Table 1 List of the target triplets, amino acid residues at the position −1 ~ +6 of the α-helix of each single-zinc-finger domain of active 7ZFPs and their binding activity
Active 7ZFPs and their variants marked with the line number | p16 DNA binding activitya | N-terminal Sp1three-zinc-finger domain (3ZF) and its mutants | Middle Sp1-liketwo-zinc-finger modules (2ZF) | C-terminal Sp1-liketwo-zinc-finger modules (2ZF) | C-terminal linkerbetween finger-7& 6 × His-tag | ||||
|---|---|---|---|---|---|---|---|---|---|
finger-1 | finger-2 | finger-3 | finger-4 | finger-5 | finger-6 | finger-7 | |||
5′-ggg-3′ | 5′-gcg-3′ | 5′-ggg-3′ | 5′-aac-3′ | 5′-gga-3′ | 5′-gaa-3′ | 5′-gag-3′ | |||
● Finger motif replacement | |||||||||
1_ Active-A (consensus)b | (++) | KTSHLRA | RSDELQR | RSDHLSK | ESDNLSQ | QSGHLQR | QSSNLQR | RSDNLAR | TGEKP |
2_Variant-A (f3) | (++) | −−−−−−− c | −−−−−−− | Q−G−−QR | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−− |
3_ Active-B | (++) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−T− | −−−−− |
4_Variant-B (f3) | (++) | −−−−−−− | −−−−−−− | Q−G−−QR | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−T− | −−−−− |
5_Variant-B (f6f7) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−R− | Q−GH−Q− | −−−−− |
6_ Active-C | (++) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−T−−−− | −−−−−−− | −−−−−T− | −−−−− |
7_Variant-C (f6f7) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−T−−−− | D−G−−R− | Q−GH−Q− | −−−−− |
● Single key amino acid mutation | |||||||||
8_ Active-D | (+) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−V− | −−−A−Q− | −−−−− |
9_Variant-D (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−R−−V− | −−−A−Q− | −−−−− |
10_ Active-F | (++) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−T−−−− | −−−−−V− | −−−−−T− | −−−−− |
11_Variant-F (f6)d | (−) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−T−−−− | −−R−−V − | −−−−−T− | −−−−− |
12_ Active-G | (++) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RV | −−−−−−− | −−−−−V− | −−−−−T− | −−−−− |
13_Variant-G (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RV | −−−−−−− | −−R−−V − | −−−−−T− | −−−−− |
14_ Active-Hc | (+) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RV | −−−−−−− | −−−−−V− | −−−A−Q− | −−−−− |
15_Variant-H (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RV | −−−−−−− | −−R−−V− | −−−A−Q− | −−−−− |
16_ Active-I | (++) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−−−−−− | −−−−−V− | −−−−−T− | −−−−− |
17_Variant-I (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−−−−−− | −−R−−V − | −−−−−T− | −−−−− |
18_ Active-J | (+) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−T−−−− | −−−−−V− | −−−A−Q− | −−−−− |
19_Variant-J (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−T−−−− | −−R−−V− | −−−A−Q− | −−−−− |
20_ Active-K | (++) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−T−−−− | −−−−−V− | −−−−−T− | −−−−− |
21_Variant-K (f6) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | D−G−−RR | −−T−−−− | −−R−−V − | −−−−−T− | −−−−− |
● C -terminal linker deletion | |||||||||
22_ Active-K | (++) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−V− | −−−−−T− | −−−−− |
23_Variant-K (linker) | (−) | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−−− | −−−−−V− | −−−−−T− | TG |