Single chain Fv-based anti-HIV proteins: Potential
and Limitations
Anthony P. West, Jr., Rachel P. Galimidi, Priyanthi N. P. Gnanapragasam, and
Pamela J. Bjorkman
Supplemental Tables S1-S3
Supplemental Figures S1-S3
Supplemental Table S1. IC50 values (g/mL) for IgG, Fab, IA, and scFv forms of
VRC01, PG9, and PG16. IC50 values in nM are presented in Table 1.
1
Supplemental Table S2. IC50 values (nM) for PG16 IA (VH-VL) (opposite orientation
to the PG16 IA (VL-VH) reagent in Table 1). †Due of the incomplete strain coverage of
these data, a comparison of the mean potency with PG16 IgG is not given. ND = not
determined.
PG16
Clade
B
B
B
B
B
B
B
B
B
B
B
B
IgG
285
>680*
120
89.5
19.4
0.22
0.11
25.5
0.095
140
85.5
3.4
Fab
1800
>2000
850
260
390
1.2
26
240
0.51
590
2000
53
IA (VL-VH)
1.9
>930
200
>930
>930
3.7
45
11
1.4
>930
>930
>390
IA (VH-VL)
650
>930
>930
>930
620
ND
ND
ND
<0.46
200
ND
43
Du156.12
Du172.17
Du422.1
ZM197M.PB7
ZM214M.PL15
ZM233M.PB6
ZM249M.PL1
ZM53M.PB12
ZM109F.PB4
ZM135M.PL10a
CAP45.2.00.G3
CAP210.2.00.E8
C
C
C
C
C
C
C
C
C
C
C
C
0.16
0.41
0.95
8.1
>680*
0.034
0.81
0.25
134
>680*
0.027
0.47
0.67
2.6
28
73
>2000
0.081
4.3
1.1
>2000
>2000
0.061
14
4.9
12
94
33
>930
0.88
3.7
3.2
15
>930
9.0
51
<0.46
0.65
61
ND
ND
<0.46
ND
<0.46
ND
ND
ND
<0.46
Q23.17
Q842.d12
Q259.d2.17
3718.v3.c11
0330.v4.c3
3415.v1.c1
A
A
A
A
A
A
0.014
0.18
0.068
0.068
0.14
0.14
0.20
0.63
19
2.1
0.69
2.3
0.45
3.5
3.4
24
2.8
22
<0.46
<0.46
ND
1.0
<0.46
2.0
1.2
12.7
10.5
18.9
15.5
†
†
Virus
6535.3
QH0692.42
SC422661.8
PVO.4
TRO.11
AC10.0.29
REJO4541.67
TRJO4551.58
WITO4160.33
CAAN5342.A2
THRO4156.18
RHPA4259.7
Geometric mean(*)
MNR
2
Supplemental Table S3. IC50 values (nM) for VRC01scFv-PG16 reagents with the
VRC01 scFv attached to the N-terminus of the PG16 light chain (the reagent shown
in Table 1) or the N-terminus of the PG16 heavy chain.
VRC01scFv-PG16
Virus
Clade
scFv attached to:
LC
HC
6535.3
QH0692.42
SC422661.8
PVO.4
TRO.11
AC10.0.29
REJO4541.67
TRJO4551.58
WITO4160.33
CAAN5342.A2
THRO4156.18
RHPA4259.7
B
B
B
B
B
B
B
B
B
B
B
B
48
14
2.1
15
5.4
0.34
0.15
0.44
0.15
24
7.9
1.0
7.1
7.5
3.4
13
3.9
1.1
0.10
0.73
0.20
23
18
0.73
Du156.12
Du172.17
Du422.1
ZM197M.PB7
ZM214M.PL15
ZM233M.PB6
ZM249M.PL1
ZM53M.PB12
ZM109F.PB4
ZM135M.PL10a
CAP45.2.00.G3
CAP210.2.00.E8
C
C
C
C
C
C
C
C
C
C
C
C
0.29
2.3
19
5.6
>49
0.05
0.34
0.34
3.1
37
0.05
2.8
0.44
8.3
21
4.7
31
0.15
0.24
0.63
1.4
8.6
0.05
16
Q23.17
Q842.d12
Q259.d2.17
3718.v3.c11
0330.v4.c3
3415.v1.c1
A
A
A
A
A
A
<0.024
0.15
1.6
1.4
0.24
0.59
<0.05
0.15
1.7
1.4
0.34
1.02
1.40
3.16
3.22
1.56
1.80
9.24
Geometric mean
Model fit for veff
Model fit for peff
3
Supplemental Fig. S1. DNA and amino acid sequences of constructs. In DNA
sequences, surrounding non-coding sequences are shown in italics. For amino acid
sequences, hydrophobic leader sequences are listed in the line above the sequence
of the mature protein.
The following sequences are listed:
1) VRC01 IA (scFv(VH-VL)–Fc) – plasmid VRC9713 – from G. Nabel
2) PG9 IA (scFv(VH-VL)–Fc)
3) PG16 IA (scFv(VL-VH)–Fc)
4) PG16 IA (scFv(VH-VL)–Fc)
5) VRC01 HC–2A–LC plasmid VRC9715 – from G. Nabel
6) PG9 scFv-His6 tagged
7) VRC01scFv-E51 HC
8) VRC01scFv-PG16 LC
9) VRC01scFv-ZAG3-Fc
10) VRC01 Fab HC (Fd)- His7 tagged
11) PG9 Fab HC (Fd)- His7 tagged
12) PG16 Fab HC (Fd)- His7 tagged
13) VRC01scFv–PG16 HC
1) VRC01 IA (VH-(G3S)4-VL-Fc (from hIgG1)) (plasmid VRC9713)
DNA:
TCTAGACCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGGTG
TACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGTCGA
TGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGGATT
CGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGGGCC
GTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCCGAC
ACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGTACT
AGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCCGGTC
ATCGTCTCATCAGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGATC
CGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAAACAGCCAT
CATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGGCCCGGCCA
GGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAGACAGGTT
CAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGTCGGGAGA
TTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAGGTCCAGGT
CGACATTAAAGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACC
TGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCAT
GATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGA
GGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGC
GGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGG
ACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCA
TCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGC
CCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCT
TCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACA
AGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGT
4
GGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCT
GCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGATGA
Protein:
MGWSCIILFLVATATGVHS
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSS
GGGSGGGSGGGSGGGS
EIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPRLVIYSGSTRAAGIPDRFSGS
RWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQV_DIK
EPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN
WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKT
ISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP
VLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
2) PG9 IA (VH-(G4S)3-VL-Fc(from hIgG2))
DNA:
GAATTCGCCGCCACCATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTTTCTTAAGAG
GTGTCCAGTGTCAGCGATTAGTGGAGTCTGGGGGAGGCGTGGTCCAGCCTGGGTCGTCCC
TGAGACTCTCCTGTGCAGCGTCCGGATTCGACTTCAGTAGACAAGGCATGCACTGGGTCC
GCCAGGCTCCAGGCCAGGGGCTGGAGTGGGTGGCATTTATTAAATATGATGGAAGTGAG
AAATATCATGCTGACTCCGTATGGGGCCGACTCAGCATCTCCAGAGACAATTCCAAGGAT
ACGCTTTATCTCCAAATGAATAGCCTGAGAGTCGAGGACACGGCTACATATTTTTGTGTG
AGAGAGGCTGGTGGGCCCGACTACCGTAATGGGTACAACTATTACGATTTCTATGATGG
TTATTATAACTACCACTATATGGACGTCTGGGGCAAAGGGACCACCGTGACAGTCTCGAG
CGCCTCCGGTGGCGGTGGCTCCGGAGGTGGTGGGAGCGGTGGCGGCGGATCTCAGTCTGC
CCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTCGATCACCATCTCCTGCAAT
GGAACCAGCAATGATGTTGGTGGCTATGAATCTGTCTCCTGGTACCAACAACATCCCGGC
AAAGCCCCCAAAGTCGTGATTTATGATGTCAGTAAACGGCCCTCAGGGGTTTCTAATCGC
TTCTCTGGCTCCAAGTCCGGCAACACGGCCTCCCTGACCATCTCTGGGCTCCAGGCTGAG
GACGAGGGTGACTATTACTGCAAGTCTCTGACAAGCACGAGACGTCGGGTTTTCGGCACT
GGGACCAAGCTGACCGTTCTAGAGCGCAAATGTTGTGTCGAGTGCCCACCGTGCCCAGCA
CCACCTGTGGCAGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGA
TCTCCCGGACCCCTGAGGTCACGTGCGTGGTGGTGGACGTGAGCCACGAAGACCCCGAGG
TCCAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCACGGG
AGGAGCAGTTCAACAGCACGTTCCGTGTGGTCAGCGTCCTCACCGTTGTGCACCAGGACT
GGCTGAACGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGGCCTCCCAGCCCCCATCG
AGAAAACCATCTCCAAAACCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCC
CATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCT
ACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAG
ACCACACCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGG
ACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGC
ACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGAGCGGCCGC
5
Protein:
MEFGLSWVFLVAFLRGVQC
QRLVESGGGVVQPGSSLRLSCAASGFDFSRQGMHWVRQAPGQGLEWVAFIKYDGSEKYH
ADSVWGRLSISRDNSKDTLYLQMNSLRVEDTATYFCVREAGGPDYRNGYNYYDFYDGYY
NYHYMDVWGKGTTVTVSSAS
GGGGSGGGGSGGGGS
QSALTQPASVSGSPGQSITISCNGTSNDVGGYESVSWYQQHPGKAPKVVIYDVSKRPSGVS
NRFSGSKSGNTASLTISGLQAEDEGDYYCKSLTSTRRRVFGTGTKLTVLERK
CCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGV
EVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQ
PREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
3) PG16 IA (VL-(G4S)3-VH-Fc (from hIgG2))
DNA:
ATGGCCTGGGCTCTGCTATTCCTCACCCTCTTCACTCAGGGCACAGGGTCCTGGGGCCAG
TCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGACGATCACCATCTCCT
GCAATGGAACCAGCAGTGACGTTGGTGGATTTGACTCTGTCTCCTGGTACCAACAATCCC
CAGGGAAAGCCCCCAAAGTCATGGTTTTTGATGTCAGTCATCGGCCCTCAGGTATCTCTA
ATCGCTTCTCTGGCTCCAAGTCCGGCAACACGGCCTCCCTGACCATCTCTGGGCTCCACAT
TGAGGACGAGGGCGATTATTTCTGCTCTTCACTGACAGACAGAAGCCATCGCATATTCGG
CGGCGGGACCAAGGTGACCGTTCTAGGTGGCGGTGGCTCCGGAGGTGGTGGGAGCGGTGG
CGGCGGATCTCAGGAACAACTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCGGGGGGGTC
CCTGAGACTCTCCTGTTTAGCGTCTGGATTCACGTTTCACAAATATGGCATGCACTGGGT
CCGCCAGGCTCCAGGCAAGGGCCTGGAGTGGGTGGCACTCATCTCAGATGACGGAATGAG
GAAATATCATTCAGACTCCATGTGGGGCCGAGTCACCATCTCCAGAGACAATTCCAAGAA
CACTCTTTATCTGCAATTCAGCAGCCTGAAAGTCGAAGACACGGCTATGTTCTTCTGTGC
GAGAGAGGCTGGTGGGCCAATCTGGCATGACGACGTCAAATATTACGATTTTAATGACG
GCTACTACAACTACCACTACATGGACGTCTGGGGCAAGGGGACCACGGTCACCGTCTCGA
GCGCCTCCGAGCGCAAATGTTGTGTCGAGTGCCCACCGTGCCCAGCACCACCTGTGGCAG
GACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCC
TGAGGTCACGTGCGTGGTGGTGGACGTGAGCCACGAAGACCCCGAGGTCCAGTTCAACTG
GTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCACGGGAGGAGCAGTTCA
ACAGCACGTTCCGTGTGGTCAGCGTCCTCACCGTTGTGCACCAGGACTGGCTGAACGGCA
AGGAGTACAAGTGCAAGGTCTCCAACAAAGGCCTCCCAGCCCCCATCGAGAAAACCATCT
CCAAAACCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGG
AGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTACCCCAGCGACA
TCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACACCTCCCA
TGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGT
GGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACA
CGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA
Protein:
MAWALLFLTLFTQGTGSWG
6
QSALTQPASVSGSPGQTITISCNGTSSDVGGFDSVSWYQQSPGKAPKVMVFDVSHRPSGIS
NRFSGSKSGNTASLTISGLHIEDEGDYFCSSLTDRSHRIFGGGTKVTVL
GGGGSGGGGSGGGGS
QEQLVESGGGVVQPGGSLRLSCLASGFTFHKYGMHWVRQAPGKGLEWVALISDDGMRKY
HSDSMWGRVTISRDNSKNTLYLQFSSLKVEDTAMFFCAREAGGPIWHDDVKYYDFNDGY
YNYHYMDVWGKGTTVTVSSASERK
CCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGV
EVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQ
PREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
4) PG16 IA (VH-(G4S)3-VL-Fc (from hIgG2))
DNA:
GAATTCGCCGCCACCATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTTTCTTAAGAG
GTGTCCAGTGTCAGGAACAACTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCGGGGGGG
TCCCTGAGACTCTCCTGTTTAGCGTCTGGATTCACGTTTCACAAATATGGCATGCACTGG
GTCCGCCAGGCTCCAGGCAAGGGCCTGGAGTGGGTGGCACTCATCTCAGATGACGGAATG
AGGAAATATCATTCAGACTCCATGTGGGGCCGAGTCACCATCTCCAGAGACAATTCCAAG
AACACTCTTTATCTGCAATTCAGCAGCCTGAAAGTCGAAGACACGGCTATGTTCTTCTGT
GCGAGAGAGGCTGGTGGGCCAATCTGGCATGACGACGTCAAATATTACGATTTTAATGA
CGGCTACTACAACTACCACTACATGGACGTCTGGGGCAAGGGGACCACGGTCACCGTCTC
GAGCGCCTCCGGTGGCGGTGGCTCCGGAGGTGGTGGGAGCGGTGGCGGCGGATCTCAGTC
TGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGACGATCACCATCTCCTGC
AATGGAACCAGCAGTGACGTTGGTGGATTTGACTCTGTCTCCTGGTACCAACAATCCCCA
GGGAAAGCCCCCAAAGTCATGGTTTTTGATGTCAGTCATCGGCCCTCAGGTATCTCTAAT
CGCTTCTCTGGCTCCAAGTCCGGCAACACGGCCTCCCTGACCATCTCTGGGCTCCACATTG
AGGACGAGGGCGATTATTTCTGCTCTTCACTGACAGACAGAAGCCATCGCATATTCGGCG
GCGGGACCAAGGTGACCGTTCTAGAGCGCAAATGTTGTGTCGAGTGCCCACCGTGCCCAG
CACCACCTGTGGCAGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCAT
GATCTCCCGGACCCCTGAGGTCACGTGCGTGGTGGTGGACGTGAGCCACGAAGACCCCGA
GGTCCAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCACG
GGAGGAGCAGTTCAACAGCACGTTCCGTGTGGTCAGCGTCCTCACCGTTGTGCACCAGGA
CTGGCTGAACGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGGCCTCCCAGCCCCCAT
CGAGAAAACCATCTCCAAAACCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCC
CCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTT
CTACCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAA
GACCACACCTCCCATGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTG
GACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTG
CACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGAGCGGCCGC
Protein:
MEFGLSWVFLVAFLRGVQC
QEQLVESGGGVVQPGGSLRLSCLASGFTFHKYGMHWVRQAPGKGLEWVALISDDGMRKY
HSDSMWGRVTISRDNSKNTLYLQFSSLKVEDTAMFFCAREAGGPIWHDDVKYYDFNDGY
YNYHYMDVWGKGTTVTVSSAS
7
GGGGSGGGGSGGGGS
QSALTQPASVSGSPGQTITISCNGTSSDVGGFDSVSWYQQSPGKAPKVMVFDVSHRPSGIS
NRFSGSKSGNTASLTISGLHIEDEGDYFCSSLTDRSHRIFGGGTKVTVLERK
CCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGV
EVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQ
PREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
5) VRC01-HC-2A-LC plasmid (hIgG1) (Plasmid VRC9715)
DNA:
TCTAGACCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGGTG
TACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGTCGA
TGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGGATT
CGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGGGCC
GTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCCGAC
ACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGTACT
AGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCCGGTC
ATCGTCTCATCACCGTCGACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGA
GCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGG
TGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCC
TACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGG
GCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGA
AAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAAC
TCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTC
CCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAA
GTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGG
AGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGC
TGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGA
AAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCAT
CCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATC
CCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACC
ACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACA
AGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACA
ACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAACGGCGGCGGCGGGCCCCCG
TGAAGCAGACCCTGAACTTCGACCTGCTGAAGCTGGCCGGCGACGTGGAGAGCAACCCCG
GCCCCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGGTGTACATT
CAGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAAACAGCCA
TCATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGGCCCGGCC
AGGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAGACAGGT
TCAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGTCGGGAG
ATTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAGGTCCAGG
TCGACATTAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGC
AGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTACCCCAGAGAAG
CCAAAGTGCAGTGGAAGGTGGACAACGCCCTGCAGAGCGGAAACAGCCAGGAAAGCGTG
ACAGAGCAGGATTCCAAGGATTCCACATACAGCCTGAGCAGCACACTGACACTGTCCAAG
8
GCCGACTACGAGAAGCACAAGGTGTACGCCTGCGAAGTGACACACCAGGGACTGTCCTCC
CCTGTGACAAAGAGCTTCAACAGAGGAGAATGCTGATAGGATCC
Protein:
MGWSCIILFLVATATGVHS
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSPSTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS
SGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGP
SVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYN
STYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDEL
TKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
RRRRAPVKQTLNFDLLKLAGDVESNPGP
MGWSCIILFLVATATGVHS
EIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPRLVIYSGSTRAAGIPDRFSGS
RWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQVDIKRTVAAPSVFIFPPSDEQLKSG
TASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEK
HKVYACEVTHQGLSSPVTKSFNRGEC
6) PG9 scFv-His6 tagged:
DNA:
GAATTCGCCGCCACCATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTTTCTTAAGAG
GTGTCCAGTGTCAGCGATTAGTGGAGTCTGGGGGAGGCGTGGTCCAGCCTGGGTCGTCCC
TGAGACTCTCCTGTGCAGCGTCCGGATTCGACTTCAGTAGACAAGGCATGCACTGGGTCC
GCCAGGCTCCAGGCCAGGGGCTGGAGTGGGTGGCATTTATTAAATATGATGGAAGTGAG
AAATATCATGCTGACTCCGTATGGGGCCGACTCAGCATCTCCAGAGACAATTCCAAGGAT
ACGCTTTATCTCCAAATGAATAGCCTGAGAGTCGAGGACACGGCTACATATTTTTGTGTG
AGAGAGGCTGGTGGGCCCGACTACCGTAATGGGTACAACTATTACGATTTCTATGATGG
TTATTATAACTACCACTATATGGACGTCTGGGGCAAAGGGACCACcGTgACaGTCTCGAG
CGGTGGCGGTGGCTCCGGAGGTGGTGGGAGCGGTGGCGGCGGATCTCAGTCTGCCCTGAC
TCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTCGATCACCATCTCCTGCAATGGAAC
CAGCAATGATGTTGGTGGCTATGAATCTGTCTCCTGGTACCAACAACATCCCGGCAAAGC
CCCCAAAGTCGTGATTTATGATGTCAGTAAACGGCCCTCAGGGGTTTCTAATCGCTTCTC
TGGCTCCAAGTCCGGCAACACGGCCTCCCTGACCATCTCTGGGCTCCAGGCTGAGGACGA
GGGTGACTATTACTGCAAGTCTCTGACAAGCACGAGACGTCGGGTTTTCGGCACTGGGAC
CAAGCTGACCGTTCTACATCATCACCATCACCACTGAGCGGCCGC
Protein:
MEFGLSWVFLVAFLRGVQC
QRLVESGGGVVQPGSSLRLSCAASGFDFSRQGMHWVRQAPGQGLEWVAFIKYDGSEKYH
ADSVWGRLSISRDNSKDTLYLQMNSLRVEDTATYFCVREAGGPDYRNGYNYYDFYDGYY
NYHYMDVWGKGTTVTVSSGGGGSGGGGSGGGGSQSALTQPASVSGSPGQSITISCNGTSND
VGGYESVSWYQQHPGKAPKVVIYDVSKRPSGVSNRFSGSKSGNTASLTISGLQAEDEGDYY
CKSLTSTRRRVFGTGTKLTVLHHHHHH
9
7) VRC01scFv-E51 HC:
DNA:
AATTCGCCGCCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGG
TGTACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGTC
GATGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGGA
TTCGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGGG
CCGTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCCG
ACACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGTA
CTAGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCCGG
TCATCGTCTCATCAGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGA
TCCGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAAACAGCC
ATCATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGGCCCGGC
CAGGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAGACAGG
TTCAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGTCGGGA
GATTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAGGTCCAG
GTCGACATTAAAGCCGGCGGCAGCGGAGGAGGATCTGGGGGAGGGTCCGGAGGCGGATC
CGGGGGCGGTTCAGGTGGGGGTAGCGGAGGGGGCTCAGGCGGAGCTAGCGAGGTGCAGC
TGGTGCAGTCTGGGGCTGAGGTGAACAAGCCTGGGTCCTCGGTGAAGGTCTCCTGTCAGG
CTTCTGGAGCCACCTTAAACAGTCATGCTTTCAGCTGGGTGCGACAGGCCCCTGGACAAG
GGCTTGAGTGGATGGCAGGGATCATCCCTATATTTGGTTCATCACACTACGCACAAAAGT
TCCAGGGCAGAGTCACGATTACCGCGGACGAGTCCACGCGCACAGTGTATTTGCATCTGA
GAGGCCTGAGATCTGATGACACGGCCGTTTATTACTGTGCGAGTAATTCTATTGCCGGAG
TAGCAGCTGCCGGAGACTACGCTGACTACGATGGGGGCTACTACTACGATATGGATGTCT
GGGGCCAAGGGACCACGGTCACCGTCTCCTCAGCCTCCACCAAGGGCCCATCGGTCTTCCC
CCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAA
GGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGT
GCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACC
GTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGC
AACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACGTGCCCA
CCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCA
AGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCC
ACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCC
AAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGGGTGGTCAGCGTCCTCACC
GTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCC
CTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGTCAGCCCCGAGAACCACAG
GTGTACACCCTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGC
CTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCG
GAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTAC
AGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTG
ATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA
TGA
Protein:
MGWSCIILFLVATATGVHS
10
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSGGGSGGGSGGGSGGGSEIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPR
LVIYSGSTRAAGIPDRFSGSRWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQVDIKAG
GSGGGSGGGSGGGSGGGSGGGSGGGSGGASEVQLVQSGAEVNKPGSSVKVSCQASGATLNS
HAFSWVRQAPGQGLEWMAGIIPIFGSSHYAQKFQGRVTITADESTRTVYLHLRGLRSDDT
AVYYCASNSIAGVAAAGDYADYDGGYYYDMDVWGQGTTVTVSSASTKGPSVFPLAPSSKS
TSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQ
TYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTP
EVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLN
GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHY
TQKSLSLSPGK
8) VRC01scFv–PG16 LC:
DNA:
AATTCGCCGCCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGG
TGTACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGTC
GATGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGGA
TTCGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGGG
CCGTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCCG
ACACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGTA
CTAGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCCGG
TCATCGTCTCATCAGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGA
TCCGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAAACAGCC
ATCATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGGCCCGGC
CAGGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAGACAGG
TTCAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGTCGGGA
GATTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAGGTCCAG
GTCGACATTAAAGCCGGCGGCAGCGGAGGAGGATCTGGGGGAGGGTCCGGAGGCGGATC
CGGGGGCGGTTCAGGTGGGGGTAGCGGAGGGGGCTCAGGCGGAGCTAGCCAGTCTGCCCT
GACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGACGATCACCATCTCCTGCAATGG
AACCAGCAGTGACGTTGGTGGATTTGACTCTGTCTCCTGGTACCAACAATCCCCAGGGAA
AGCCCCCAAAGTCATGGTTTTTGATGTCAGTCATCGGCCCTCAGGTATCTCTAATCGCTT
CTCTGGCTCCAAGTCCGGCAACACGGCCTCCCTGACCATCTCTGGGCTCCACATTGAGGA
CGAGGGCGATTATTTCTGCTCTTCACTGACAGACAGAAGCCATCGCATATTCGGCGGCGG
GACCAAGGTGACCGTTCTAGGTCAGCCCAAGGCTGCCCCCTCGGTCACTCTGTTCCCGCCC
TCCTCTGAGGAGCTTCAAGCCAACAAGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC
CCGGGAGCCGTGACAGTGGCCTGGAAGGCAGATAGCAGCCCCGTCAAGGCGGGAGTGGAG
ACCACCACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCAGCTACCTGAGCCTG
ACGCCTGAGCAGTGGAAGTCCCACAAAAGCTACAGCTGCCAGGTCACGCATGAAGGGAGC
ACCGTGGAGAAGACAGTGGCCCCTACAGAATGTTCATAG
Protein:
MGWSCIILFLVATATGVHS
11
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSGGGSGGGSGGGSGGGSEIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPR
LVIYSGSTRAAGIPDRFSGSRWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQVDIKAG
GSGGGSGGGSGGGSGGGSGGGSGGGSGGASQSALTQPASVSGSPGQTITISCNGTSSDVGGF
DSVSWYQQSPGKAPKVMVFDVSHRPSGISNRFSGSKSGNTASLTISGLHIEDEGDYFCSSLT
DRSHRIFGGGTKVTVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKAD
SSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHKSYSCQVTHEGSTVEKTVAPTECS
9) VRC01scFv–ZAG3–E51 HC:
DNA:
GAATTCCCTCTAGACCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCA
ACCGGTGTACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGC
GAGTCGATGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAA
TTGGATTCGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGG
GGGGGCCGTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTA
TTCCGACACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTT
TTGTACTAGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCA
CCCCGGTCATCGTCTCATCAGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGAAGCGGA
GGGGGATCCGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAA
ACAGCCATCATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGG
CCCGGCCAGGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAG
ACAGGTTCAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGT
CGGGAGATTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAG
GTCCAGGTCGACATTAAATCCGGAGGTGGCGGTAGTGGGGGAGGCGACCCGCCAAGCGTG
GTGGTCACATCCCATCAAGCCCCTGGTGAAAAGAAGAAGCTCAAATGTTTGGCCTACGAT
TTCTATCCAGGGAAGATCGACGTGCACTGGACTAGAGCCGGAGAAGTCCAGGAGCCGGA
ACTCAGGGGTGATGTCCTGCACAACGGCAACGGCACTTACCAGTCTTGGGTCGTCGTAGC
CGTGCCACCTCAGGACACGGCGCCATACTCCTGCCATGTGCAACATTCTTCCCTCGCACAG
CCCCTGGTCGTGCCTTGGGAGGCCTCCGGTGAGCCCAAATCTTGTGACAAAACTCACACA
TGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAA
AACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACG
TGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCAT
AATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTC
CTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAAC
AAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAA
CCACAGGTGTACACCCTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTG
ACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGG
CAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTC
CTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGC
TCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCG
GGTAAATGATGAGGATCC
Protein:
MGWSCIILFLVATATGVHS
12
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSGGGSGGGSGGGSGGGSEIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPR
LVIYSGSTRAAGIPDRFSGSRWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQVDIKSG
GGGSGGGDPPSVVVTSHQAPGEKKKLKCLAYDFYPGKIDVHWTRAGEVQEPELRGDVLH
NGNGTYQSWVVVAVPPQDTAPYSCHVQHSSLAQPLVVPWEASGEPKSCDKTHTCPPCPA
PELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKP
REEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTL
PPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
10) VRC01 Fab HC (Fd)- His7 tagged
DNA:
ATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCGGTGTACATTCCCAG
GTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGTCGATGAGAATTTC
TTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGGATTCGTCTGGCCCC
CGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGGGCCGTCAACTACG
CACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCCGACACAGCCTTTT
TGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGTACTAGGGGAAAAA
ACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCCGGTCATCGTCTCAT
CACCGTCGACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGG
GGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTC
GTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTC
AGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGAC
CTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCC
CAAATCTTGTGACAAACATCATCACCATCACCACCAT
Protein:
MGWSCIILFLVATATGVHS
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSPSTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS
SGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKHHHHHHH
11) PG9 Fab HC (Fd)- His7 tagged
DNA:
ATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTTTCTTAAGAGGTGTCCAGTGTCAG
CGATTAGTGGAGTCTGGGGGAGGCGTGGTCCAGCCTGGGTCGTCCCTGAGACTCTCCTGT
GCAGCGTCCGGATTCGACTTCAGTAGACAAGGCATGCACTGGGTCCGCCAGGCTCCAGGC
CAGGGGCTGGAGTGGGTGGCATTTATTAAATATGATGGAAGTGAGAAATATCATGCTGA
CTCCGTATGGGGCCGACTCAGCATCTCCAGAGACAATTCCAAGGATACGCTTTATCTCCA
AATGAATAGCCTGAGAGTCGAGGACACGGCTACATATTTTTGTGTGAGAGAGGCTGGTG
GGCCCGACTACCGTAATGGGTACAACTATTACGATTTCTATGATGGTTATTATAACTACC
ACTATATGGACGTCTGGGGCAAAGGGACCACCGTGACAGTCTCGAGCGCCTCCACCAAGG
13
GCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCT
GGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGC
CCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTC
AGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTG
AATCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAAATCTTGTGACAA
ACATCATCACCATCACCACCAT
Protein:
MEFGLSWVFLVAFLRGVQC
QRLVESGGGVVQPGSSLRLSCAASGFDFSRQGMHWVRQAPGQGLEWVAFIKYDGSEKYH
ADSVWGRLSISRDNSKDTLYLQMNSLRVEDTATYFCVREAGGPDYRNGYNYYDFYDGYY
NYHYMDVWGKGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCD
KHHHHHHH
12) PG16 Fab HC (Fd)- His7 tagged
DNA:
ATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGCAACTCTGTTAAGAGTTGTGAAGTGTCAG
GAACAACTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCGGGGGGGTCCCTGAGACTCTCC
TGTTTAGCGTCTGGATTCACGTTTCACAAATATGGCATGCACTGGGTCCGCCAGGCTCCA
GGCAAGGGCCTGGAGTGGGTGGCACTCATCTCAGATGACGGAATGAGGAAATATCATTC
AGACTCCATGTGGGGCCGAGTCACCATCTCCAGAGACAATTCCAAGAACACTCTTTATCT
GCAATTCAGCAGCCTGAAAGTCGAAGACACGGCTATGTTCTTCTGTGCGAGAGAGGCTGG
TGGGCCAATCTGGCATGACGACGTCAAATATTACGATTTTAATGACGGCTACTACAACTA
CCACTACATGGACGTCTGGGGCAAGGGGACCACGGTCACCGTCTCGAGCGCCTCCACCAA
GGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCC
CTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGC
GCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCC
TCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACG
TGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAAATCTTGTGAC
AAACATCATCACCATCACCACCAT
Protein:
MEFGLSWVFLATLLRVVKC
QEQLVESGGGVVQPGGSLRLSCLASGFTFHKYGMHWVRQAPGKGLEWVALISDDGMRKY
HSDSMWGRVTISRDNSKNTLYLQFSSLKVEDTAMFFCAREAGGPIWHDDVKYYDFNDGY
YNYHYMDVWGKGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSW
NSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSC
DKHHHHHHH
13) VRC01scFv–PG16 HC:
DNA:
GAATTCGCCGCCACCATGGGATGGTCATGTATCATCCTTTTTCTAGTAGCAACTGCAACCG
GTGTACATTCCCAGGTGCAGCTGGTGCAGTCTGGAGGTCAGATGAAGAAGCCTGGCGAGT
14
CGATGAGAATTTCTTGTCGGGCTTCTGGATATGAATTTATTGATTGTACGCTAAATTGG
ATTCGTCTGGCCCCCGGAAAAAGGCCTGAGTGGATGGGATGGCTGAAGCCTCGGGGGGGG
GCCGTCAACTACGCACGTCCACTTCAGGGCAGAGTGACCATGACTCGAGACGTTTATTCC
GACACAGCCTTTTTGGAGCTGCGCTCGTTGACAGTAGACGACACGGCCGTCTACTTTTGT
ACTAGGGGAAAAAACTGTGATTACAATTGGGACTTCGAACACTGGGGCCGGGGCACCCC
GGTCATCGTCTCATCAGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGGGAAGCGGAGGGG
GATCCGAAATTGTGTTGACACAGTCTCCAGGCACCCTGTCTTTGTCTCCAGGGGAAACAG
CCATCATCTCTTGTCGGACCAGTCAGTATGGTTCCTTAGCCTGGTATCAACAGAGGCCCG
GCCAGGCCCCCAGGCTCGTCATCTATTCGGGCTCTACTCGGGCCGCTGGCATCCCAGACA
GGTTCAGCGGCAGTCGGTGGGGGCCAGACTACAATCTCACCATCAGCAACCTGGAGTCGG
GAGATTTTGGTGTTTATTATTGCCAGCAGTATGAATTTTTTGGCCAGGGGACCAAGGTCC
AGGTCGACATTAAAGCCGGCGGCAGCGGAGGAGGATCTGGGGGAGGGTCCGGAGGCGGA
TCCGGGGGCGGTTCAGGTGGGGGTAGCGGAGGGGGCTCAGGCGGAGCTAGCCAGGAACA
ACTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCGGGGGGGTCCCTGAGACTCTCCTGTTT
AGCGTCTGGATTCACGTTTCACAAATATGGCATGCACTGGGTCCGCCAGGCTCCAGGCAA
GGGCCTGGAGTGGGTGGCACTCATCTCAGATGACGGAATGAGGAAATATCATTCAGACT
CCATGTGGGGCCGAGTCACCATCTCCAGAGACAATTCCAAGAACACTCTTTATCTGCAAT
TCAGCAGCCTGAAAGTCGAAGACACGGCTATGTTCTTCTGTGCGAGAGAGGCTGGTGGGC
CAATCTGGCATGACGACGTCAAATATTACGATTTTAATGACGGCTACTACAACTACCACT
ACATGGACGTCTGGGGCAAGGGGACCACGGTCACCGTCTCGAGCGCCTCCACCAAGGGCC
CATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGG
CTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCT
GACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGC
AGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAAT
CACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCCAAATCTTGTGACAAAAC
TCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTC
CCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTG
GTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGA
GGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGG
TCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGG
TCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGC
CCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGG
TCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGA
GCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCT
CCTTCTTCCTCTATAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCT
TCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCC
TGTCTCCGGGTAAATGAGCGGCCGC
Protein:
MGWSCIILFLVATATGVHS
QVQLVQSGGQMKKPGESMRISCRASGYEFIDCTLNWIRLAPGKRPEWMGWLKPRGGAVN
YARPLQGRVTMTRDVYSDTAFLELRSLTVDDTAVYFCTRGKNCDYNWDFEHWGRGTPVI
VSSGGGSGGGSGGGSGGGSEIVLTQSPGTLSLSPGETAIISCRTSQYGSLAWYQQRPGQAPR
LVIYSGSTRAAGIPDRFSGSRWGPDYNLTISNLESGDFGVYYCQQYEFFGQGTKVQVDIKAG
GSGGGSGGGSGGGSGGGSGGGSGGGSGGASQEQLVESGGGVVQPGGSLRLSCLASGFTFHK
YGMHWVRQAPGKGLEWVALISDDGMRKYHSDSMWGRVTISRDNSKNTLYLQFSSLKVE
15
DTAMFFCAREAGGPIWHDDVKYYDFNDGYYNYHYMDVWGKGTTVTVSSASTKGPSVFP
LAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVP
SSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVL
HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMH
EALHNHYTQKSLSLSPGK
16
Supplemental Fig. S2. Biosensor analyses of gp120 binding to immobilized VRC01
reagents. Sensorgrams (colored curves) of injected gp120 binding to VRC01 IA (A),
VRC01scFv-ZAG3-Fc (B), and VRC01 IgG (C) immobilized using covalently attached
protein A. Best fit binding curves (assuming a 1:1 binding model) are shown as thin
black lines.
17
Supplemental Fig. S3. Measured versus modeled IC50 values of VRC01scFv–PG16. One
parameter (PG16 or VRC01) or two parameter (combined fit; PG16 and VRC01) best
fits based on IC50s for PG16 IgG and VRC01 IgG. A 1:1 correlation between the
modeled IC50 and the actual IC50 data is indicated by the y = x line. The parameters
derived from the combined fit indicate that VRC01scFv–PG16 has ~1/3 of the activity
of PG16 IgG plus ~1/3 the activity of VRC01 IgG. The values of the correlation
coefficient R for the combined, PG16 only, and VRC01 only fits are 0.877, 0.825, and
0.414, respectively. A reduced vs. full model F-test comparing the PG16 only fit with
the 2-parameter fit demonstrates that the 2-parameter model is superior (F-test
analysis significance P<0.01).
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