Table 2.

Substitutions made in related bEBPs within the highly conserved GAFTGA motif^a

bEBP	Residue	Change	Description	Reference
NtrC	G215	V	Fails to activate transcription in vivo or in vitro	131
DctD	G220	D	Fails to activate transcription in vivo	214
NtrC	A216	V	Fails to activate transcription despite little reduction in ATPase activity	154
NtrC	A216	C	Sufficient ATPase; increased oligomerization state; binds enhancer, but activity is perturbed by DNA	229
NifA	A306	D/N	Fails to activate transcription in vivo	86
DctD	A221	V/D	Fails to activate transcription in vivo	214
NtrC	F217	L	Fails to activate transcription in vivo or in vitro	131
NifA	F307	A/L/I/P/R/H/N	Fails to activate transcription in vivo	86
NifA	F307	Y	<20% activity in vivo	86
DctD	F222	L	Fails to activate transcription in vivo/in vitro; ATPase 13% of WTΔNTD	214
DmpR	F312	L	Fails to activate transcription in vivo; ATPase 75-85% of WTΔNTD (ATP) and 23-25% WTΔNTD (dATP)	222
PspF	F85	A/E/R	<1% activity of WT in in vitro transcription assays; does not form ADP.AlFx-dependent trapped complex; decreased ATPase activity; defective for oligomerization	21
PspF	F85	C	<1% activity of WT in in vitro transcription assays; does not form ADP.AlFx-dependent trapped complex; decreased ATPase activity; oligomerizes in the presence/absence of nucleotide	234
PspF	F85	H/I/W	<1% activity of WT in in vitro transcription assays; does not form ADP.AlFx-dependent trapped complex; WT ATPase activity; nucleotide-dependent oligomerization (∼WT)	234
PspF	F85	L/Q	<1% activity of WT in in vitro transcription assays; does not form ADP.AlFx-dependent trapped complex; decreased ATPase activity; nucleotide-dependent oligomerization (∼WT)	234
PspF	F85	Y	<1% activity of WT in in vitro transcription assays; forms ADP.AlFx-dependent trapped complex; decreased ATPase activity; oligomerizes in the presence/absence of nucleotide; cannot form activator-DNA-σ54 complex (phenotype rescued by G4L substitution in σ54)	234
NorR	F264	Y	Partial “escape” from repression in vivo; shows WT-like response	36
NtrC	T218	A/N	Fails to activate transcription in vivo/in vitro	131
NifA	T308	A/L/M/P/R/V/G/C/S	Fails to activate transcription in vivo	86
DctD	T223	I	Fails to activate transcription in vivo/in vitro; ATPase 123% of WTΔNTD	214
DctD	T223	A	Fails to activate transcription; significant ATPase retained	214
PspF	T86	A	<1% activity of WT in in vitro transcription assays; wild-type ATPase activity	49
			Does not form ADP.AlFx-dependent trapped complex	21
PspF	T86	S	52% activity of WT in in vitro transcription assays; in vivo, ∼25% of WT	49
			Region IG4L σ54 substitution restores transcription activation activity	21
			WT ATPase activity	20
			Forms ADP.AlFx-dependent trapped complex	58
PspF	T86	V	<1% activity of WT in in vitro transcription assays; WT ATPase activity; does not form ADP.AlFx-dependent trapped complex	49
NtrC	G219	K	50% ATPase but fails to activate transcription; improved DNA binding	154
NtrC	G219	C	Increased ATPase due to increased oligomerization; binding of enhancer DNA prevents transcription	229
NorR	G266	D/N	Complete “escape” from repression in vivo; shows increased activity relative to that of WT	36
NorR	G266	C/S/Q/M	Partial “escape” from repression in vivo; shows WT-like activities	36
NorR	G266	K/R/P/A/E/L/V/T/H/I/W/Y/F	Fails to activate transcription in vivo	36
NtrC	A220	T	Fails to activate transcription despite little reduction in ATPase activity	154
NtrC	A220	V	Does not activate transcription in vivo but shows “hyperactivity” at low concentrations in vitro	131
NifA	A310	S	<20% activity in vivo	86
NifA	A310	N/D/G	Fails to activate transcription in vivo	86
DctD	A225	T	Fails to activate transcription in vivo/in vitro; reduced ATPase	214
DmpR	A315	T	Fails to activate transcription in vivo; WT ATPase activity	222

^aWT, wild type; NTD, N-terminal domain.