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Biomedical subjects

H Blobel

Publications and source records attributed to H Blobel.

At least 55 records · Page 3Linked to original sources
PubMed1987

A simple method for quantitative determination of bacterial adherence to human and animal epithelial cells.

A simple quantitative method for determining bacterial adherence to epithelial cells was devised. The method involved incubation of fluorescein-labeled bacteria with oral epithelial cells. Non-adherent bacteria were subsequently removed by Percoll density centrifugation. The degree of bacterial adherence was then determined fluorospectrophotometrically by measuring the fluorescein extracted from the adherent bacteria. The degree of adherence corresponded well with the microscopic counts of adherent bacteria. The method proved to be suitable for assaying adherence of both gram-positive and gram-negative bacteria to human as well as animal epithelial cells.

Animals↗
PubMed1987

Specific binding of the human S protein (vitronectin) to streptococci, Staphylococcus aureus, and Escherichia coli.

Specific binding of the 125I-labeled human S protein (vitronectin) which has been shown to be identical with serum-spreading factor, was observed with group A, C, and G streptococci as well as with Staphylococcus aureus and Escherichia coli. The specific binding of S protein to group A, C, and G streptococci was high, whereas the binding to S. aureus and E. coli cultures was moderate. In contrast, group B streptococci and a number of other bacterial species tested did not interact with S protein. The binding of S protein to bacteria was saturable and could be inhibited only by unlabeled S protein but not by albumin. Trypsinization and heat treatment of bacteria destroyed the S-protein binding capacity for group G streptococci, S. aureus, and E. coli but not for group A and C streptococci. Likewise, unlabeled human fibronectin and heparin inhibited the binding of labeled S protein to group G streptococci, S. aureus, and E. coli, but did not influence the binding to group A and C streptococci. Double-reciprocal plots of S-protein binding to group G streptococci indicated that fibronectin inhibited the binding in a competitive manner, while heparin acts in a noncompetitive manner. Moreover, the binding of S protein to G streptococci could be partially by the synthetic peptide Gly-Arg-Gly-Asp-Ser, which contains the cell attachment site of S protein. Trypsin-treated S protein had similar binding activity as untreated S protein for group G streptococci, S. aureus, and E. coli, but showed reduced binding to group A and C streptococci. The present data are indicative of two different types of bacterial binding sites in S protein. The binding to group G streptococci, S. aureus, and E. coli is mediated in part through a domain in the S protein containing the sequence Arg-Gly-Asp, whereas a different site is responsible for the binding to group A and C streptococci.

Bacterial Adhesion↗
PubMed1987

Novel complex formed between a nonproteolytic cell wall protein of group A streptococci and alpha 2-macroglobulin.

Binding of 125I-labeled alpha 2-macroglobulin (alpha 2M) to streptococci belonging to serological groups A, B, C, and G was studied. Streptococci of groups A and G interacted only with native alpha 2M, and those of group C reacted only with alpha 2M-trypsin complex. Binding of alpha 2M to group A streptococci was saturable and reversible. The dissociation constant was 2.02 X 10(-7) M, and the number of binding sites was calculated to be 18,000 per streptococcus. The alpha 2M-binding protein could be solubilized by treatment of group A streptococci with a murolytic enzyme and subsequently purified by affinity chromatography and high-pressure liquid chromatography. The purified protein was homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and had a molecular weight of 78,000. It possessed no proteolytic activity and interacted with native alpha 2M in Western blots (immunoblots). Interaction of purified binding protein with alpha 2M led to a change in the conformation of alpha 2M similar to that obtained by alpha 2M-protease complexes. Reversible binding of a nonproteolytic streptococcal component of alpha 2M is thus a novel feature of alpha 2M reactivity.

Bacterial Proteins↗
PubMed1987

Streptococcal group B type antigen X in group L streptococci.

Extracts from 19 of 29 group L streptococcal cultures reacted distinctly with antiserum against group B streptococcal type antigen X in coagglutination and immunodiffusion tests. The X antigen corresponded to those obtained by streptococci of serological groups B and G.

Agglutination Tests↗
PubMed1986

Screening for bacterial Fc-receptor activity on nitrocellulose membranes.

Fc receptors released from staphylococci and streptococci could be readily detected by direct cultivation of the bacteria on nitrocellulose membranes or by microfiltration of their culture supernatants. The nitrocellulose membranes used in both assays were subsequently treated with human immunoglobulin (Ig) G. The reactions of the Fc receptor with IgG could be demonstrated by use of the respective peroxidase-labelled antibodies against IgG. The resulting color formation, which indicated Fc-receptor activity of the bacterial culture, closely corresponded to the 125I-labelled IgG-binding reactivity.

Collodion↗
PubMed1986

Release of Fc-receptors after streptococcal lysis induced by a lytic enzyme from Streptomyces globisporus.

Streptomyces globisporus produced an enzyme which had lytic effects on streptococci and was able to solubilize their Fc-receptors. A new method for rapid purification of this enzyme was described. It consisted of adsorption to Amberlite CG 50 and subsequent chromatography on CM-cellulose Whatman CM 52. The purified enzyme was free of proteases and had lysed streptococci of serological groups A, B, C, G and L. It released Fc-receptors from the streptococcal surface in a biologically active form.

Enzymes↗
PubMed1986

Binding of human haptoglobin to streptococci of serological group A.

All 11 streptococcal cultures of serological group A possessing antigen T4 bound human 125I-labelled haptoglobin (hp) and fibronectin (fbn). Most of these cultures also interacted with alpha 2-macroglobulin (alpha 2m), immunoglobulin G (IgG) and fibrinogen (fib), none with albumin. The binding of the labelled hp could be completely inhibited by unlabelled hp. It was not significantly affected by the other plasma proteins, except alpha 2m. Trypsinization of the streptococci reduced their binding activities for hp, fbn, fib and alpha 2m, but not for IgG. Extraction with guanidine hydrochloride removed non-covalently bound substances from the streptococci without loss in binding-activity for hp.

Antigens, Bacterial↗
PubMed1986

beta-D-galactosidase activity in streptococci of serological group B.

Group B streptococci isolated from humans differed significantly in beta-D-galactosidase-activity from those of bovine mastitis. This could be demonstrated in a relatively simple and rapid test using a fluorogenic 4-methylumbelliferyl-beta-D-galactoside conjugate. Only 10 (12%) of 82 group B streptococcal cultures from human produced beta-D-galactosidase. On the other hand, 74 (96%) of 77 "bovine" cultures formed this enzyme. Thus, beta-D-galactosidase activity could be used as an additional marker for the differentiation between group B streptococci of human and bovine origin.

Acetylglucosaminidase↗
PubMed1986

Partial characterization of leukocidin from Fusobacterium necrophorum.

Leukocidin from Fusobacterium necrophorum was produced in the diffusate of a dialysis culture. It was free from deoxyribonuclease, fibrinolysin, gelatinase, haemolysin, lipase, caseinase and endotoxin. The leukocidin had a molecular weight between 10,000 and 5,000 as estimated by membrane partition chromatography. It formed precipitin lines with anti-leukocidin-serum in double immunodiffusion tests. Mouse peritoneal cells were characteristically damaged by the leukocidin, as revealed by scanning electron microscopy. The damaged cells lost microvilli and suffered partial destruction of their cell membranes.

Animals↗
PubMed1986

Specific binding sites for monomeric and aggregated beta 2-microglobulin on surface of groups A, B, C, and G streptococci.

Human beta 2-microglobulin (beta 2-m) was isolated from urine samples of patients with tubular dysfunctions and aggregated with glutaraldehyde. Four aggregates with molecular weights of 800,000, 480,000, 260,000, and 60,000 were separated by filtration on Sephacryl S-300. The aggregates and monomeric beta 2-m (11,800 MW) were subsequently labeled with 125I and tested for binding to streptococci. Group A streptococci bound only aggregated beta 2-m with a mean binding of 44.5%. Most of the group G streptococci, on the other hand, bound only monomeric beta 2-m with a mean binding of 58%. Among group B streptococci the serotypes with protein antigens interacted mainly with monomeric beta 2-m and those without protein antigens preferentially with aggregated beta 2-m. Nontypable group B streptococcal serotypes did not bind monomeric or aggregated beta 2-m. Of the streptococci belonging to group C, S. equisimilis reacted with monomeric beta 2-m and S. dysgalactiae with aggregated beta 2-m. S. equi did not interact with monomeric beta 2-m or aggregated beta 2-m. Bindings of monomeric beta 2-m and aggregated beta 2-m were saturable and could be inhibited by the respective unlabeled forms of beta 2-m. Fibrinogen, fibronectin, alpha 2-macroglobulin, haptoglobin, or immunoglobulin G did not inhibit the binding of either form of beta 2-m. The binding sites for monomeric beta 2-m were more susceptible to trypsin than those for aggregated beta 2-m. Treatment of streptococci with pronase destroyed their binding activities for monomeric and aggregated beta 2-m. Both monomeric beta 2-m and aggregated beta 2-m binding sites were sensitive to heat. The Scatchard plots of monomeric beta 2-m and aggregated beta 2-m were linear with Kd of 1.29 X 10(-9) M and 1.9 X 10(-9) M respectively. The number of binding sites per bacterium were estimated to be 81,000 for monomeric beta 2-m and 1,210 for aggregated beta 2-m.

Binding Sites↗
PubMed1986

Rapid differentiation of streptococci isolated from cows with mastitis.

The demonstration of some species-specific streptococcal enzymes with 4-methylumbelliferyl-conjugated beta-D-glucuronide, N-acetyl-beta-D-glucosaminide, and beta-D-mannoside and agglutination with the lectin from Dolichos biflorus allowed rapid tentative identification of streptococci isolated from cows with mastitis.

Acetylglucosaminidase↗
PubMed1985

Purification of oligomeric staphylococcal alpha-toxin by affinity chromatography on digitonin-sepharose.

An effective concentration of alpha-toxin from Staphylococcus aureus Wood 46, directly from the culture supernatant, could be achieved by adsorption on digitonin-sepharose and elution with 3 mol/l sodium thiocyanate (NaSCN). The toxin was further purified by gelchromatography. The purified product yielded 1 single protein band upon SDS-polyacrylamide electrophoresis. It was nonhemolytic, but reacted with anti-alpha-toxin under complement fixation. Dialysis against 0.14 mol/l NaCl with hydrophobic amino acids partially reactivated the alpha-hemolytic activity of the toxin. Ultracentrifugal analysis yielded sedimentation coefficients for the purified toxin of approximately 3,7 S when dissolved in 3 mol/l NaSCN and of about 12 S after dialysis against 0.14 mol/l NaCl (Table 1). The spontaneous oligomerization of the alpha-toxin during dialysis against 0.14 mol/l NaCl possibly resulted from a change in configuration induced by its adsorption to digitonin-sepharose.

Bacterial Toxins↗