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Infection and Immunity, April 1999, p. 1708-1714, Vol. 67, No. 4
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Protein H, an Antiphagocytic Surface Protein in
Streptococcus pyogenes
Britt-Marie
Kihlberg,*
Mattias
Collin,
Arne
Olsén, and
Lars
Björck
Section for Molecular Pathogenesis,
Department of Cell and Molecular Biology, Lund University, Lund,
Sweden
Received 19 June 1998/Returned for modification 26 August
1998/Accepted 17 December 1998
 |
ABSTRACT |
Surface-associated M protein is a major virulence factor in
Streptococcus pyogenes which confers bacterial
resistance to phagocytosis. However, many S. pyogenes strains also express additional structurally related so-called M-like proteins. The strain
studied here is of the clinically important M1 serotype and expresses
two structurally related surface proteins, the M1 protein and
protein H. Mutants were generated that expressed only one or none of
these proteins at the bacterial surface. For survival in human blood
either protein H or M1 protein was sufficient, whereas the double
mutant was rapidly killed. The protein-binding properties of protein H,
M1 protein, and the mutants suggest that bacterial binding of
immunoglobulin G and factor H or factor H-like protein 1, which are
regulatory proteins in the complement system, contribute to the
antiphagocytic property.
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INTRODUCTION |
Streptococcus pyogenes
(group A streptococcus) is an important human pathogen, with the
ability to cause a variety of infections. Primary disease
manifestations include pharyngitis, impetigo, and erysipelas, which may
lead to serious sequelae like rheumatic fever and glomerulonephritis
(7). Since the late 1980s there has been an increase in
severe invasive infections of skin and soft tissues (8),
associated with streptococcal toxic shock syndrome, with high mortality
(54). This type of severe infection has mainly been reported
to be associated with the serotypes M1 and M3 (22, 38, 41).
Clinical isolates of S. pyogenes grow rapidly in human
blood, a property associated with resistance against phagocytosis
(33). This antiphagocytic activity has been ascribed to the
expression of surface-associated M protein (for references, see
reference 15). There are more than 80 different
antigenically distinguishable M proteins, and immunity against
S. pyogenes is known to be M type specific. Each strain
was originally believed to express a single antiphagocytic M protein.
However, the M proteins have been shown to be members of a larger
family of structurally related proteins, the M-like proteins, that
includes three subtypes of proteins known as Mrp, Emm, and Enn
(29), and many S. pyogenes strains express
more than one M-like protein. The expression of these proteins is under
the control of the trans-acting regulator Mga (11, 30,
39, 47). Many M proteins with known antiphagocytic activities
belong to the Emm subfamily (14, 44, 50). Recently, Mrp
proteins also were shown to contribute to S. pyogenes
resistance against phagocytosis (48, 56). However, not all M
proteins have antiphagocytic activity, as demonstrated for Arp4, which belongs to the Emm subfamily (25).
The M and M-like proteins have affinity for several human plasma
proteins and often display different ligand-binding properties. For
instance, many of the proteins have affinity for factor H or
C4b-binding protein (C4BP) (24, 55), which are inhibitors of
complement. Both of these proteins have been suggested to contribute to
the resistance to phagocytosis (23, 56). Moreover, many Emm
and Mrp proteins bind fibrinogen, another human plasma protein that has
been implicated in the resistance to phagocytosis (14, 59).
Several M and M-like proteins also bind human immunoglobulin G (IgG)
and/or IgA through the constant Fc region (19, 20, 36, 53),
but the importance of the nonimmune interaction with Ig-Fc for the
virulence of S. pyogenes is unclear (12).
However protein H, an IgG-Fc-binding M-like protein, was shown to
inhibit complement activation at the bacterial surface, suggesting that Ig binding could contribute to resistance to phagocytosis
(6).
The S. pyogenes strain used in this study, AP1, is of
the clinically important M1 serotype. An insertional inactivation of Mga in AP1 rendered this strain sensitive to phagocytosis. The transcriptional regulator Mga in AP1 was shown to coregulate the expression of the M1 protein, protein H, protein SIC, and the C5a
peptidase (30). The proteins M1 and H are structurally
related and belong to the Emm subfamily, but these proteins have
different ligand-binding properties. Protein SIC has been shown to
interfere with complement-mediated cell lysis (4), and the
C5a peptidase is a cell wall-anchored enzyme with the ability to
inactivate C5a, a chemotactic factor of complement (58).
Here we evaluate the importance of the M1 protein and protein H for the
resistance to phagocytosis. This was done by generating mutants that
were selectively affected in the expression of one or both of these proteins, followed by analyzing the properties of the mutants.
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MATERIALS AND METHODS |
Bacteria, proteins, and plasmids.
S. pyogenes AP1
is strain 40/58 of the M1 serotype from the World Health Organization
Streptococcal Reference Laboratory in Prague and has a nonmucoid
morphology. S. pyogenes was cultured in Todd-Hewitt
(TH) medium supplemented with 0.2% yeast extract (THY medium) at
37°C. Luria-Bertani medium (51) was used for the culturing
of Escherichia coli. Unless otherwise indicated, antibiotics
were used at the following concentrations: tetracycline at 5 µg/ml,
streptomycin at 1,000 µg/ml, kanamycin at 25 µg/ml for E. coli and 500 µg/ml for S. pyogenes, and
erythromycin at 300 µg/ml for E. coli and 1 µg/ml for
S. pyogenes.
Human fibrinogen and human IgG3
were from Sigma (St. Louis,
Mo.). Purified human factor H was kindly provided by U. Sjöbring, and a rabbit antiserum against human factor H was a
generous gift from A. Sjöholm. Proteins were labeled with
125I by using the Bolton and Hunter reagent (Amersham,
Little Chalfont, England) or the chloramine-T method.
Plasmid pMC10 (
13) is a recombinant vector consisting of a
960-bp internal PCR fragment from
emm1, the gene encoding
the
M1 protein, ligated into the vector pFW13 (
49). To
generate
the truncated
emm1 sequence the oligonucleotide
primers 5'-ATAGAAGATCTAGAAGCAAACAAT-3'
and
5'-TCAGCTTTTTCTAGATCTGTTAATTTCTTG-3' were used in a PCR
experiment
with chromosomal DNA from AP1 as the template. After
XbaI digestion
of the PCR-generated
emm1 product,
this fragment was ligated into
the
SpeI site in the multiple
cloning site II of plasmid pFW13.
pBK37 was generated by the insertion
of an 803-bp internal PCR
fragment from
sph, the gene
encoding protein H, into pJRS233 (
46),
a derivative of the
temperature-sensitive plasmid pG
+host4 (
37). A
detailed description of pBK37 has been published
previously
(
6). pBH was generated by cloning the complete nucleotide
sequence for
sph, including promoter and termination
sequences,
into the
EcoRI site in pLZ12-21K (
45).
To generate the
sph sequence
the primers hybridizing to the
following sequences were used:
5'-GCTATCACTTTGTAATACTGAGTG-3'
and 5'-GTGACCTCTCCTTAACCTCATTC-3'.
Generation of S. pyogenes mutants.
Transposon mutants of S. pyogenes AP1 were generated by
Tn916 mutagenesis. Tn916 was transferred by
conjugation essentially as described previously (10), with
some modifications as described previously (30).
Transconjugants potentially lacking surface-associated M1 protein or
protein H were identified by colony blotting, as previously described
(30), by their reduced binding of fibrinogen and IgG3,
respectively. Chromosomal DNA from the transconjugants was digested
with HindIII and hybridized with a
Tn916-specific probe to analyze the number of transposons
that had been integrated.
Insertional inactivation of
sph in AP1, generating BM27.6,
has been described previously (
6). Briefly,
S. pyogenes was
transformed with the temperature-sensitive
recombinant plasmid
pBK37, which in BM27.6 was shown to be integrated
into
sph by
a single homologous recombination event
(
6). The insertional
inactivation of
sph in
BMJ11, generating BM22.1, was performed
in the same
way.
The insertional inactivation of
emm1 in AP1, generating
MC25, was performed as described previously (
13). Briefly,
the
emm1 mutant was generated by a single homologous
crossover recombination
event with plasmid pMC10. Recombinant clones
were selected by
plating on TH agar containing 150 µg of kanamycin
per
ml.
The bacterial strains used, the bacterial mutants generated, and the
plasmids used in this study are described in Table
1.
DNA preparation and other DNA techniques.
Chromosomal DNA
was prepared from S. pyogenes as previously described
(9), with the addition that the cells were treated with 500 U of mutanolysin (Sigma) per ml at 37°C for 2 h and then lysed
with 1% sodium dodecyl sulfate (SDS) and 0.2% Tween 20 on ice for
2 h.
Plasmid DNA preparations and other standard recombinant DNA techniques
were performed as described previously (
51). PCR
(
26) were performed with
Taq DNA polymerase
(Promega, Madison,
Wis.) in the presence of 1.5 mM MgCl
2
and 0.1 µM concentrations
of each primer (25 cycles were performed at
an annealing temperature
of 55°C and 1 to 3 min of extension at
72°C, depending on the
length of the product).
E. coli
JM109 was used to propagate plasmids
and was made competent for
transformation according to the procedure
of Nishimura et al.
(
42).
S. pyogenes was transformed by
electroporation
(
45).
Binding of human plasma proteins and analysis of protein
expression in S. pyogenes.
Bacteria to be analyzed in a
direct binding assay were grown overnight, washed twice in PBST (10 mM
sodium phosphate buffer, pH 7.2, containing 0.12 M NaCl and 0.05%
Tween 20), and resuspended in PBST to a concentration of 2 × 109 bacteria/ml. A serial dilution of bacteria (200 µl)
was mixed with 25 µl of 125I-labeled protein
(104 cpm) and incubated at room temperature for 1 h.
Cells were spun down, and the radioactivity associated with the pellet
was determined in a gamma counter. All samples were run in duplicate.
The absorption of human plasma proteins to S. pyogenes
was performed by incubating the bacteria in human heparinized plasma at
37°C as described previously (1). Plasma proteins bound to
the bacteria were eluted with 0.1 M glycine-HCl buffer, pH 2.0, and
then neutralized.
CNBr treatment was used to release surface-associated proteins M1 and
H, as previously described (
43). M1 protein released
into
the medium was precipitated with 70% (wt/vol)
(NH
4)
2SO
4,
and this material was
further purified on a fibrinogen-Sepharose
CL-4B column as described
previously (
1). Protein SIC was isolated
from the growth
medium of
S. pyogenes by precipitation with 30%
(wt/vol) (NH
4)
2SO
4, followed by gel
filtration as previously described
(
4). C5a peptidase was
solubilized from the bacterial surface
with streptococcal cysteine
proteinase as described previously
(
5).
SDS-polyacrylamide gel electrophoresis (PAGE) and Western blotting to
polyvinylidene difluoride membranes (Millipore, Bedford,
Mass.) were
performed according to standard methods (
32,
57).
Gels were
stained with Coomassie
blue.
Bactericidal assay.
The ability of S. pyogenes strains to survive in blood was tested essentially as
described previously (46). Cells grown to early mid-log
phase (A620 = 0.15) were serially diluted in TH
medium, and 100 µl of the bacterial solution was mixed with 1 ml of
heparin-treated blood from a donor lacking type-specific antibodies and
rotated end over end for 3 h at 37°C. In some experiments purified M1 protein was added to a concentration of 50 µg/ml. Samples
of 100 µl were withdrawn at indicated time points, added to 2.5 ml of
TH medium with 0.5% agar, spread on TH agar plates, and incubated at
37°C overnight. For strains complemented with plasmid pBH, the blood
and the plates were supplemented with kanamycin at a concentration of
500 µg/ml.
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RESULTS |
Generation and characterization of emm and
sph mutants.
Transposon mutagenesis of AP1 was used to
generate mutants affected in the expression of proteins H and M1. Two
types of mutants were generated with the Tn916 conjugative
transposon. The first type was deficient in the expression of both
proteins, and a detailed description and characterization of this
mutant have been published (30). The second type was
deficient in M1 protein, whereas the expression of protein H was
unaffected. One of the mutants, BMJ11, was selected for further
studies. Despite extensive screening, a mutant that had selectively
lost its ability to express surface-associated protein H could not be
detected. Such a protein H mutant, BM27.6, was therefore generated from
AP1 by homologous recombination (6). The same technique was
used to isolate the double mutant BM22.1 from BMJ11, which lacks both
surface-expressed M1 protein and protein H.
Further analysis of BMJ11 demonstrated that only a single transposon
was present in this mutant. However, the transposon in
BMJ11 was not
inserted either into or close to the M1 protein
gene (
emm1)
(data not shown). Another M1 protein mutant, named
MC25, was therefore
generated from AP1 by deleting the 3' end
of the
emm1 gene
by homologous recombination (
13), and the MC25
strain is the
M1 protein-negative mutant used in this
study.
The M1 protein expressed by AP1 has been shown to bind fibrinogen, and
it also has affinity for human IgG. Compared to protein
H, which does
not bind fibrinogen, the affinity for IgG is lower,
especially for the
IgG3 subclass (
2,
3). Therefore, the
binding of human
fibrinogen and IgG3 to intact AP1 bacteria was
used as a marker for
surface-expressed M1 protein and protein
H, respectively. A direct
binding assay demonstrated that MC25
had no affinity for fibrinogen,
whereas only a slight reduction
was seen in the affinity for IgG3 (Fig.
1). The
sph mutant BM27.6
bound fibrinogen, but the binding of IgG3 was significantly reduced
(Fig.
1). The reduction of IgG3 binding seen with MC25, as well
as the
residual IgG3 binding exhibited by BM27.6, is explained
by the affinity
between M1 protein and IgG (see above). The double-mutant
strain BM22.1
had no affinity for either IgG3 or fibrinogen (Fig.
1). This shows that
MC25 lacks surface-associated M1 protein,
whereas BM27.6 does not
express protein H. BM22.1, finally, does
not express M1 protein or
protein H at its surface. When BM27.6
was complemented with pBH, a
plasmid containing the
sph gene,
including promoter and
termination sequences, it regained its
ability to bind IgG3 (data not
shown). BM22.1
trans-complemented
with pBH bound IgG3 almost
as well as MC25 (Fig.
1). This shows
that the IgG3-binding property is
predominately associated with
the expression of protein H.
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FIG. 1.
Binding of human fibrinogen or IgG3 to S. pyogenes bacteria. 125I-labeled proteins were
incubated with indicated strains of bacteria at a concentration of
2 × 109 bacteria/ml. Data are means + standard
deviations of at least three independent binding experiments.
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Since the
trans-acting regulator Mga in AP1 is known to
coregulate the expression of M1 protein, protein H, protein SIC, and
the C5a peptidase (
30), the expression of the two latter
proteins
in the mutants was also studied. Protein SIC is an
extracellular
protein and can be purified from the growth medium
(
4), whereas
C5a peptidase can be released from the
streptococcal surface by
a cysteine proteinase produced by
S. pyogenes (
5). The same
amounts of protein
SIC and C5a peptidase could be purified from
AP1 and the mutants (data
not shown), demonstrating that the expression
of M1 protein and protein
H was selectively affected in the mutants
studied
here.
Characterization of protein expression in the mutants.
To
investigate whether the binding data described above correlate with the
expression of proteins M1 and H, we investigated the presence of
surface-associated proteins by cyanogen bromide (CNBr) treatment. This
treatment releases three protein fragments of 54, 49, and 44 kDa from
wild-type AP1 bacteria (Fig. 2, lane A).
The 54- and 49-kDa fragments originate from the M1 protein, and the
44-kDa band represents a protein H fragment (30). CNBr treatment of MC25 solubilized only the 44-kDa protein H-related fragment (Fig. 2, lane B). CNBr released the 54- and 49-kDa M1 fragments from BM27.6, whereas the 44-kDa band was missing (Fig. 2,
lane C), which is consistent with an insertional mutation of sph in this strain. CNBr treatment of BM27.6
trans-complemented with pBH released the same three protein
bands seen in extracts from wild-type AP1 bacteria (data not shown). No
protein band was released by CNBr treatment of the double mutant
BM22.1, whereas the 44-kDa fragment was seen after trans
complementation of this strain with pBH (Fig. 2, lanes D and E). These
results confirmed the binding data described above.
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FIG. 2.
Analysis of surface-anchored M1 protein and protein H. AP1 (A), MC25 (B), BM27.6 (C), BM22.1 (D), or BM22.1/pBH (E) bacteria
were treated with CNBr. Molecular mass markers are shown in lane F. Solubilized proteins were separated on SDS-10% PAGE gels followed by
Coomassie blue staining.
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In the MC25 strain the mutant M1 protein was expected to be exported
into the growth medium due to the absence of the COOH-terminal
cell
wall-anchoring LPXTGE amino acid motif. Therefore, growth
medium from
MC25 was purified on a Sepharose CL-4B column coupled
with human
fibrinogen. The eluted material contained a single
protein component
with a molecular mass of approximately 42 kDa
(Fig.
3). The molecular masses of M proteins
are overestimated
by SDS-PAGE (
21), and the value of 42 kDa
is therefore compatible
with the mass of 38 kDa predicted from the
sequence of the truncated
M1 protein.
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FIG. 3.
Analysis of M1 protein released into the growth medium
by MC25 bacteria. Culture supernatant from MC25 was affinity purified
on Sepharose CL-4B coupled with human fibrinogen. (A) Absorbance at 280 nm of eluted fractions. (B) Eluted fractions 4 to 11 were separated on
SDS-10% PAGE gels and stained with Coomassie blue. Lane M, purified
recombinant M1 protein. Molecular mass markers are indicated to the
left.
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|
Surface-expressed protein H is sufficient for the survival of AP1
in human blood.
The antiphagocytic property of S. pyogenes is associated with the type-specific M protein (for
references, see references 15 and
29). However, M-like proteins have also been shown to contribute to the antiphagocytic property (48, 56). The MC25 mutant expressed protein H but not M protein, and by using the
classical bactericidal test (34) it was found that MC25 survived in human blood (Table 1), although the rate of multiplication was reduced compared to that of the wild-type AP1 strain. This demonstrates that surface-associated protein H is sufficient for the
antiphagocytic property. MC25 secretes a fragment of the M1 protein
into the growth medium (Fig. 3). To investigate whether the soluble M1
protein fragment contributes to the resistance to phagocytosis, an AP1
mutant carrying a transposon in mga was tested in the
bactericidal assay. This mutant expresses no M or M-like proteins
(30), and it was rapidly killed in human blood also in the
presence of added purified M protein (data not shown), further
emphasizing that protein H alone is sufficient for the antiphagocytic
property. When BM27.6 was tested in the bactericidal test it was shown
that it survived and multiplied rapidly, whereas BM22.1 was killed
(Table 2). Also BM22.1/pBH survived in
human blood (Table 2) but multiplied less well than the parent strain BMJ11. Thus, it can be concluded that M1 protein and protein H both
contribute to resistance against phagocytosis.
Analysis of the binding of human factor H to the M1 protein and
protein H mutants.
The resistance of S. pyogenes
to phagocytosis has been linked to the expression of M protein and the
ability of this protein to bind fibrinogen and/or factor H (24,
59). MC25 has selectively lost its fibrinogen-binding activity.
However, soluble protein H, as well as protein M1, has been shown to
bind purified factor H (30), which could explain the
antiphagocytic property of MC25. Moreover, it is known that wild-type
AP1 bacteria absorb albumin, fibrinogen, and IgG from human plasma
(3). To investigate whether factor H could bind to AP1 and
the mutants also in plasma environment, the different strains were
separately incubated with human plasma. Following incubation,
bacteria were washed, and proteins bound to the surface were
eluted and subjected to SDS-PAGE and Western blot analysis with
antibodies against factor H (Fig. 4). The
most efficient absorption of factor H and its naturally occurring
splice variant, factor H-like protein 1 (FHL-1), was seen with
wild-type AP1 bacteria. All mutants also showed affinity for FHL-1,
whereas factor H was clearly detected in the eluates from mutant
bacteria expressing protein H (BM22.1/pBH and MC25). The data suggest
that protein H and M1 protein both contribute to the binding of factor H-FHL-1 to the surface of AP1 bacteria, but other surface components also show affinity. Finally, AP1 mutants that survive in fresh human
blood bind both factor H-FHL-1 and IgG (Table
3).
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FIG. 4.
Absorption of factor H (FH) and FHL-1 from human plasma
by wild-type AP1 bacteria and various AP1 mutants. Bacteria were
incubated with human plasma, and the proteins absorbed were eluted and
subjected to SDS-10% PAGE, followed by electrotransfer to a
polyvinylidene difluoride membrane. The blot was incubated with
antibodies against factor H, and the antibodies were visualized with
peroxidase-labeled protein A.
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| |
DISCUSSION |
We have previously shown that the insertional inactivation of the
regulator gene mga in the AP1 strain of the M1 serotype resulted in the loss of its antiphagocytic property (30).
Most opacity factor-negative S. pyogenes strains
express a single Emm protein (29), whereas AP1 bacteria
express two structurally related Emm proteins, the M1 protein and
protein H, with different ligand-binding properties (3, 17,
18). The present work demonstrates that these proteins both
contribute to the phagocytic resistance. Mutants expressing either the
M1 protein or protein H multiplied rapidly, whereas the double mutant
lacking both proteins was killed in human blood. The double mutant
trans-complemented with protein H on a plasmid (BM22.1/pBH)
regained the ability to survive in human blood. However, BM22.1/pBH
multiplied less well than the other mutants. This could be due to the
insertion of several extra DNA segments. Both double-mutant
bacteria and BM22.1/pBH grew somewhat slower in THY media than the
other strains studied. The fact that BM22.1/pBH, but not the double
mutant, is resistant to phagocytosis also indicates that proteins M1
and H are the only factors expressed by AP1 that are
directly involved in the resistance to phagocytosis in human blood.
The M and M-like proteins are structurally related and are coexpressed
but often display different ligand-binding properties. Our results, in
line with the results of previous studies (48, 56),
demonstrate that apart from the type-specific M protein, other M-like
proteins also can be involved in the resistance to phagocytosis.
Opacity factor-positive S. pyogenes strains express M-like proteins known as the Mrp proteins (29). The Mrp
protein, as well as the coexpressed M protein, has been shown to be
antiphagocytic in several strains (48, 56). However, the
abilities of the individual proteins to confer resistance against
phagocytosis seem to vary. Furthermore, not all M-like proteins are
antiphagocytic. The IgA-binding protein Arp, an M-like protein
expressed by an M4 strain (35), does not have this property
(25). Attempts to complement heterologous nonresistant
S. pyogenes JRS145, used by Husmann et al.
(25), with pBH did not succeed. JRS145 could be transformed
with pBH, but the recombinant strain expressed very small amounts of
protein H, and this transformed strain was killed by human phagocytes
(30a).
It has been demonstrated that surface-associated protein H can block C3
deposition on the bacterial surface, due to the inhibition of the
classical complement pathway. This inhibitory activity of protein H was
dependent on its ability to bind IgG (6). A significantly
higher amount of C3 deposition was measured on the mutant of AP1 that
expresses the M1 protein but lacks protein H (BM27.6) than on wild-type
AP1 (6). However, despite the observed C3 deposition, BM27.6
was found to be resistant against phagocytosis in this study. Protein H
has also been shown to bind C4BP (28), an inhibitor of the
classical complement pathway, and purified protein H and M1 protein
both bind factor H (30), another inhibitor of complement.
Moreover, it was recently demonstrated that FHL-1, a splice variant of
factor H (40), interacts with M5 and M6 proteins
(31) by binding to their hypervariable
NH2-terminal regions, whereas the factor H-binding site is
separate (27) and located further towards the COOH-terminal
end (16, 27, 52). In the case of M5 protein, fibrinogen was
found to inhibit the binding of factor H (27). The binding
of fibrinogen to M1 protein could therefore explain the low degree of
absorption of factor H from plasma by the protein H-negative mutant
expressing M1 protein. Among the tested strains, wild-type AP1 bacteria
absorbed both factor H and FHL-1 most efficiently. However, the fact
that the mutants, including the double mutant, still had FHL-1-binding activity demonstrates that AP1 bacteria have surface structures apart
from protein H and M1 protein that also mediate FHL-1 binding. Thus,
the relative importance of the binding of factor H, FHL-1, C4BP, and
IgG for phagocytosis resistance remains unclear.
There is no doubt that the molecular basis for the antiphagocytic
property of S. pyogenes is highly complex.
Nevertheless, some conclusions can be made from the present
observations, at least concerning the M1 strain used in this study.
Firstly, proteins H and M1 are both antiphagocytic, and the
presence of one of them at the bacterial surface is sufficient for the
survival of bacteria in human blood. Secondly, the binding of
fibrinogen to the bacterial surface is not required. Thus, the M1
protein mutant (MC25) devoid of fibrinogen-binding activity still
survived. Similar observations were made for a strain of the M22
serotype (56). In contrast, fibrinogen binding was found to
be necessary for the antiphagocytic activity of M proteins of some
other serotypes (14). Thirdly, the surviving mutants bind
both factor H-FHL-1 and IgG, suggesting that the presence of these host
proteins at the surface of the M1 strain used here is crucial for its
antiphagocytic property. As mentioned above, it has been demonstrated
that the binding of IgG to the streptococcal surface through
interaction with the Fc region of IgG inhibits the activation of the
classical pathway (6), whereas factor H binding interferes
with the activation of the alternative pathway of complement
(24). Therefore, the present data suggest that the
antiphagocytic property of S. pyogenes could be based
on the interference with both complement pathways.
| |
ACKNOWLEDGMENTS |
The excellent technical assistance of Ingbritt Gustafsson and
Ulla Johannesson is acknowledged.
This work was supported by grants from the Swedish Medical Research
Council (projects 7480 and 13062), the Foundations of Crafoord,
Kock, Lundberg, Nilson, and Österlund, the Göran Gustafsson Foundation for Research in Natural Sciences and Medicine, ACTINOVA Ltd., and the Medical Faculty, Lund University.
| |
FOOTNOTES |
*
Corresponding author. Present address: Department of
Cell and Molecular Biology, Umeå University, S-901 87 Umeå, Sweden.
Phone: 46-90-7852689. Fax: 46-90-771420. E-mail:
Britt-Marie.Kihlberg{at}cmb.umu.se.
Editor:
V. A. Fischetti
| |
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Abstract
Introduction
