogenous tumor antigens is impeded by immune tolerance. portrayed in the growth microenvironment. We present that our healing proteins packed antigenic epitope onto the surface area of mesothelin-expressing growth cells particularly, object rendering tumors prone to antigen-specific cytotoxic Compact disc8+ Testosterone levels lymphocytes (CTL)-mediated eliminating and and and prospects to MHC course I demonstration of Ovum peptide to OVA-specific Compact disc8+ Capital t cells We produced a chimeric antihuman mesothelin scFv (Meso-scFv) conjugated with Fc (IgG2a) proteins made up of ovalbumin (Ovum) peptide flanked by furin cleavage sites (Meso-scFv-ROR-Fc). Physique 1a displays the schematic diagram of chimeric Meso-scFv-ROR-Fc create, control Meso-scFv-Fc proteins without Ovum peptide, and control Meso-scFv-O-Fc proteins without furin cleavage sites. As demonstrated in Physique 1b, just the furin-expressing baby hamster kidney 21 cells transfected ...

The potency of immunotherapies targeting endogenous tumor antigens is impeded by immune tolerance. portrayed in the growth microenvironment. We present that our healing proteins packed antigenic epitope onto the surface area of mesothelin-expressing growth cells particularly, object rendering tumors prone to antigen-specific cytotoxic Compact disc8+ Testosterone levels lymphocytes (CTL)-mediated eliminating and and and prospects to MHC course I demonstration of Ovum peptide to OVA-specific Compact disc8+ Capital t cells We produced a chimeric antihuman mesothelin scFv (Meso-scFv) conjugated with Fc (IgG2a) proteins made up of ovalbumin (Ovum) peptide flanked by furin cleavage sites (Meso-scFv-ROR-Fc). Physique 1a displays the schematic diagram of chimeric Meso-scFv-ROR-Fc create, control Meso-scFv-Fc proteins without Ovum peptide, and control Meso-scFv-O-Fc proteins without furin cleavage sites. As demonstrated in Physique 1b, just the furin-expressing baby hamster kidney 21 cells transfected ...

The severe acute respiratory coronavirus 2 (SARS-CoV-2) spike (S) protein is the target of vaccine design efforts to end the coronavirus disease 2019 (COVID-19) pandemic. Despite a low mutation rate, isolates with the D614G substitution in the S protein appeared early during the pandemic and are now the dominant form worldwide. Here, we explore S conformational changes and the effects of the D614G mutation on a soluble S ectodomain construct. Cryoelectron microscopy (cryo-EM) structures reveal altered receptor binding domain (RBD) disposition; antigenicity and proteolysis experiments reveal structural changes and enhanced furin cleavage efficiency of the G614 variant. Furthermore, furin cleavage alters the up/down ratio of the RBDs in the G614 S ectodomain, demonstrating an allosteric effect on RBD positioning triggered by changes in the SD2 region, which harbors residue 614 and the furin cleavage site. Our results elucidate SARS-CoV-2 S conformational landscape and allostery and have ...

Furin cleavage assays of fluorogenic peptides. A) Synthetic fluorogenic peptides were generated with sequences matching consensus feline enteric coronavirus and

TY - JOUR. T1 - Evidence for furin-type activity-mediated C-terminal processing of profibrillin-1 and interference in the processing by certain mutations. AU - Lönnqvist, L.. AU - Reinhardt, D.. AU - Sakai, Lynn. AU - Peltonen, L.. PY - 1998/12/1. Y1 - 1998/12/1. N2 - Fibrillin-1 is a major component of the 10 nm microfibrils of the extracellular matrix (ECM). It is synthesized as an ~350 kDa precursor molecule, profibrillin-1, which is proteolytically processed into its biologically active ~320 kDa form. Furin, a calcium-dependent endoprotease of the subtilisin family, which is known to be the processing enzyme for a variety of proproteins, is believed to be responsible for the N-terminal proteolytic cleavage of profibrillin-1. In this article we provide several lines of evidence that the C-terminal trimming of profibrillin-1 also occurs via a furin-type activity. Edman degradation of a small recombinant C-terminal subdomain of fibrillin-1 revealed complete processing of the peptide ...

Two stable cell lines expressing the hBMP2 gene, CHO-BMP2 and HEK-BMP2, were cultured in the presence of IND-1 in short-term (24 h, multi-well) and long-term (two-month, perfusion flasks) cultures. The rhBMP-2 produced was characterized by Western blot and its activity assessed using the C2C12 cell-based assay. The amount of proBMP-2 and mature BMP-2 produced was quantified by ELISA. The mRNA level of BMP-2 and furin in cells treated with or without IND-1 was compared by real-time RT-PCR. Cellular uptake of IND-1 was estimated by measuring the fluorescence of cell lysates following incubation with FITC labeled IND-1. Cellular PC activity post IND-1 incubation was measured using the Boc-RVRR-AMC substrate. Furin-specific siRNA was used to knock down the furin expression in CHO-BMP2 cells and its effect on the rhBMP-2 production was determined. ...

The outbreak of the COVID-19 pandemic is the reason of the current global health crisis. The development of effective antiviral compounds and vaccines requires detailed descriptive studies of SARS-CoV-2 proteins. The SARS-CoV-2 spike (S) protein mediates virion binding to human cells through its interaction with the cell surface receptor ACE2 and is one of the major immunization targets. The functional virion consists of three S1 and three S2 subunits formed by the furin cleavage of the spike protein at R682, a polybasic cleavage site that differs from the animal version of the protein as well as from the SARS-CoV spike protein 2002. We analyzed the glycoprotein using our newly developed methodology based on low-energy fragmentation and cyclic ion mobility. Our analysis confirms the O-glycosylation of the spike protein on a threonine (T678) located near the furin cleavage site. This site is occupied by core-1 and core-2 structures. Two other predicted O-glycosites are unoccupied. We identified ...

The -secretase (BACE1) is initially synthesized as a partially active zymogen containing a prodomain which can be further activated through proteolytic cleavage of the prodomain by a furin-like protease. The active site of BACE1 is large and although a number of high-affinity active-site inhibitors of BACE1 have been described, most of these compounds are large, polar and do not cross the blood-brain barrier. However, it may be possible to target other regions of the protein which regulate BACE1 allosterically. We have found that proBACE1 can be stimulated by relatively low concentrations (e.g. 1 µg/ml) of heparin. Heparin initially increases proBACE1 activity, probably by binding to the prodomain, which decreases steric inhibition at the active site. However, the heparin-activated zymogen also undergoes autocatalysis, which ultimately leads to a loss of enzyme activity. We speculate that proBACE1 can be regulated by endogenous heparan sulfate proteoglycans and that drugs which target this ...

Peptides , SARS-CoV2,COVID-19 Peptides , 96-well Overlapping Peptide Library; Specifications: Spike glycoprotein from SARS-CoV-2: NCBI the RefSeq (YP_009724390.1) of 1273 aa Peptide length: 15 aa, 9aa for the C-terminal peptide. Gross peptide weight: not quantified Synthesis scale: 5 mol Offset number: 7 aa Format: lyophilized in 96-well plate (P-DW-20-C Axygen ) Counter ion: TFA QC validation: MALDI-TOF QC on 10 % of peptidesCoronavirus Spike glycoproteins mediate the virus attachment to host cell surface receptors and facilitate the virus entry by assisting fusion between viral and host cell membranes. They are composed of two subunits, S1 and S2. The Spike protein of SARS-CoV-2 harbors furin cleavage sites, one being located at the boundary between S1 and S2.

In this study, 48% of the human melanoma cell lines tested were sensitive to PA-L1/LF. Although our studies indicated that modification of the furin cleavage site did slightly reduce PA potency, we have successfully showed the enhanced selectivity of PA-L1/LF. We were able to show that all PA-L1/LF-sensitive melanoma cell lines exhibited high PA-L1 activation as determined by a FRET disruption-based assay (13). Subsequently, we showed that these cell lines exhibited high cell lysate MMP-2 and/or MMP-9 activity. Furthermore, we showed that 11 of 12 PA-L1/LF-sensitive melanoma cell lines carried the B-RAF V600E mutation. In a study including nonmelanoma human tumors, cells carrying this specific mutation were similarly highly susceptible to LF-mediated MAPK inhibition (11). These results show that the B-RAF V600E is closely associated with sensitivity to LF-mediated cell death. Furthermore, small molecular weight inhibitors of MEK1 show similar B-RAF V600E-dependent melanoma cell cytotoxicity ...

Theres another aspect of the furin cleavage site that narrows the path for a natural emergence origin even further.. As everyone knows (or may at least recall from high school), the genetic code uses three units of DNA to specify each amino acid unit of a protein chain. When read in groups of 3, the 4 different kinds of DNA unit can specify 4 x 4 x 4 or 64 different triplets, or codons as they are called. Since there are only 20 kinds of amino acid, there are more than enough codons to go around, allowing some amino acids to be specified by more than one codon. The amino acid arginine, for instance, can be designated by any of the six codons CGU, CGC, CGA, CGG, AGA or AGG, where A, U, G and C stand for the four different kinds of unit in RNA.. Heres where it gets interesting. Different organisms have different codon preferences. Human cells like to designate arginine with the codons CGT, CGC or CGG. But CGG is coronaviruss least popular codon for arginine. Keep that in mind when looking at ...

The Wuhan institutes most recent chimeric virus used a very different coronavirus as its genetic backbone. Looking at the body of research produced there, its clear that scientists were laser-focused on the bat viruses related to SARS-CoV, which spurred research on coronaviruses worldwide after it emerged in 2003 because of its pandemic potential. Theres just no trace of SARS-CoV-2 in the lab, and if the SARS-CoV-2 progenitor or its building blocks werent in the lab before the pandemic, the pandemic could not have started there - even accidentally. This precludes the possibility that SARS-CoV-2 evolved via serial passage in cell culture, or repeated rounds of infection of other cells in a lab, as do other observations about the virus. In standard cell culture, features like the furin cleavage site that are crucial for transmission and disease in humans are rapidly lost as the virus begins adapting to the vervet monkey kidney cells typically used to grow it. For the past 18 months, ...

SARS-CoV-2 is the only coronavirus with a furin cleavage site, making it unique among coronaviruses, the smoking gun that proves SARS-CoV-2 was lab-created.

Generating a soluble and native-like trimeric envelope glycoprotein (Env) with high efficacy as an immunogen has been a major focus for developing an effective vaccine against HIV-1. The Env immunogen is a heavily glycosylated protein composed of 3 identical surface gp120 and gp41 subunits that form into a trimer o

A common feature of transmembrane collagens is the presence of two forms of the molecule: a full-length membrane-bound form and an ectodomain shed form. This characteristic can be also applicable to collagen XXIII. The distribution of both collagen XXIII forms is tissue-specific, since there are organs such as the brain where the shed form is predominant, whereas in the lungs the molecule is generally found as the full-length form. It has been reported that the cell is able to regulate the amounts of collagen XXIII in the membrane-bound form and in the secreted shed form, influencing the production of one form or the other when it is needed. For that reason, the shedding process of collagen XXIII has been described as a selective proteolysis, carried out principally by furin,[4] although there are other enzymes, like serine and cysteine proteases, which are able to shed the molecule too. When collagen XXIII is inside the Golgi apparatus, furin proteases act, cleaving the protein and originating ...

To maintain its high infectivity while keeping its RBD less accessible, SARS-CoV-2 relies on a second strategy-host protease activation. Host protease activation is a significant determinant of coronavirus infection and pathogenesis, and a significant target for host immune surveillance and human intervention strategies. Using a combination of mutagenesis, protease inhibitors, and siRNA approaches, here we showed that furin preactivation enhances SARS-CoV-2 pseudovirus entry into different types of hACE2-expressing cell lines, including lung epithelial and lung fibroblast cell lines. We also showed that cell surface protease TMPRSS2 and lysosomal cathepsins activate SARS-CoV-2 pseudovirus entry and that both TMPRSS2 and cathepsins have cumulative effects with furin on SARS-CoV-2 entry. In comparison, SARS-CoV pseudovirus entry is activated by TMPRSS2 and cathepsins, but not furin. Furin preactivation allows SARS-CoV-2 to be less dependent on target cells, enhancing its entry into some target ...

H5N1, an influenza, binds well onto to &alpha2-3 sialic acid. It contains RNA segments that help produce disease. For example, one RNA segment turns off a component of the immune system. H5 is cleaved by the protease furin, which is present in all cells. Therefore, H5N1 is a pantropic virus able to infect all tissues of avians. It is so pathogenic because, although most human lung cells express α2-6 sialic acid, there are a few expressing α2-3 sialic acid. The H5N1 virus infects these cells and causes a potentially fatal very strong immune response. It can infect poultry, and may adapt to be transmissble from human-to-human. ...

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Reactome is pathway database which provides intuitive bioinformatics tools for the visualisation, interpretation and analysis of pathway knowledge.

The second notable feature of SARS-CoV-2 is a polybasic cleavage site (RRAR) at the junction of S1 and S2, the two subunits of the spike8 (Fig. 1b). This allows effective cleavage by furin and other proteases and has a role in determining viral infectivity and host range12. In addition, a leading proline is also inserted at this site in SARS-CoV-2; thus, the inserted sequence is PRRA (Fig. 1b). The turn created by the proline is predicted to result in the addition of O-linked glycans to S673, T678 and S686, which flank the cleavage site and are unique to SARS-CoV-2 (Fig. 1b). Polybasic cleavage sites have not been observed in related lineage B betacoronaviruses, although other human betacoronaviruses, including HKU1 (lineage A), have those sites and predicted O-linked glycans13. Given the level of genetic variation in the spike, it is likely that SARS-CoV-2-like viruses with partial or full polybasic cleavage sites will be discovered in other species.. The functional consequence of the ...

The second notable feature of SARS-CoV-2 is a polybasic cleavage site (RRAR) at the junction of S1 and S2, the two subunits of the spike8 (Fig. 1b). This allows effective cleavage by furin and other proteases and has a role in determining viral infectivity and host range12. In addition, a leading proline is also inserted at this site in SARS-CoV-2; thus, the inserted sequence is PRRA (Fig. 1b). The turn created by the proline is predicted to result in the addition of O-linked glycans to S673, T678 and S686, which flank the cleavage site and are unique to SARS-CoV-2 (Fig. 1b). Polybasic cleavage sites have not been observed in related lineage B betacoronaviruses, although other human betacoronaviruses, including HKU1 (lineage A), have those sites and predicted O-linked glycans13. Given the level of genetic variation in the spike, it is likely that SARS-CoV-2-like viruses with partial or full polybasic cleavage sites will be discovered in other species.. The functional consequence of the ...

The FDA recently announced approval of two cholesterol lowering drugs which both work by inhibiting PCSK9 or proprotein convertase subtilisin/kexin type 9. PCSK9 is an enzyme from the subtilisin-like proprotein convertase family which include proteases that process proteins trafficking through various constitutive secretory pathways. PCSK9 increases blood cholesterol levels by binding to low densitiy lipoprotein (LDL) receptors, targeting them for lysosomal…

The MSLN gene encodes a precursor protein that is cleaved into two products, megakaryocyte potentiating factor (MPF) and mesothelin. Mesothelin is a glycosylphosphatidylinositol (GPI)-anchored cell-surface protein that may function as a cell-adhesion protein. Mesothelin is cleaved by a furin-like protease to yield fragments of approximately 31 kDa and 40 kDa. In humans, mesothelin is overexpressed in epithelial mesotheliomas, ovarian cancers, and in some types of squamous cell carcinomas. The ERC (expressed in renal carcinoma) gene, which was identified in a rat model of renal cancer, has been proposed as a homolog of the human MSLN gene. It plays a role in cell adhesion and shape. Rat mesothelin was also shown to be dynamically expressed in the developing pancreas. In rat and mouse, mesothelin is also known as ERC/mesothelin, protein expressed in renal carcinoma, and pre-pro-megakaryocyte-potentiating factor.. ...

Many studies have verified that posttranslational modification determines whether FGF23 polypeptide is secreted as an intact biologically active molecule or subjected to intracellular proteolysis (37). FGF23 is O-glycosylated at the residue T178 by the enzyme ppGalNAc-T3 (polypeptide N-acetylgalactosaminyltransferase 3), and this modification blocks its proteolysis and is required for the release of iFGF23 (38). Furthermore, a secretory kinase, FAM20C, phosphorylates FGF23 on S180 within the furin recognition motif R176XXR179/S180, and this phosphorylation blocks the O-glycosylation of FGF23 and renders it susceptible to proteolysis by furin (14, 39). It seems that these dynamic posttranslational processes including phosphorylation, glycosylation, and proteolysis play a critical role in the intracellular regulation of FGF23 homeostasis. In light of these findings, it is important to note that both tPA and uPA can cleave glycosylated FGF23, because glycosylated rhFGF23 was used in this study. ...

Supplementary Components1. sites of Personal computers 4C7 relative to furin. Our findings suggest a new approach for developing selective inhibitors of Personal computers using 1PDX like a scaffold, as evidenced by our capability Bisoprolol fumarate to engineer particular and selective inhibitors of furin and Computers 4C7 extremely. Launch Proprotein convertases (Computers) are ubiquitous calcium mineral reliant serine proteases from the subtilisin flip. In mammals, Computers are complicated multi-domain proteins that perform the proteolytic posttranslational adjustment of several secreted proteins and peptides, and regulate central mobile processes like development and proliferation (1, 2). The Computers from the Kexin-like subtype, furin, Computer4, Computer5, PC7 and PACE4, localize towards the trans-Golgi network and endosomes from the constitutive proteins secretion pathway and cleave precursors of a big diversity of protein at polybasic sites comprising the overall P4Arg-X-X-P1Arg ...

Myostatin, a key regulator of muscle mass in vertebrates, is biosynthesised as a latent precursor in muscle and is activated by sequential proteolysis of the pro-domain. To investigate the molecular mechanism by which pro-myostatin remains latent, we have determined the structure of unprocessed pro-myostatin and analysed the properties of the protein in its different forms. Crystal structures and SAXS analyses show that pro-myostatin adopts an open, V-shaped structure with a domain-swapped arrangement. The pro-mature complex, after cleavage of the furin site, has significantly reduced activity compared with the mature growth factor and persists as a stable complex that is resistant to the natural antagonist follistatin ...

SARS-CoV-2 infection is initiated by virus binding to ACE2 cell surface receptors1-4, followed by fusion of virus and cell membranes to release the virus genome into the cell. Both receptor binding and membrane fusion activities are mediated by the virus Spike glycoprotein, S5-7. As with other class I membrane fusion proteins, S is post-translationally cleaved, in this case by furin, into S1 and S2 components that remain associated following cleavage8-10. Fusion activation following receptor binding is proposed to involve the exposure of a second proteolytic site (S2), cleavage of which is required for the fusion peptide release11,12. We have investigated the binding of ACE2 to the furin-cleaved form of SARS-CoV-2 S by cryoEM. We classify ten different molecular species including the unbound, closed spike trimer, the fully open ACE2-bound trimer, and dissociated monomeric S1 bound to ACE2. The ten structures describe ACE2 binding events which destabilise the spike trimer, progressively opening ...

Human site-1-protease (S1P, MEROPS S08.8063), also widely known as subtilisin/kexin isozyme 1 (SKI-1), is a membrane bound subtilisin-related serine protease, that belongs to a group of nine mammalian proprotein convertases. Among these proteases, S1P displays unique substrate specificity, by showin …

Glypican (GPC)-3 inhibits cell proliferation and regulates cell survival during development. This action is demonstrated by GPC3 loss-of-function mutations in h

Immunization with Human Papillomavirus (HPV) L1 virus‐like particles or L2 capsid protein elicits neutralizing antibodies that mediate protection

Growing numbers of scientists studying COVID-19 globally are suspecting it was man-made in Chinese labs. The UK Daily Telegraph ran a report in early June inter-viewing leading immunologists and geneticists. While COVID-19s genetic code is 96% similar to a bat-coronavirus strain, it is the other 4% that makes it so infectious to humans. This virus is able to bind with human cells 100x to 1000x more efficiently than SARS. And it binds with an ACE2 receptor in human cells far more strongly than in bats! It targets an enzyme in human cells called furin, which works to activate new proteins by cleaving them off after they are synthesized. Thats not how normal bat coronaviruses work, but other highly-contagious diseases like HIV and Ebola target furin. COVID-19 was found to have other HIV-like properties too. Human cells have a molecule called MHC on their surfaces. It acts like a flag to signal the immune system, with changes alerting T cells to kill virus-infected cells. But both HIV and COVID-19 ...

In addition to influencing GF bioactivity, the pro-domains appear to often play a role in protein localisation. Latent-TGF-βs are the best characterised in this way. These proteins associate with components of the extracellular matrix (ECM) and are disulfide-bonded to latent-TGF-β binding proteins (LTBPs) or Leucine rich repeat containing 32 (LRRC32, aka GARP; Figure 2) [25-28]. These inactive, but furin-cleaved, proteins can activated by diverse mechanism, e.g., by mechanical forces, proteolysis and through interaction with other proteins. Interaction with ECM on one end and with integrin binding to the RGD motif on the other end of the pro-domain, can induce mechanical force that distorts the pro-domain releasing the bioactive mature dimer [14] (Figure 2). Thromospondin-1 can bind both to mature and latent forms of TGF-β and activate the latent form both in vitro and in vivo [29, 30]. The BMP-4 pro-domain interacts with fibrillin, targeting the complex to the extracellular matrix where the ...

The structure of the precursor of the muscle mass regulator myostatin/GDF8 shows an open‐armed conformation, similar to pro‐activin A and distinct from pro‐TGF‐β1. An enhanced interface between prodomains and the mature growth factor allows pro‐myostatin to persist as a latent, antagonist resistant complex even after processing of the precursor by furin‐like proteases.. ...

Seddon, James A.; Perez-Velez, Carlos M.; Schaaf, H. Simon; Furin, Jennifer J.; Marais, Ben J.; Tebruegge, Marc; Detjen, Anne et al. (Journal of the Pediatric Infectious Diseases Society, 2013) Link to Published Version ...