Table 1. Properties of variants with substitution of conserved histidine, lysine and tyrosine residues in the C-terminal catalytic domain of the murine CAD.

Residue	Substitution	ICAD-L bindinga	Relative DNA cleavage activity (%)b	Oligomeric state	DNA bindingc	Remark
Wild-type	–	+	100	wt	wt
Lys155	Gln	+	n.d.c.	wt	wt
Lys301	Gln	+	1.4	wt	wt
Lys310	Gln	+	2.3	wt	wt
Tyr170	Phe	+	107.0	wt	wt	Fragment pattern
Tyr247	Phe	+	1.2	wt	wt
His127	Asnd	+	25.8	wt	n.d.e
His242	Asnd	+	6.3	Deviating	n.d.e	Fragment pattern
	Glu	+	<1.0	wt	wt
	Arg	+	<1.0	wt	wt
His263	Asnd	+	0.7	wt	n.d.e
	Asp	+	<1.0	wt	wt
	Arg	+	<1.0	wt	wt
His304	Asnd	+	14.5	wt	n.d.e
His308	Asnd	+	9.3	wt	n.d.e	Fragment pattern
	Asp	+	∼1–2	wt	wt	Residual activity at pH 5.0
	Arg	+	<1.0	wt	wt
His313	Asnd	+	1.3	wt	n.d.e
	Asp	+	<1.0	wt	wt
	Arg	+	109.0	wt	wt	Fragment pattern

n.d., not determined; n.d.c., no detectable cleavage; wt, wild-type-like.

^aICAD-L binding measured by co-purification of untagged hICAD-L with GST-mCAD.

^bMeasured by the disappearance of supercoiled plasmid DNA in steady state cleavage experiments.

^cBinding of free GST-mCAD to DNA–cellulose.

^dThe properties of these variants were described in Meiss et al. (17).

^eSakahira et al. (16) have demonstrated DNA binding for the alanine substitutions at the indicated positions in murine CAD. With the exception of H242A, all other variants bound to DNA–cellulose, similar to wild-type CAD.