Streptococcal opacity factor: a family of bifunctional proteins with lipoproteinase and fibronectin-binding activities.

The serum opacity factor (SOF) of Streptococcus pyogenes is a type-specific lipoproteinase of unknown biological significance. We have sequenced the sof gene and characterized the corresponding SOF protein from a strain of type M63. It was found that sof63 is related to sof22 and that, similar to SOF22 [25], SOF63 binds fibronectin. Moreover, we demonstrate opacity factor activity in a Streptococcus dysgalactiae fibronectin-binding protein FnBA that is structurally related to the SOF proteins of S. pyogenes. Sequence analysis of these three SOF proteins showed a unique periodical pattern of conserved and variable regions. The enzymatically active part of SOF63 was localized to the fragment corresponding to the entire set of conserved and variable sequences, while for fibronectin-binding a single repeat in the C terminal part of the protein was sufficient. The results show that streptococcal SOF proteins form a novel family of bifunctional proteins with lipoproteinase and fibronectin-binding activities.

Sequencing of genes within the vir regulon of Streptococcus pyogenes type M15--an opacity factor-positive serotype with low opacity factor expression.

Major virulence determinants of group A streptococci, such as M-protein, immunoglobulin Fc-receptors (FcRA, EmmL) and C5a peptidase, appear to be genetically co-regulated, their genes being located within a vir regulon. We studied the organization of these genes in a group A, type M15 strain of Streptococcus pyogenes, previously defined as OF-, by hybridization analysis of chromosomal DNA and of an S. pyogenes gene library in Escherichia coli, and by gene sequencing. Within the vir regulon, in addition to the virR and scpA genes, three so-called emm-related genes were found: fcrA, emmL and enn. Whereas IgG Fc-binding proteins were encoded by fcrA and emmL, the product of enn was not identified. The presence of three emm-related genes in this region is reminescent of vir regulon organization in OF+ rather than OF- strains as earlier defined by others. Furthermore, analysis of the deduced product of the emmL gene showed deletions and amino acid substitutions within the PGTS-rich domain and membrane anchor, which thus resembles corresponding products of OF+ rather than OF- strains. In view of these findings, the opacity factor (OF) activity of the strain was tested using growth supernatant, with negative outcome. However, a concentrated SDS cell extract revealed definite OF activity. One of two other type M15 reference strains also showed definite OF activity in SDS extracts. We therefore propose that type M15 strains belong to the OF+ category but often show low levels of expression of OF.

Haemolytic and phospholipase C (PLC) activities of Rhodococcus equi.

Rhodococcus equi, an intracellular organism causing pneumonia and lung abscesses in foals, is generally thought to be non-haemolytic. In the present study, however, 13 of 14 representative isolates were found to be haemolytic when tested on agar media containing washed red blood cells rather than whole blood. Red cells of rabbits, dogs, horses and man were more sensitive to lysis than were those of ruminants. Two new enzymatic activities of the species were defined: a lecithinase and a phosphatidylinositol-specific phospholipase C (PI-PLC). As judged from tests for trypsin, temperature and ethanol sensitivity, the haemolytic activity was primarily dependent on PI-PLC though the participation of lecithinase seemed probable. The haemolytic activity of growing strains, but not of cell-free preparations, was partially inhibited by lecithin but enhanced by cholesterol; however, cholesterol oxidase (CO) activity, known to mediate cooperative lysis of RBC sensitized with sphingomyelin-specific phospholipases C or D of some other species, did not contribute to the direct haemolysis caused by R. equi as demonstrated here.

M-like, immunoglobulin-binding protein of Streptococcus pyogenes type M15.

An M-like protein from Streptococcus pyogenes type M15 strain EF1949 (EMML15) was cloned in Escherichia coli and sequenced. Recombinant EMML15 protein revealed a unique binding pattern for human IgG subclasses not described previously. Comparative analysis of the EMML15 amino acid sequence with those of other M-like proteins of opacity factor positive (OF+) serotypes and protein H, and IgG receptor from OF- serotype M1, showed that IgG-binding proteins with common binding of IgG3 were closely related and distinct from streptococcal IgG receptors not binding IgG3. Thus, the Ig-binding proteins from S. pyogenes were subdivided into two main categories according to binding pattern, protein structure, and gene location.

[The cloning and expression of the gene for Staphylococcus aureus beta-hemolysin]. / Klonirovanie i ékspressia gena beta-gemolizina Staphylococcus aureus.

The gene coding the synthesis of beta-hemolysin (phospholipase C) has been cloned from S.aureus strain 126/89. A highly active Escherichia coli producer has been obtained, its capacity of synthesizing phospholipase C exceeding that of the natural strain 37-fold. Phospholipase C (beta-hemolysin) obtained from the natural and gene-engineering producers have been found to be identical in their enzymatic and molecular characteristics. The nucleotide sequence of the full plc gene which codes peptide consisting of 333 acid radicals has been established. In S.aureus strains of different origin this gene has been found to have a conservative character.