J. gen. Virol. (1986), 67, 2721-2729. Printedin Great Britain
2721
Key words: RS virus~antigenicvariation~structuralpolypeptides
Structural Differences between Subtype A and B Strains of Respiratory
Syncytial Virus
By E R L I N G N O R R B Y , 1. M A U R I C E A. M U F S O N 2
AND H O O S H M A N D S H E S H B E R A D A R A N 1
1Department of Virology, Karolinska Institute, School of Medicine, SBL, S-105 21 Stockholm,
Sweden and 2Department of Medicine, Marshall University School of Medicine and Veterans
Administration Medical Center, Huntington, West Virginia, U.S.A.
(Accepted 28 August 1986)
SUMMARY
Differences in the properties of homologous intracellular structural components of
eight strains of subtype A and eight strains of subtype B of human respiratory syncytial
(RS) virus were examined. The size of the fusion (F) protein cleavage products and the
phosphoprotein (P) showed systematic differences between virus strains representing
the two subtypes. The apparent mol. wt. in SDS-polyacrylamide gels under reducing
conditions was 48000 (48K) and 46K to 47K for the cleavage product FI in subtype A
and B strains, respectively. The size of the F2 protein was 18K to 20K. The subtype B
strains showed a slightly higher tool. wt. of this protein compared to the subtype A
strains. The size of the P protein was 36K in subtype A strains, but only 34K in subtype
B strains. Variations also occurred in the size of the glycoprotein (G) and the 22K to
24K structural protein. These variations did not correlate with the virus subtypes, but
were strain-specific. The size of non-glycosylated forms of the F protein cleavage
products was determined by use of material from tunicamycin-treated cells. A 44K to
45K non-glycosylated form of the F 1 protein was detected with subtype A virus strains,
but the corresponding protein of subtype B strains was not reproducibly identified,
presumably due to instability in the absence of glycosylation or altered antigenicity.
Monoclonal antibody immunosorbent-bound viral glycoproteins were partially
digested with proteases. The pattern of breakdown products of the F1 protein was
distinctly different between subtype A and B strains, but it was similar among strains of
the same subtype. No subtype-specific pattern was seen in proteolytic digests of
monoclonal antibody-bound G protein.
INTRODUCTION
For the 3 decades since its discovery human respiratory syncytial (RS) virus has been
considered to represent a single homogeneous serotype. Although cross-neutralization tests with
animal hyperimmune sera demonstrated differences between isolates of the virus (Coates et al.,
1963; Wulffet al., 1964; Doggett & Taylor-Robinson, 1964) no corresponding differences were
observed with infant sera and they were therefore not considered to be of any relevance. With
the advent of monoclonal antibodies (MAbs) the antigenic stability of RS virus could be
assessed again. Studies with such reagents revealed that isolates of RS virus exhibited variations
in epitope occurrence (Gimenez et al., 1984) but that these variations were more extensive than
could be explained by incidental mutational changes (Anderson et al., 1985; Mufson et al.,
1985). The isolates could be separated into two categories based on their characteristic reaction
pattern with different MAbs. The two groups were named RS virus subtype A and B (Mufson et
al., 1985). These subtypes showed different properties in at least four different structural
proteins emphasizing that they had become ecologically adjusted after a period of separate
evolution. Characterization of the epidemiological occurrence of the two RS virus subtypes
0000-7332 © 1986 SGM
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2722
E. NORRBY, M. A. MUFSON AND H. SHESHBERADARAN
confirms that they may circulate concurrently, but that both temporal and geographical
clustering can be distinguished (Hendry et al., 1986; Akerlind & Norrby, 1986).
Previous studies have shown that different strains of RS virus have phosphoproteins (P) of
different sizes (Cash et al., 1977). The occurrence of such differences was confirmed in this study
and the size variation could be correlated with subtype A or B characteristics of different virus
strains. In addition we observed distinct structural differences between other cell-associated
components of RS virus subtype A and B strains by use of physicochemical methods. This
presentation describes subtype-specific differences in the size of homologous structural proteins
of both the fusion (F) protein cleavage products and P protein, the stability of the F glycoprotein
in a non-glycosylated form, and partial protease digestion patterns of MAb-anchored virus F
glycoprotein.
METHODS
Virus strains. The subtype A strains included the prototype laboratory strains A2 and Long, the strain CH287
which was evaluated as a parenterally administered live vaccine (Belshe et al., 1982), the strains WV9894 and
WV12138 characterized in a previous study (Mufson et al., 1985) and three strains (V214, V316, V401) isolated in
Bristol, U.K. between 1983 and 1984 and provided by Dr E. J. Stott, Institute for Research on Animal Diseases,
Compton, Newbury, U.K. The subtype B strains included two strains isolated during the early 1960s, CH18537
(Coates et al., 1963) obtained from Dr R. M. Chanock, NIAID, National Institutes of Health, Bethesda, Md.,
U.S.A. and a Swedish strain, 8/60 (Doggett & Taylor-Robinson, 1964), obtained from Dr E. J. Stott (both have
been defined as B subtypes; E. Norrby, unpublished), the strains WV1293, WV3212, WV4843 and WV6873
characterized previously (Mufson et al., 1985) and two strains (V422 and V463) isolated in Bristol between 1983
and 1984 and provided by Dr E. J. Stott. The LEC strain of measles virus (see Sheshberadaran et al., 1983) was
used as a control in experiments with tunicamycin-treated cells.
Cells. Veto and HeLa cells were grown in Eagle's MEM supplemented with 3 ~ heat-inactivated (56 °C for
30 min) foetal calf serum. The cells were maintained in medium with 1 ~ serum.
Antibody-containing samples. Ascites materials prepared by injection of hybridoma cells producing RS virusspecific MAbs described in an earlier publication (Mufson et al., 1985) were used. Antibodies with specificities for
the glycoprotein (G), F, nucleocapsid (NP), P and matrix (M) proteins were employed. In addition two MAbs
specific for the 22K to 24K envelope-associated protein (Huang et al., 1985) were kindly provided by Dr G. L.
Toms, Department of Virology, University of Newcastle Upon Tyne, U.K. and human convalescent sera from
children with serologically defined RS virus infections were obtained from the National Bacteriological
Laboratory, Stockholm, Sweden.
Radioimmune precipitation assay (RIPA). Viral structural proteins were labelled for 24 h with [3sS]methionine or
[3H]glucosamine (50 ~tCi/ml of medium) prior to development of extensive cytopathic effects in Vero and HeLa
cells. Cells were scraped into the supernatant, pelleted, washed twice with cold phosphate-buffered saline and
dissociated on ice with a buffer containing 2% Triton X-100, 0.15 M-NaC1, 0.6 M-KC1, 0.5 mM-MgC12, 1 mMPMSF and 1% trasylol aprotinin in 0.01 M-Tris-HC1 pH 7.8 (RIPA buffer). The supernatant was collected,
clarified in an Eppendorf centrifuge and then stored at - 2 0 ° C until used. The antigen preparation was
precipitated with MAbs in the form of ascites fluid and fractionated by SDS PAGE under reducing conditions.
The details of these procedures were described previously (Orvell & Norrby, 1980; Sheshberadaran et al., 1983).
Proteinfootprinting. This method in brief was as follows. Isotope-labelled antigen and MAb were allowed to bind
on ice for at least 2 h and thereafter the immunocomplexes were precipitated with Staphylococcus aureus Protein
A-Sepharose CL-4B beads. The beads were washed three times with RIPA buffer and twice with TBS (0.01 MTris HCI, 0.15 M-NaCI pH 8.0) containing 1.2% Triton X-100 and 0.1% SDS. The excess buffer was removed
from the pelleted beads and 30 ~tl of stock trypsin or V8 protease solution (1 mg/ml in H20) was added per sample.
The samples were held at 37 °C with occasional agitation for the time periods stated in the text. At the end of the
digestion samples were placed on ice and washed twice with RIPA buffer and once with TBS. Excess buffer was
removed and beads were dried at 37 °C. Samples were analysed by SDS-PAGE under reducing conditions unless
otherwise stated.
The above procedure was employed for anti-F protein MAb footprinting. For G protein footprinting the
procedure was modified because the single available anti-G protein MAb reacting with both subtypes was
degraded by the proteases too rapidly. Prior to addition of protease, 20 i11 of undiluted rabbit anti-mouse
immunoglobulins was added per sample and the beads were held at 4 °C for 1 h and occasionally agitated. Protease
was then added and digestion performed as described above. At the end of the digestion period, 20 ~tl of fivefold
concentrated SDS-PAGE reducing sample buffer was added to each sample which was then held at 100 °C for
3 min. Samples were analysed by SDS-PAGE without further treatment.
Isotopes, molecular weight markers and chemicals. [3sS]Methionine (sp. act. 1000 to 1200 Ci/mmol), I>[1,6Downloaded from www.microbiologyresearch.org by
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Differences between RS virus subtypes
2723
3H(N)]glucosaminehydrochloride (sp. act. 31 to 33 Ci/mmol), and reference 14C-methylatedprotein mixture were
purchased from New England Nuclear. The reference protein mixture contained phosphorylase b (97400 mol.wt.,
97-4K), bovine serum albumin (69K), ovalbumin (46K), carbonic anhydrase (30K), lactoglobulin A (18.4K) and
insulin (5-8K). S. aureus V8 protease, TPCK-treated trypsin, tunicamycin, PMSF and trasylot aprotinin were
purchased from Sigma, Protein A-Sepharose CL-4B beads were bought from Pharmacia, and the rabbit antimouse immunoglobulins from Dako, Copenhagen, Denmark.
RESULTS
Sizes of cell-associated structural proteins of RS virus subtypes A and B strains in S D S - P A G E
R I P A with [35S]methionine-labelled virus strains incubated with polyclonal h u m a n
convalescent sera and a M A b specific for the F protein is illustrated in Fig. 1 (a). No variations
in apparent mol. wt. in S D S - P A G E of the N P and M proteins of six virus strains representing
different subtypes was found, but there was subtype-specific variation in the size of the F
proteins. Thus the size of the F 1 protein was 48K in all eight subtype A strains, but only 46K to
47K in all eight subtype B strains (see also Fig. 2, Table i). Further, the characteristics of lower
mol. wt. F cleavage polypeptides also differed (Fig. 1 a and tests with anti-F MAbs in Fig. 2 and
Fig. 3). In subtype A strains two bands were seen in the 18K to 23K range. As is shown below
(Table 1) these were apparently the 18K to 20K bona fide F2 protein and a 23K non-glycosylated
fragment of the F1 protein. In subtype B strains only one band (18.5K to 20K) was visible in this
(a)
(b)
(c)
WV4843 CH18537
II
A B A B
A B R
A
B A B
1
2
3
I
1
2 3
~---97K
97K
",~---69K
69K
-~---46K
G
~--69K
NP
F1
46K
,~--97K
P
NP
",~---30K
~--46K
22K-24K
M
30K
~----18K
-~---30K
F2
Fig. 1. Comparison of the size of homologous proteins of RS virus subtype A and B strains in RIPA.
Viral proteins were labelled in Vero cells with psS]methionine (a and b) and in HeLa cells with
[3H]glucosamine(c). (a) The four samples to the left [virus strains CH287 and WV 12138 (subtype A),
and CH 18537 and WV4843 (subtype B)] were precipitated with a human convalescent serum and the
samples on the right (virus strains WV 12138, WV4843)with an anti-F MAb (B46). (b) The two left lanes
contain one subtype A strain (CH287) and one subtype B strain (WV3212) precipitated with an anti-P
MAb (C771) and the two right lanes contain the corresponding antigen preparations precipitated with
an anti-22K-24K MAb (5H5). The 41K band represents co-precipitated NP protein and the low tool.
wt. band in the two left lanes represents a P protein breakdown product or co-precipitated 22K-24K
protein. (c) Two subtype B strains labelled on three different occasions and precipitated with an anti-G
MAb (C793). Samples were analysed on 10% (a, c) or 12.5~ (b) gels under reducing conditions. Lane R
shows mol. wt. markers in (a) and arrows indicate the position of these markers in (b) and (c).
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2724
E. N O R R B Y , M. A. M U F S O N A N D H. S H E S H B E R A D A R A N
(a)
(b)
A
B
A
B
97K
69K
F1
46K
30K
F2
18K
Fig. 2. Comparison of the size of F cleavage products of a subtype A strain (CH287) and a subtype B
strain (WV4843) grown in ¥ero cells in the absence (a) and presence (b) of 15 p.g/ml of tunicamycin.
Samples were analysed in a 12.5% gel under reducing conditions. Mol. wt. markers are indicated by
arrows.
Summary of size characteristics of homologous structural proteins of RS virus subtype A
and B strains, including the size of the non-glycosylatedform of the F1 protein and of different
proteolytic breakdown products of this protein
T a b l e 1.
Molecular weight
f
Component
G
F1
F1, non-glycosylated
F1, trypsin breakdown fragments
F2
NP
P
M
22K-24K
A
RS virus subtype A
90K*
48K
44K-45K
32K, 23K
18K-20K
41K
36K
26K
23K-24K*
RS virus subtype B
90K*
46K-47K
(43K-44K)t
30K, 21K
t8.5K-20K
41K
34K
26K
22K-24K*
* Certain strain-specific variations.
, Detected in only one out of four experiments.
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Differences between RS virus subtypes
2725
mol. wt. range but this band contains two polypeptides which migrate closely (see below). One of
these, the F2 glycoprotein, in repeated experiments had a marginally higher tool. wt. than the
corresponding protein in subtype A strains (not readily observable in Fig. 1 a or by comparison
of lanes 1 to 3 and 4 to 5 in Fig. 3 b, but better seen in Fig. 2). Other consistent differences were
detected in the size of the P protein in four strains representing each subtype (exemplified in Fig.
1b); 36K in subtype A virus strains but only 34K in subtype B virus strains (Table 1).
Occasional variations were seen also in the size of other proteins in different strains, but they
did not correlate with the subtype A and B classification. The 22K to 24K protein of unknown
function had an apparent size of 22K in the subtype B strain WV3212, but in another subtype B
strain, WV4843, as well as in two subtype A strains (A2 and CH287), it had a size of about 23K
to 24K (exemplified in Fig. 1 b). The relative size of the G protein was compared in RIPAs with
[3H]glucosamine-labelled HeLa cell material with five strains each representing subtype A and
subtype B strains. There was a tendency for the broad G band to migrate slightly more slowly in
subtype A strains than in subtype B strains, but one subtype B strain, WV4843, had a G protein
similar in size to that of most subtype A strains (exemplified in Fig. 5 a). This difference in size of
the G proteins of WV4843 and another subtype B strain was seen in three consecutive
experiments (Fig. 1c). Therefore the size of the G protein may be strain- but not subtypespecific.
Relative size in SDS-polyacrylamide gels of non-glycosylated F protein cleavage products of RS
virus subtypes A and B
The F protein of RS virus contains N-linked oligosaccharides as evidenced by the effect of
treatment of infected cells with tunicamycin (Gruber & Levine, 1985). Since the size of the F
protein cleavage products differs between subtype A and B strains it was considered of interest
to compare the relative size of these proteins in their non-glycosylated form. Cells showing
partial cytopathic effects were transferred to [3SS]methionine medium containing 5 or 15 ~tg/ml
of tunicamycin or as control without addition of the drug. Cell harvests were made 24 h later.
For comparison Vero cell cultures infected with the LEC strain of measles virus were
maintained on the corresponding medium with and without tunicamycin. The virus-specific
polypeptides were identified by immunoprecipitation with MAbs. The measles virus
haemagglutinin was reduced in relative size from 79K to 65K or 66K (not illustrated) by
treatment with tunicamycin as described previously (Bellini et al., 1983), but in the presence of
even the lower concentration of tunicamycin, 5 ~tg/ml, no non-glycosylated forms of the 40K and
18K measles virus F1 and F2 proteins were detectable. This was interpreted to reflect an
accentuated sensitivity to proteolysis of these forms of the F products.
The effect of tunicamycin treatment on the F cleavage products of RS virus differed for the
subtype A and B strains. A distinct 44K to 45K non-glycosylated form of the F1 protein was
detected in cells infected with subtype A virus strains (Fig. 2) confirming previously published
data (Gruber & Levine, 1985). However, in cells infected with subtype B strains one out of four
experiments revealed a low concentration of a 43K to 44K protein, but in the three other
experiments no non-glycosylated F cleavage products could be discerned as in the case of
measles virus F protein in tunicamycin-treated cells. Fig. 2 also shows a weak protein band with
a high migration rate ( < 10K) in tunicamycin-treated cells infected with subtype A RS virus.
This protein may represent the non-glycosylated F2 protein, but it may also represent a
degradation product of the F1 protein. In most experiments with subtype A strains this low
molecular weight protein was not seen and it was never observed in tunicamycin-treated RS
subtype B virus-infected cells.
Footprinting of F and G glycoproteins : proteolytic cleavage of MAb-bound antigen
MAb immunosorbent-bound F protein was partially digested with protease and washed to
remove unbound fragments. [3sS]Methionine- and [3H]glucosamine-radiolabelled F protein
from subtype A and B strains were compared. The anti-F MAbs reacting with epitope 1 on the F
protein (Mufson et al., 1985) generated the digest pattern shown in Fig. 3 in which the major
MAb-bound Fl-derived fragment was 32K in subtype A and 30K in subtype B strains. The
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2726
E. N O R R B Y , M. A. M U F S O N A N D H. S H E S H B E R A D A R A N
(b)
(a)
ul
u2
1
2
3
4
5
6
1
2
3 4
5
ul
u2
59K
F1
46K
30K
J
:lgll
5.8K
Fig. 3. Footprinting of MAb B46-bound F protein using [35S]methionine- (a) or [3H]glucosamine- (b)
radiolabelled antigens from virus-infected Vero cells. Lanes u show RIPA of control undigested
material of subtype A strain CH287 (ul) and subtype B strain CH18537 (u2). Footprinting was
performed using trypsin (digestion time 2 h) on subtype A strains A2 (lanes 1), CH287 (lanes 2),
WV9894 (lanes 3) and on subtype B strains WV9894 (lanes 4), WV6873 (lanes 5) and CH 18537 (lane 6;
no glucosamine-labelled sample included). Mol. wt. markers are indicated by arrows. Samples were
analysed on 10 to 20 ~ gradient gels under reducing conditions.
molecular weight difference in the undigested F1 of the two subtypes therefore was maintained
in this fragment. This 30K/32K F1 fragment was non-glycosylated in both subtypes.
The single anti-F MAb which reacts with a second, distinct epitope (B151 ; Mufson et al.,
1985) generated a slightly different F digest pattern. This major Fl-derived fragment was a nonglycosylated fragment ofmol, wt. 23K in subtype A and 21K in subtype B strains (Fig. 4a). This
23K fragment was seen also in RIPA with undigested F protein of subtype A strains, but its
equivalent in subtype B strains (21K fragment) was commonly masked in such RIPAs due to its
close migration with subtype B F2 (Fig. 4b). As with the 32K/30K F1 fragment, the molecular
weight difference between the undigested F1 of the two subtypes was maintained in the
23K/21K fragment (Table 1). In addition to this fragment 13K fragments were generated.
Further, the 30K fragment was seen with MAb B 151 in subtype B strains, but the corresponding
32K fragment of subtype A strains was absent. The MAb-bound F digests were examined also
by non-reducing SDS-PAGE. The results (Fig. 4b) indicate that the 32K/30K, 23K/21K and F2
proteins are disulphide-bonded in both subtypes (data for a subtype B strain not illustrated).
Comparison of partial proteolytic digests of MAb bound G protein of the two subtypes did not
reveal any subtype-specific pattern as seen for the F protein (Fig. 5).
DISCUSSION
The results of this study further emphasize the distinctive nature of RS virus subtypes A and
B. Previously it was shown by use of monoclonal antibodies that subtypes A and B differed in the
antigenic properties of at least four structural proteins (Mufson et al., 1985). The present study
demonstrates that distinct differences exist in the apparent size in SDS-PAGE of the
homologous cell-associated F1 and P proteins of RS subtype A and B viruses. Furthermore the
footprinting analysis by proteolysis of MAb-bound anti-F proteins shows patterns of breakdown
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Differences between R S virus subtypes
2727
(b)
(a)
CHl8537
A2
1 I
2
u
1
2
MET
GLU
I
i
I
R
u
1
2
r
1
u
1 2
~TK
97K
69K
FI
46K
F1
30K
F2
18K
6K
10K
F2
5-8K
8K
i.8K
Fig. 4. Footprinting of MAb151-bound F protein. (a) [3sS]Methionine-labelledsubtype A strain A2 and
subtype B strain CH18537. (b) [3H]glucosamine(GLU)- and [35S]methionine (MET)-labelled subtype
A strain CH287. All labelling was performed in Vero cells. In all panels lanes u show the undigested
control material, lanes 1 show footprints after digestion for I h and lanes 2 after digestion for 6 h with
trypsin. Lane R in (a) shows tool. wt. markers (see text) and arrows indicate the positions of these
markers in (b). Sampleswere analysedon 10 to 20~ gradient gels under reducing conditions except the
MET samples in (b), which were run without addition of mercaptoethanol.
products characteristic of subtype A and B viruses. In studies of one RS virus strain from a local
epidemic, Ward et al. (1983) noted that their anti-F MAb immunoprecipitated a nonglycosylated 23K protein in addition to the glycopolypeptides F1 and F2. Judging from the
present data it is likely that the epidemic studied was caused by a subtype A virus strain.
As with other paramyxoviruses, both surface glycoproteins G and F may play a role in
immune protection (Walsh et al., 1984a, b, 1985). However, it appears that the involvement of
RS virus F protein is relatively more important both for neutralization phenomena in vitro and
for events restricting replication in vivo. It remains to be determined whether the difference
between the homologous and heterologous neutralization titres of strain-specific polyclonal
hyperimmune sera (Coates et al., 1963; Wulff et al., 1964, Doggett & Taylor-Robinson, 1964)
which exemplify differences between subtype A and B strains (E. Norrby, unpublished data)
reflect antigenic differences between G or F components. Since it is more likely that the F
antigen is involved, the finding in this study of distinct differences between the F components of
subtypes A and B assume a special significance.
The identification of antigenic fragments after proteolytic digestion may allow dissection of
structural-functional relationships of the F component with reference to subtype-unique and
subtype-common antigenic sites. Information on the primary structure of the F polypeptide
from cloning of RS virus subtype A genes (Collins et al., 1984; Elango et al., 1985) and
forthcoming similar data on the F gene of subtype B strains will be of considerable value in this
context.
Concerning the cell-associated G protein, no reproducible size differences between subtype A
and B strains (five of each were examined) were found, although in the majority of strains the
protein of subtype A strains was larger. The footprinting experiments using proteolysis of MAbbound antigen also did not reveal any uniform pattern characteristic of this protein in subtype A
and B strains. The protein part of the G protein is a 33K polypeptide (Satake et al., 1985; Wertz
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E. NORRBY, M. A. MUFSON AND H. SHESHBERADARAN
(a)
(b)
1
2
3
4
5
1
2
3
4
-,~----97K
,~--69K
(
46K
5
--97K
--69K
46K
30K
Fig. 5. Footprinting of MAb C793-bound G protein. [3H]Glucosamine-labelled virus antigen grown in
HeLa cells was used. Subtype A strains included CH287 (lanes 1) and WV9894 (lanes 2) and subtype B
strains WV6873 (lanes 3), CH18537 (lanes 4) and WV4843 (lanes 5). Samples in (a) were untreated and
in (b) digested for 2-5 h with V8 protease. Mol. wt. markers are indicated by arrows. Samples were
analysed on 10 to 20% gradient gels under reducing conditions.
et al., 1985) and the 90K apparent mol. wt. in S D S - P A G E is due to heavy glycosylation. This
glycosylation involves both N- and O-linked oligosaccharides (Gruber & Levine, 1985). Since
mutational events could readily change any of the many sites of glycosylation, a variation in
molecular weight of the G protein in different strains would be expected to occur. Repeated
analysis of individual strains showed that the G protein had stable size characteristics.
The stability of non-glycosylated F proteins of subtype A and B strains was different. By
analogy with other systems it would appear that glycosylation contributes to the resistance of the
RS virus F proteins to proteolysis or possibly alters the capacity of the protein to react with the
MAbs employed in RIPA. In most experiments the use of Vero cells, which show a relatively
high endogenous proteolytic activity, may have assisted in revealing the difference in stability of
the F proteins of RS virus subtype A and B strains. It can not be deduced whether the differences
in glycosylation which cause the high sensitivity of non-glycosylated RS virus subtype B F1
protein to proteolysis is quantitative or qualitative. The present data do not indicate major
differences in glycosylation of the F1 and F2 proteins of viruses representing the two subtypes,
suggesting that the main difference may be qualitative. In fact this finding may not be a
reflection of differences in glycosylation, but may be due to variations in protein tertiary
structure.
During the preparation of this manuscript an article representing an extension of previously
published data (Gimenez et al., 1984), dealing with antigenic variation between human RS virus
isolates appeared (Gimenez et al., 1986). Twenty-one out of 42 strains lacked the capacity to
react with one of two anti-P MAbs. The mol. wt. of the P protein of representative isolates of
these 21 strains was 3K higher (35K compared to 32K) than in other strains. Judging from the
present data the former 21 strains represent RS subtype A viruses. As a corollary it can be
concluded that the strain RSN-2 used to generate the MAbs has subtype B characteristics. Two
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Differences between R S virus subtypes
2729
o f t h e t h r e e M A b s u s e d t h e r e f o r e a r e s u b t y p e B - s p e c i f i c . I n r e c e n t s t u d i e s in t h i s l a b o r a t o r y a
l a r g e c o l l e c t i o n o f s u b t y p e B - s p e c i f i c M A b s h a v e b e e n g e n e r a t e d (C. O r v e l l et al., u n p u b l i s h e d
results).
The excellent technical assistance of Mariethe Ehnlund, Anita Bergstr6m and Barbara Akerblom is gratefully
acknowledged. This work was supported by a grant from the Swedish Medical Research Council (project no B8616X-00116-22C) and the WHO Programme for Vaccine Development.
REFERENCES
L~ERLIND, B. & NORRBY, E. (1986). Occurrence of respiratory syncytial (RS) virus subtype A and B strains in
Sweden. Journal of Medical Virology 19, 241-247.
ANDERSON,L. I., HIERHDLZER,J. C., TSOU,C., HENDRY,R. M., FERNIE, B. F., STONE,Y. & McINTOSH,K. (1985) Antigenic
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(Received 30 June 1986)
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