LamB as a carrier molecule for the functional exposition of IgG-binding domains of the Staphylococcus aureus Protein A at the surface of Escherichia coli K12

1993 ◽  
Vol 236-236 (2-3) ◽  
pp. 187-192 ◽  
Author(s):  
Lothar Steidler ◽  
Erik Remaut ◽  
Walter Fiers
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Protein A ◽  
Escherichia Coli K12 ◽  
Binding Domains ◽  
Igg Binding ◽  
Carrier Molecule

Comparative sequence analysis of spa gene of Staphylococcus aureus isolated from bovine mastitis: characterization of an unusual spa gene variant

2006 ◽  
Vol 73 (3) ◽  
pp. 322-327 ◽  
Author(s):  
Amr El-Sayed ◽  
Jörg Alber ◽  
Christoph Lämmler ◽  
Amir Abdulmawjood ◽  
Michael Zschöck ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Protein A ◽  
Bovine Mastitis ◽  
Clinical Mastitis ◽  
Comparative Sequence Analysis ◽  
Border Region ◽  
Binding Domains ◽  
Igg Binding ◽  
Spa Gene

The protein A encoding gene spa of four Staphylococcus aureus strains isolated from bovine clinical mastitis was amplified by PCR and sequenced. The four strains were selected after an initial screening of spa gene of 41 strains isolated from mastitic cows and were subjected to detailed investigations. According to the sequencing results the spa gene of three strains (M1, M2, M3) appeared with gene segments encoding five (E, D, A, B and C) and four (E, A, B and C) IgG binding domains for two (M1, M3) and one (M2) strain, respectively and with gene segments encoding four, two and two repeats of the octapeptide Xr-repeats for the strains M1, M2 and M3, respectively. For the remaining Staph. aureus strain (M4) gene segments encoding IgG binding domains E, D and A and a new domain BC with a size of 219 bp could be observed. The BC domain appears, with a deletion of a 123 bp segment from the border region between both domains, as fused domain of both previously characterized domains. The Xr-region of this strain had 11 octapeptide repeats.

scholarly journals Recombinant Escherichia coli Strain Produces a ZZ Domain Displaying Biopolyester Granules Suitable for Immunoglobulin G Purification

2006 ◽  
Vol 72 (11) ◽  
pp. 7394-7397 ◽  
Author(s):  
Jane A. Brockelbank ◽  
Verena Peters ◽  
Bernd H. A. Rehm
Keyword(s):  
Escherichia Coli ◽  
Immunoglobulin G ◽  
Binding Capacity ◽  
Protein A ◽  
Coli Strain ◽  
Pha Synthase ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ionization Time ◽  
Flight Mass Spectrometry ◽  
Igg Binding

ABSTRACT The immunoglobulin G (IgG) binding ZZ domain of protein A from Staphylococcus aureus was fused to the N terminus of the polyhydroxyalkanoate (PHA) synthase from Cupriavidus necator. The fusion protein was confirmed by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry and mediated formation of ZZ domain-displaying PHA granules in recombinant Escherichia coli. The IgG binding capacity of isolated granules was assessed using enzyme-linked immunosorbent assay and could be enhanced by the overproduction of the ZZ-PHA synthase. ZZ-PHA granules enabled efficient purification of IgG from human serum.

scholarly journals Fibroblast adhesion to recombinant tropoelastin expressed as a protein A-fusion protein

1991 ◽  
Vol 273 (3) ◽  
pp. 517-522 ◽  
Author(s):  
L E Grosso ◽  
W C Parks ◽  
L J Wu ◽  
R P Mecham
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Fusion Protein ◽  
Developmental Regulation ◽  
Protein A ◽  
Receptor Binding Site ◽  
Elastin Receptor ◽  
Cnbr Cleavage ◽  
Fibroblast Adhesion ◽  
Nuchal Ligament

A bovine tropoelastin cDNA encoding exons 15-36 that includes the elastin-receptor binding site was expressed in Escherichia coli as a fusion protein with Protein A from Staphylococcus aureus. After isolation of the fusion protein by affinity chromatography on Ig-Sepharose, the tropoelastin domain was separated from plasmid-pR1T2T-encoded Protein A (Protein A') by CNBr cleavage. Cell-adhesion assays demonstrated specific adhesion to the recombinant tropoelastin. Furthermore, the data indicate that interactions involving the bovine elastin receptor mediate nuchalligament fibroblast adhesion to the recombinant protein. In agreement with earlier studies of fibroblast chemotaxis to bovine tropoelastin, nuchal-ligament fibroblast adhesion demonstrated developmental regulation of the elastin receptor.

scholarly journals The Sbi Protein Is a Multifunctional Immune Evasion Factor of Staphylococcus aureus

2011 ◽  
Vol 79 (9) ◽  
pp. 3801-3809 ◽  
Author(s):  
Emma Jane Smith ◽  
Livia Visai ◽  
Steven W. Kerrigan ◽  
Pietro Speziale ◽  
Timothy J. Foster
Keyword(s):  
Staphylococcus Aureus ◽  
Immune Evasion ◽  
Capsular Polysaccharide ◽  
Cell Envelope ◽  
Protein A ◽  
Biologically Active ◽  
Complement Protein ◽  
Content Type ◽  
Binding Domains ◽  
Terminal Domains

ABSTRACTThe second immunoglobulin-binding protein (Sbi) ofStaphylococcus aureushas two N-terminal domains that bind the Fc region of IgG in a fashion similar to that of protein A and two domains that can bind to the complement protein C3 and promote its futile consumption in the fluid phase. It has been proposed that Sbi helps bacteria to avoid innate immune defenses. By comparing a mutant defective in Sbi with mutants defective in protein A, clumping factor A, iron-regulated surface determinant H, and capsular polysaccharide, it was shown that Sbi is indeed an immune evasion factor that promotes bacterial survival in whole human blood and the avoidance of neutrophil-mediated opsonophagocytosis. Sbi is present in the culture supernatant and is also associated with the cell envelope.S. aureusstrains that expressed truncates of Sbi lacking N-terminal domains D1 and D2 (D1D2) or D3 and D4 (D3D4) or a C-terminal truncate that was no longer retained in the cell envelope were analyzed. Both the secreted and envelope-associated forms of Sbi contributed to immune evasion. The IgG-binding domains contributed only when Sbi was attached to the cell, while only the secreted C3-binding domains were biologically active.

scholarly journals Protein A-Specific Monoclonal Antibodies and Prevention of Staphylococcus aureus Disease in Mice

2012 ◽  
Vol 80 (10) ◽  
pp. 3460-3470 ◽  
Author(s):  
Hwan Keun Kim ◽  
Carla Emolo ◽  
Andrea C. DeDent ◽  
Fabiana Falugi ◽  
Dominique M. Missiakas ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Monoclonal Antibodies ◽  
Immune Responses ◽  
B Cell ◽  
Protein A ◽  
Abscess Formation ◽  
Staphylococcal Protein A ◽  
Content Type ◽  
Humoral Immune ◽  
Binding Domains

ABSTRACTStaphylococcus aureusis a leading cause of human soft tissue infections and bacterial sepsis. The emergence of antibiotic-resistant strains (methicillin-resistantS. aureus[MRSA]) has prompted research into staphylococcal vaccines and preventive measures. The envelope ofS. aureusis decorated with staphylococcal protein A (SpA), which captures the Fcγ portion of immunoglobulins to prevent opsonophagocytosis and associates with the Fab portion of VH3-type B cell receptors to trigger B cell superantigen activity. Nontoxigenic protein A (SpAKKAA), when used as an immunogen in mice, stimulates humoral immune responses that neutralize the Fcγ and the VH3+Fab binding activities of SpA and provide protection from staphylococcal abscess formation in mice. Here, we isolated monoclonal antibodies (MAbs) against SpAKKAAthat, by binding to the triple-helical bundle fold of its immunoglobulin binding domains (IgBDs), neutralize the Fcγ and Fab binding activities of SpA. SpAKKAAMAbs promoted opsonophagocytic killing of MRSA in mouse and human blood, provided protection from abscess formation, and stimulated pathogen-specific immune responses in a mouse model of staphylococcal disease. Thus, SpAKKAAMAbs may be useful for the prevention and therapy of staphylococcal disease in humans.

scholarly journals The first report on the sortase-mediated display of bioactive protein A from Staphylococcus aureus (SpA) on the surface of the vegetative form of Bacillus subtilis

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Samira Ghaedmohammadi ◽  
Gholamreza Ahmadian
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Bacillus Subtilis ◽  
Binding Capacity ◽  
Surface Display ◽  
Protein A ◽  
Production Costs ◽  
Rabbit Serum ◽  
Experimental Conditions ◽  
Binding Domains

AbstractProtein A (SpA) is one of the most important Staphylococcus aureus cell wall proteins. It includes five immunoglobulin (Ig)-binding domains which can bind to immune complexes through the Fc region of immunoglobulins. The binding of SpA to the polymeric supports can be used to prepare affinity chromatography resins, which are useful for immunoprecipitation (IP) of antibodies. Protein A is also used to purify many anti-cancer antibodies. In this study, SpA was displayed on the surface of Bacillus subtilis cells using a sortase-mediated system to display the target protein to the B. subtilis cell wall. A series of plasmids consisting of cassettes for cell wall-directed protein A as well as negative controls were constructed and transformed into B. subtilis WASD (wprA sigD) cells. SDS-PAGE, western blot, flow cytometry, functional IgG purification assay, and a modified ELISA assay were used to confirm the surface display of SpA and evaluate its function. Semi-quantitative ELISA results showed that the binding capacity of lyophilized Bs-SpA is 100 μg IgG from rabbit serum per 1 mg of cells under optimal experimental conditions. Low production costs, optimal performance, and the use of a harmless strain compared to a similar commercial product predict the possible use of SpA immobilization technology in the future.

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