Isolation and partial characterization of a complementary protein from the mycoparasite Piptocephalis virginiana that specifically binds to two glycoproteins at the host cell surface

1997 ◽  
Vol 43 (7) ◽  
pp. 625-632 ◽  
Author(s):  
M. S. Manocha ◽  
D. Xiong ◽  
V. Govindsamy
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Western Blot ◽  
Host Cell ◽  
Immunofluorescence Microscopy ◽  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Primary Antibody ◽  
Rabbit Igg ◽  
Host Cell Surface

Immunofluorescence microscopy was used to detect in the mycoparasite Piptocephalis virginiana the presence of a complementary glycoprotein that binds specifically to the host cell surface glycoproteins b and c, reported earlier from our laboratory. Germinated spores of P. virginiana treated with cell wall extract of the host Mortierella pusilla, primary antibody prepared against cell wall glycoproteins b and c, and fluorescein isothiocyanate (FITC) – goat anti-rabbit IgG conjugate showed fluorescence. Immunobinding analysis identified from the mycoparasite a protein of 100 kDa that binds with the host glycoproteins b and c, separately as well as collectively. Its purification was achieved by (i) 60% ammonium sulfate precipitation, (ii) heat treatment, (iii) Sephadex G-100 gel filtration, and (iv) preparative polyacrylamide gel electrophoresis (PAGE). The purity was ascertained by sodium dodecyl sulphate (SDS) – PAGE and Western blot analysis. Positive reaction to periodic acid – Schiff s reagent revealed its glycoprotein nature, and mannose was identified as a major sugar component. The specificity of the polyclonal antibody raised against electrophoretically purified complementary protein in rabbit was confirmed by dot immunobinding and Western blot analyses. Immunofluorescence microscopy revealed surface localization of the protein on the germ tubes of P. virginiana. Fluorescence was also observed at the surface of the germinated spores and hyphae of the host M. pusilla, after treatment with complementary protein from P. virginiana, primary antibody prepared against the complementary protein, and FITC – goat anti-rabbit IgG conjugate.Key words: biotrophic mycoparasite, cell surface agglutinin, glycoprotein immunobinding, immunofluorescence, mucoraceous host.

Cell surface characteristics of Mortierella species and their interaction with a mycoparasite

1984 ◽  
Vol 30 (3) ◽  
pp. 290-298 ◽  
Author(s):  
M. S. Manocha
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Characteristics ◽  
Protein Patterns ◽  
Periodic Acid Schiff ◽  
The Difference ◽  
Chitin And Chitosan

Cell surface characteristics of three Mortierella species differing in their response to a mycoparasite, Piptocephalis virginiana, were examined. Their cell wall composition was typical of mucoraceous fungi with chitin and chitosan as major polysaccharides. Electron microscopy revealed that the mycoparasite penetrated and formed haustoria in the hyphae of susceptible hosts, M. pusilla and M. isabellina. The failure of the parasite to establish contact and penetrate a hypha of the nonhost, M. candelabrum, was not due to cell wall thickness, rigidity, or chitin contents. Markedly different protein patterns obtained from crude alkali extracts of host and nonhost cell walls by sodium dodecyl sulfate – polyacrylamide gel electrophoresis might explain the difference in host and nonhost response to the mycoparasite. Whereas most of the bands differed only in intensity after staining with either Coomassie blue or periodic acid – Schiff reagent, there were two distinct bands of glycoproteins (76 000 and 74 000) observed in the host species which were absent in the nonhost species.

Isolation and partial characterization of host cell surface agglutinin and its role in attachment of a biotrophic mycoparasite

1991 ◽  
Vol 37 (5) ◽  
pp. 377-383 ◽  
Author(s):  
M. S. Manocha ◽  
Y. Chen
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Proteins ◽  
Carbohydrate Binding ◽  
Protein Extract ◽  
Host Cell Wall ◽  
Host Cell Surface ◽  
Faint Band

Cell surface proteins obtained by alkaline extraction from isolated cell walls of Mortierella pusilla and M. candelabrum, host and nonhost, respectively, of the mycoparasite Piptocephalis virginiana, were tested for their ability to agglutinate mycoparasite spores. The host cell wall protein extract had a high agglutinating activity (788 agglutination units/mg) compared with that of the nonhost extract (21 agglutination units/mg). Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of the crude extract of the host revealed four bands, a, b, c, and d, with respective Mr of 117 000, 100 000, 85 000 and 64 000; these bands except for a faint band c, were absent from the nonhost surface. Deletion of proteins b or c from the crude protein extract of the host significantly reduced its agglutinating activity. Proteins b and c, purified by a series of procedures, were shown to be glycoproteins with glucose and N-acetylglucosamine as major saccharides. The agglutinating activity of a mixture of pure proteins b and c was over 500 times that of either glycoprotein alone, suggesting an involvement of both glycoproteins in the agglutination process. Further characterization showed that the two glycoproteins were heat-resistant with respect to their agglutinin function, which could be totally inhibited by three sugars: arabinose, glucose and N-acetyglucosamine. It is suggested that glycoproteins b and c are the two subunits of a carbohydrate-binding agglutinin present at the host cell surface and involved in agglutination and attachment of the mycoparasite germ tubes. Key words: agglutinin, attachment, cell surface, sugars, glycoproteins, mycoparasitism.

Specificity of mycoparasite attachment to the host cell surface

1985 ◽  
Vol 63 (4) ◽  
pp. 772-778 ◽  
Author(s):  
M. S. Manocha
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Lectin Binding ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Host Recognition ◽  
Binding Reaction ◽  
Choanephora Cucurbitarum ◽  
Host Cell Surface ◽  
Germ Tubes

The use of isolated cell wall fragments of Choanephora cucurbitarum (Berk. & Rav.) Thaxter (a host), and of Linderina pennispora Raper and Fennell (a nonhost), has provided not only a convenient method to quantify attachment of the parasite, Piptocephalis virginiana Leadbeater and Mercer, by the artificial inoculation and washing-off procedure, but also an excellent material for investigations on the molecular basis of specificity and host recognition. The parasite germ tubes are attached to the cell wall fragments of the host but not of the nonhost. Attachment was inhibited by the addition of sugars, chitobiose and chitotriose, and by treatment with acid or alkali indicating the involvement of proteins or glycoproteins in recognizing sugar residues at the cell surface. Both host and nonhost showed a positive binding reaction with fluorescent lectins specific for N-acetyl-D-glucosamine oligomer. The cell surface of the nonhost also contains D-galactose and N-acetyl-D-galactosamine residues as lectin binding sites. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis of cell wall extracts of host and nonhost revealed four bands of glycoproteins common to both fungi and two were specific to the host.

Involvement of cell surface sugars in recognition, attachment, and appressorium formation by a mycoparasite

1990 ◽  
Vol 36 (11) ◽  
pp. 771-778 ◽  
Author(s):  
M. S. Manocha ◽  
Y. Chen ◽  
N. Rao
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Host Cell ◽  
Germ Tube ◽  
Lectin Binding ◽  
Fluorescein Isothiocyanate ◽  
Appressorium Formation ◽  
Obvious Effect ◽  
Surface Attachment ◽  
Host Cell Surface

Fluorescein isothiocyanate labeled lectin binding techniques have revealed differences in the distribution pattern of glycosyl residues at the cell wall level between fungi that are hosts and those that are nonhosts of the mycoparasite Piptocephalis virginiana, and at the protoplast level between compatible and incompatible hosts. The cell wall of the compatible hosts (Choanephora cucurbitarum and Mortierella pusilla) and an incompatible host (Phascolomyces articulosus), as well as that of the mycoparasite itself, contains glucose and N-acetylglucosamine. However, the cell wall of a nonhost (Mortierella candelabrum) tested positive with lectins specific for various sugars, including not only glucose and N-acetylglucosamine, but also fucose, N-acetylgalactosamine, and galactose. These latter sugars could also be exposed at the surfaces of hosts and of the mycoparasite, but only after mild treatment with proteinase or when grown in a liquid culture. Pretreatment of the mycoparasite with glucose and N-acetylglucosamine inhibited its attachment to the host cell surface, but had no obvious effect on appressorium formation. On the other hand, appressorium formation was inhibited by heat treatment of host cell wall fragments which still permitted attachment, thus indicating that the factors responsible for attachment and for appressorium formation are different. The protoplast surfaces of compatible hosts contained all the sugars listed above and these protoplasts could attach to the germ tube of the mycoparasite. Only lectins specific for N-acetylglucosamine and for glucose were bound at the protoplast surface of the incompatible host; these protoplasts did not attach to the mycoparasite germ tube. Key words: mycoparasite, appressorium formation, lectins, host cell surface, attachment, protoplast surface.

scholarly journals A Single-Pass Type I Membrane Protein from the Apicomplexan Parasite Cryptosporidium parvum with Nanomolar Binding Affinity to Host Cell Surface

2021 ◽  
Vol 9 (5) ◽  
pp. 1015
Author(s):  
Tianyu Zhang ◽  
Xin Gao ◽  
Dongqiang Wang ◽  
Jixue Zhao ◽  
Nan Zhang ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Host Cell ◽  
Binding Affinity ◽  
Cryptosporidium Parvum ◽  
Host Cells ◽  
Watery Diarrhea ◽  
Type I ◽  
Single Pass ◽  
Host Cell Surface

Cryptosporidium parvum is a globally recognized zoonotic parasite of medical and veterinary importance. This parasite mainly infects intestinal epithelial cells and causes mild to severe watery diarrhea that could be deadly in patients with weakened or defect immunity. However, its molecular interactions with hosts and pathogenesis, an important part in adaptation of parasitic lifestyle, remain poorly understood. Here we report the identification and characterization of a C. parvum T-cell immunomodulatory protein homolog (CpTIPH). CpTIPH is a 901-aa single-pass type I membrane protein encoded by cgd5_830 gene that also contains a short Vibrio, Colwellia, Bradyrhizobium and Shewanella (VCBS) repeat and relatively long integrin alpha (ITGA) N-terminus domain. Immunofluorescence assay confirmed the location of CpTIPH on the cell surface of C. parvum sporozoites. In congruence with the presence of VCBS repeat and ITGA domain, CpTIPH displayed high, nanomolar binding affinity to host cell surface (i.e., Kd(App) at 16.2 to 44.7 nM on fixed HCT-8 and CHO-K1 cells, respectively). The involvement of CpTIPH in the parasite invasion is partly supported by experiments showing that an anti-CpTIPH antibody could partially block the invasion of C. parvum sporozoites into host cells. These observations provide a strong basis for further investigation of the roles of CpTIPH in parasite-host cell interactions.

scholarly journals Evidence for a Host Cell Surface Antigen on the Envelope of Avian Tumour Viruses

1975 ◽  
Vol 27 (2) ◽  
pp. 151-161 ◽  
Author(s):  
M. Aupoix ◽  
P. Vigier
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Surface Antigen ◽  
Cell Surface Antigen ◽  
Host Cell Surface

scholarly journals Extracellular Bacterial Pathogen Induces Host Cell Surface Reorganization to Resist Shear Stress

2009 ◽  
Vol 5 (2) ◽  
pp. e1000314 ◽  
Author(s):  
Guillain Mikaty ◽  
Magali Soyer ◽  
Emilie Mairey ◽  
Nelly Henry ◽  
Dave Dyer ◽  
...  
Keyword(s):  
Shear Stress ◽  
Cell Surface ◽  
Host Cell ◽  
Bacterial Pathogen ◽  
Host Cell Surface

scholarly journals Heparan Sulfate Facilitates Spike Protein-Mediated SARS-CoV-2 Host Cell Invasion and Contributes to Increased Infection of SARS-CoV-2 G614 Mutant and in Lung Cancer

2021 ◽  
Vol 8 ◽  
Author(s):  
Jingwen Yue ◽  
Weihua Jin ◽  
Hua Yang ◽  
John Faulkner ◽  
Xuehong Song ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cell Surface ◽  
Heparan Sulfate ◽  
Host Cell ◽  
Host Cells ◽  
Cell Surface Receptor ◽  
Spike Protein ◽  
Effective Prevention ◽  
Heparin Derivatives ◽  
Host Cell Surface

The severe acute respiratory syndrome (SARS)-like coronavirus disease (COVID-19) is caused by SARS-CoV-2 and has been a serious threat to global public health with limited treatment. Cellular heparan sulfate (HS) has been found to bind SARS-CoV-2 spike protein (SV2-S) and co-operate with cell surface receptor angiotensin-converting enzyme 2 (ACE2) to mediate SARS-CoV-2 infection of host cells. In this study, we determined that host cell surface SV2-S binding depends on and correlates with host cell surface HS expression. This binding is required for SARS-Cov-2 virus to infect host cells and can be blocked by heparin lyase, HS antagonist surfen, heparin, and heparin derivatives. The binding of heparin/HS to SV2-S is mainly determined by its overall sulfation with potential, minor contribution of specific SV2-S binding motifs. The higher binding affinity of SV2-S G614 mutant to heparin and upregulated HS expression may be one of the mechanisms underlying the higher infectivity of the SARS-CoV-2 G614 variant and the high vulnerability of lung cancer patients to SARS-CoV-2 infection, respectively. The higher host cell infection by SARS-CoV-2 G614 variant pseudovirus and the increased infection caused by upregulated HS expression both can be effectively blocked by heparin lyase and heparin, and possibly surfen and heparin derivatives too. Our findings support blocking HS-SV2-S interaction may provide one addition to achieve effective prevention and/treatment of COVID-19.

scholarly journals Significance of thiol-disulfide balance in SARS-CoV-2 infection

2021 ◽  
Vol 72 (3) ◽  
pp. 30-36
Author(s):  
Tatjana Simić
Keyword(s):  
Cell Surface ◽  
Host Cell ◽  
Protein Interactions ◽  
Viral Entry ◽  
Molecular Mechanisms ◽  
Alkylating Agents ◽  
Protein S ◽  
Protein Protein Interactions ◽  
Virus Surface ◽  
Host Cell Surface

Studies of the molecular mechanisms regarding interaction of different viruses with receptors on the host cell surface have shown that the viral entry depends on the specific relationship between free thiol (SH) groups and disulfides on the virus surface, as well as the thiol disulfide balance on the host cell surface. The presence of oxidizing compounds or alkylating agents, which disturb the thiol-disulfide balance on the surface of the virus, can also affect its infectious potential. Disturbed thiol-disulfide balance may also influence protein-protein interactions between SARS-CoV-2 protein S and ACE2 receptors of the host cell. This review presents the basic mechanisms of maintaining intracellular and extracellular thiol disulfide balance and previous experimental and clinical evidence in favor of impaired balance in SARS-CoV-2 infection. Besides, the results of the clinical application or experimental analysis of compounds that induce changes in the thiol disulfide balance towards reduction of disulfide bridges in proteins of interest in COVID-19 infection are presented.

Brilacidin, a COVID-19 Drug Candidate, demonstrates broad-spectrum antiviral activity against human coronaviruses OC43, 229E and NL63 through targeting both the virus and the host cell

2021 ◽  
Author(s):  
Yanmei Hu ◽  
Hyunil Jo ◽  
William DeGrado ◽  
Jun Wang
Keyword(s):  
Antiviral Activity ◽  
Cell Surface ◽  
Host Cell ◽  
Mechanism Of Action ◽  
Broad Spectrum ◽  
Drug Candidate ◽  
Host Defense Peptides ◽  
Antibiotic Drug ◽  
Host Cell Surface ◽  
Human Coronaviruses

Brilacidin, a mimetic of host defense peptides (HDPs), is currently in phase 2 clinical trial as an antibiotic drug candidate. A recent study reported that brilacidin has antiviral activity against SARS-CoV-2 by inactivating the virus. In this work, we discovered an additional mechanism of action of brilacidin by targeting heparan sulfate proteoglycans (HSPGs) on host cell surface. Brilacidin, but not acetyl brilacidin, inhibits the entry of SARS-CoV-2 pseudovirus into multiple cell lines, and heparin, a HSPG mimetic, abolishes the inhibitory activity of brilacidin on SARS-CoV-2 pseudovirus cell entry. In addition, we found that brilacidin has broad-spectrum antiviral activity against multiple human coronaviruses (HCoVs) including HCoV-229E, HCoV-OC43, and HCoV-NL63. Mechanistic studies revealed that brilacidin has a dual antiviral mechanism of action including virucidal activity and binding to coronavirus attachment factor HSPGs on host cell surface. Brilacidin partially loses its antiviral activity when heparin was included in the cell cultures, supporting the host-targeting mechanism. Drug combination therapy showed that brilacidin has a strong synergistic effect with remdesivir against HCoV-OC43 in cell culture. Taken together, this study provides appealing findings for the translational potential of brilacidin as a broad-spectrum antiviral for coronaviruses including SARS-CoV-2.

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