Nickel quercetinase, a “promiscuous” metalloenzyme: metal incorporation and metal ligand substitution studies

BMC Biochemistry

Table 2 Metal contents and activities of QueD proteins carrying ligand replacements, and K _D values of protein-quercetin complexes

Protein ^a	Metal content (equivalents per protein monomer)	Spec. activity (U mg ⁻¹ )	K _D (μM)
Ni-QueD-H69A	0.88 Ni, 0.07 Fe	b.d.^b	3.1 (0.9)
Ni-QueD-H115A	0.34 Ni, 0.29 Zn, 0.07 Fe, 0.02 Co, 0.01 Mn	b.d.	2.6 (0.6)
Ni-QueD-H71A	0.18 Ni, 0.16 Zn, 0.06 Fe	0.90 (0.04)	10.4 (2.9)
Ni-QueD-E76D	0.28 Ni, 0.24 Zn, 0.05 Fe	1.32 (0.01)	4.0 (1.0)
Ni-QueD-E76H	0.54 Ni, 0.03 Cu, 0.02 Fe, 0.01 Zn	3.44 (0.13)	1.3 (0.1)
Co-QueD-E76H	0.6 Co, 0.03 Cu, 0.02 Fe, 0.01 Zn, 0.02 Ni	0.19 (0.01)	n.d.^c

^aThe designations of the QueD metal forms refer to the metal of interest and do not imply exclusive occupancy of the protein with this metal.
^bb.d., below detection.
^cn.d., not determined.
Metal ions which contribute to ≥0.01 equivalents per protein monomer are listed. For the specific activities and K _D values, the average of 3 experiments is given with the standard deviations in brackets. The K _D value of the complex of Ni-QueD with quercetin, determined for the preparation specified in Table 1, was 10.1 ± 2.9 μM.

ISSN: 1471-2091