Transcript subtype B consensus CTRPNNNTRKSIHIGPGRAFYATGdIIGDIRQAHC BaL.26 ---------------------T--E---------JRFL ---------------------T--E---------ADA ---------------------T--E---------YU2 ------------N------L-T--E---------6535.3 ------------NL----------D---------QH0692.42 ----G-------------------D---------SC42.8 ----------G-T-----V--~--E-V-----V-PVO.04 ------------S-----------D---------TRO.11 ---------R--------------D---------AC10.29 -I--------G----------T--D---------RHPA.7 ---H--------N-----------K---------THRO.18 --------S----M---G--F---R------K-YREJO.67 --------------A---------E------K-YTRJO.58 ----H--------------W----D------K-YWITO.33 ----G----R--N-----------A------K--CAAN.A2 --------S---T-----------R------K--- subtype C consensus ZA012 TV1.29 DU156.12 DU172.17 DU422.1 ZM197M.PB7 ZM214M.PL15 ZM233M.PB6 ZM249M.PL1 ZM53MPB12 ZM109FPB4 ZM135M.PL10a CAP45.2.00.G3 CAP210.2.00.E8 CTRPNNNTRKSIRIGPGQTFYATGdIIGDIRQAHC -----------M------------D--------Y-----------V------A----NDV-------------------V------------D-------------S------V--------F---D--------------------V------------E------E---V---------V--------F---E----------M--G----R-V------------E---------------------------S-----E-V-N--E--------------------------E---K--E------G-------------A-F--TN--------Y-I--G--------L----------DV-----K-Y------------------------EV---------R----------------A----ND----------I--G----R--------A---M-D---N--E--- Supplementary Figure 1. Amino acid sequence alignments of the V3 region from subtype B (top)

subtype B consensus CTRPNNNTRKSIHIGPGRAFYATGdIIGDIRQAHC
BaL.26
---------------------T--E---------JRFL
---------------------T--E---------ADA
---------------------T--E---------YU2
------------N------L-T--E---------6535.3
------------NL----------D---------QH0692.42
----G-------------------D---------SC42.8
----------G-T-----V--~--E-V-----V-PVO.04
------------S-----------D---------TRO.11
---------R--------------D---------AC10.29
-I--------G----------T--D---------RHPA.7
---H--------N-----------K---------THRO.18
--------S----M---G--F---R------K-YREJO.67
--------------A---------E------K-YTRJO.58
----H--------------W----D------K-YWITO.33
----G----R--N-----------A------K--CAAN.A2
--------S---T-----------R------K---
subtype C consensus
ZA012
TV1.29
DU156.12
DU172.17
DU422.1
ZM197M.PB7
ZM214M.PL15
ZM233M.PB6
ZM249M.PL1
ZM53MPB12
ZM109FPB4
ZM135M.PL10a
CAP45.2.00.G3
CAP210.2.00.E8
CTRPNNNTRKSIRIGPGQTFYATGdIIGDIRQAHC
-----------M------------D--------Y-----------V------A----NDV-------------------V------------D-------------S------V--------F---D--------------------V------------E------E---V---------V--------F---E----------M--G----R-V------------E---------------------------S-----E-V-N--E--------------------------E---K--E------G-------------A-F--TN--------Y-I--G--------L----------DV-----K-Y------------------------EV---------R----------------A----ND----------I--G----R--------A---M-D---N--E---
Supplementary Figure 1. Amino acid sequence alignments of the V3 region from subtype B (top) and
subtype C (bottom) HIV-1 isolates tested in this paper. “-” depicts identical residues to the corresponding
consensus sequences. “~” depicts a gap to maintain the alignment.
A
GP group 82 immunized with a clade B V3 (BaL∆V1V2(V3-1AB))
100%
% neutralization
80%
82-1
82-2
82-3
82-4
82-6
60%
40%
20%
D
U
12
3.
6
17
6.
66
02
5
R
ZM
01
2
1.
29
B
TV
42
2
ZA
C
S
C
A
65
35
.3
10
.2
9
O
.1
1
TR
B
61
01
07
F
R
S
C
JR
F1
62
S
.2
6
aL
B
B
aL
.0
1
0%
B
GP group 83 immunized with a clade C V3 (BaL∆V1V2(ZA012V3-1AB))
100%
% neutralization
80%
60%
83-1
83-2
83-3
83-4
40%
20%
D
U
12
3.
6
17
6.
66
02
5
R
ZM
01
2
1.
29
B
TV
ZA
42
2
C
S
C
A
65
35
.3
10
.2
9
O
.1
1
61
01
R
B
TR
07
F
S
C
JR
F1
62
S
.2
6
aL
B
B
aL
.0
1
0%
GP group 86 immunized with a clade C V3 (Hx/BaL∆V1V2(ZA012V3-1AB))
100%
C
% neutralization
80%
86-1
86-2
86-3
86-4
86-5
86-6
60%
40%
20%
12
6
ZM
53
M
.P
B
B
23
3M
.P
u4
22
.1
ZM
D
u1
72
.1
7
D
02
5
R
B
1.
29
TV
01
2
ZA
65
35
.3
u1
56
.1
2
D
S
S
11
96
.1
FL
JR
F1
62
S
aL
.2
6
B
B
aL
.0
1
0%
Supplementary Figure 2. Neutralization by GP vaccine sera at 1:5. (A) Group 82 immunized with a
modified BaL Env containing the BaL V3 sequence; (B) group 83 immunized with a modified BaL Env
containing the subtype C ZA012 V3 sequence; (C) group 86 immunized with a modified chimeric
Hx/BaL Env containing the ZA012 V3 sequence. Neutralization values equal to or below 0 are plotted at
1%. Immune sera highlighted in a solid color are those described in this paper. Solid blue bar denotes
subtype B viruses; solid red bar denotes subtype C viruses.
% adjusted neutralization
447-52D
17b
% adjusted neutralization
100
100
P=0.03
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
2219
sCD4
3074
100
% adjusted neutralization
P=0.66
80
100
P=0.002
P=0.03
80
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
GP 82-2
sCD4
GP 82-4
100
100
P=0.007
P=0.005
80
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
sCD4
Supplementary Figure 3. Statistical comparison of neutralization of subtype B viruses by anti-V3
mAbs (10 ug/ml) and GP sera (1:10) without sCD4 and with an IC80 of sCD4. Bars indicate the median
neutralization value of each antibody against the set of 15 viruses. P values were generated using the
paired Wilcoxon test and are significant for the anti-V3 mAbs and GP vaccine sera.
% adjusted neutralization
447-52D
GP 83-2
100
100
P=0.21
80
P=0.09
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
% adjusted neutralization
2219
GP 86-2
100
100
P=0.05
P=0.10
80
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
3074
% adjusted neutralization
sCD4
sCD4
GP 86-6
100
100
P=0.003
80
P=0.02
80
60
60
40
40
20
20
0
0
No sCD4
sCD4
No sCD4
sCD4
Supplementary Figure 4. Statistical comparison of neutralizations of subtype C viruses by anti-V3
mAbs (10 ug/ml) and GP sera (1:10) without sCD4 and with an IC80 of sCD4. Bars indicate the
median neutralization value for each antibody against the set of 13 viruses. P values were
generated using the paired Wilcoxon test and are significant for mAbs 2219, 3074 and GP 86-6.