Isolation of Staphylococcus aureus clumping factor

Abstract

Immunochemically identical components were isolated from water-soluble phases of five Staphylococcus aureus strains by affinity chromatography on fibrinogen-linked Sepharose 4B. The elution was performed with 1 M MgCl2. The component could be isolated from sonicated preparations of whole cells, cell walls, and extracellular products of S. aureus but not from sonicated preparations of staphylococcal L-forms or from Staphylococcus epidermidis. Investigations of the eluted component by immunoelectrophoresis and Western blot analysis by use of different polyspecific antibodies to S. aureus raised in rabbits revealed only one immunoprecipitate or one band. By means of gel filtration on Sepharose CL 6B and sodium dodecyl sulfate-polyacrylamide gel electrophoresis a molecular mass of 420,000 and 360,000 was found, respectively. Chemical analysis showed a carbohydrate content of about 20% by weight. By crossed immunoelectrophoresis the isolated component was demonstrated to bind to human fibrinogen. The finding that this purified component inhibited the fibrinogen-induced clumping of staphylococci strongly suggests that the component is the S. aureus clumping factor.

IMMUNIZATION OF MICE WITH THE FIBRONECTIN‐BINDING PROTEIN AND CLUMPING FACTOR FROM STAPHYLOCOCCUS AUREUS: ANTIBODY RESPONSE AND RESISTANCE AGAINST INTRAPERITONEAL INFECTION

“…The high antibody response after immunization with C F could also be a consequence of the structure of this molecule. Analysis of C F showed a high content of polysaccharide by weight (9). C F had a relative molecular mass (M,) of about 400,000 and showed only one band on SDS-page under reduced condition (9).…”

“…Analysis of C F showed a high content of polysaccharide by weight (9). C F had a relative molecular mass (M,) of about 400,000 and showed only one band on SDS-page under reduced condition (9). It was suggested that C F is composed of repeated units of minor glycopeptides interlinked by bindings not broken by SDS in the presence of P-mercaptoetha-no1 (9).…”

“…Clumping factor. Clumping factor (CF) was isolated from strain E 2371 by aEnity-chromatography on fibrinogen-linked Sepharose 4 B as described (9). By polyacrylamide gel electrophoresis only one band was seen on unreduced gels, and no impurities were found in crossed immunoelectrophoresis in rabbit antibodies against S. aureus cell walls (9).…”

“…Clumping factor (CF) was isolated from strain E 2371 by aEnity-chromatography on fibrinogen-linked Sepharose 4 B as described (9). By polyacrylamide gel electrophoresis only one band was seen on unreduced gels, and no impurities were found in crossed immunoelectrophoresis in rabbit antibodies against S. aureus cell walls (9). The isolated component is claimed to be identical or containing the clumping factor, as it was demonstrated to bind to purified fibrinogen as demonstrated by crossed immunoelectrophoresis, and it showed competition with the fibrinogen-induced clumping of whole staphylococci (9).…”

“…A fibronectin-binding protein from S. aureus and S. aureus clumping factor have recently been isolated from S. aureus strain E 237 1 (6,9). In the present study these purified, plasma protein-binding components were used for immunization of mice and the antibody response evaluated.…”