TABLE 3.

DNA binding affinity of Ets domain constructs (Etv1 unless specified)

NA indicates undetectable binding or below threshold for regression analysis. K_d^app1 indicates apparent K_d value measured with a 2 nm DNA probe. K_d^app2 indicates apparent K_d value measured with a 0.1 nm DNA probe.

Construct (Etv1 unless specified otherwise)	Description/mutation	Kdapp1 (×10−10 m)a	Kdapp1-fold inhibitionb	Kdapp2 (× 10−10 m)a	Kdapp2-fold inhibitionb
WT	Wild type Ets domain	2.7 ± 0.28	1	5.9 ± 1.5	1
Y329S-P427S	PCR mutant Ets domain	2.5 ± 0.26	0.9	3.1 ± 0.6	0.5
K379A	DNA backbone contact	NA	NA
D387A	DNA base contact	1.7 ± 0.2	0.6	1.9 ± 0.3	0.3
R391A	DNA base contact	118 ± 14.8	43.7	86.0 ± 16.8	14.6
R394A	DNA base contact	2757 ± 627	1021	ND	ND
Y395F	DNA base contact	2.3 ± 0.5	0.9	13.5 ± 10.5	2.3
Y396F	DNA backbone contact	26390 ± 9349	9774.1
Y397F	DNA backbone contact	NA	NA
K404A	DNA backbone contact	NA	NA
S334E	Phosphoserine mimic	8.9 ± 3.2	3.3	9.4 ± 5.1	1.6
WT (phosphorylated)	PKA-phosphorylated Ser-334	521 ± 455	193
C416S	Abrogation of disulfides	4.6 ± 0.6	1.7	2.6 ± 0.6	0.4
WT dimer	Disulfide-linked dimeric WT ETV1	195 ± 148	72.2	802 ± 164	136
WT dimer (TCEP reduced)	Dimeric WT ETV1, reduced	11.2 ± 2.6	4.1	2.6 ± 0.4	0.4
WT dimer (9:1 GSH:GSSG)	Dimeric WT ETV1, reduced			171 ± 23	29
WT dimer (1:9 GSH:GSSG)	Dimeric WT ETV1, reduced			272 ± 30	47
Etv4 WT (reduced)	Wild type Ets domain, reduced			3.4 ± 0.66	1
Etv4 WT (oxidized)	Disulfide-linked dimeric WT ETV4			148 c ± 35	44
Etv5 WT (reduced)	Wild type Ets, reduced			11.6 ± 1.1	1
Etv5 WT (oxidized)	Disulfide-linked dimeric WT ETV5			790 ± 610c	68

^a K_d ± S.E.

^b Fold increase in K_d value compared with the corresponding wild type.

^c Estimated, due to binding at regression analysis threshold.