TABLE 3.

Binding of fluorescent nucleotides to wt ClpB and selected mutants.

Titrations were performed under experimental conditions that modify the oligomeric state of ClpB. Experiments were repeated at least two times. ND, not determined. −, binding of nucleotide analogs to T1N2 was not detected.

ClpB variant	MANT-ATP		MANT-ADP
	Kd	nH	Kd	nH
	μm		μm
Wt	3.6 ± 0.2a	1.7 ± 0.1a	3.4 ± 0.2b	1.7 ± 0.1b
	4.6 ± 0.2b	1.4 ± 0.1b
	3.2 ± 0.2c	1.4 ± 0.1c
T1T2	3.3 ± 0.2a	1.6 ± 0.1a	3.2 ± 0.3b	1.8 ± 0.2b
	4.4 ± 0.3b	1.3 ± 0.1b
	4.9 ± 0.4c	1.1 ± 0.1c
T1N2	–	–	–	–
N1T2	3.3 ± 0.1a	1.6 ± 0.1a	ND	ND
	4.4 ± 0.3b	1.4 ± 0.1b
	3.3 ± 0.1c	1.4 ± 0.1c

^a 10 μm protein in 15 mm KCl.

^b 10 μm protein in 50 mm KCl buffer.

^c 5 μm protein in 500 mm KCl buffer.