TABLE 1.

Purification of components of CARDO

Component	Vol (ml)	Total activitya (U)	Total protein (mg)	Sp acta (U/mg)	Yield (%)	Purifi- cation (fold)
His-tagged CarAd
Crude extract	87	28	510	0.055	100	1
HiTrap Chelating	9.1	16	12	1.3	57	24
Sephacryl S-200	3.4	12	4.8	2.4	43	44
His-tagged CarAc
Crude extract	100	400	120	3.3	100	1
HiTrap Chelating	11	330	6.5	51	85	16
Sephacryl S-200	3.2	220	1.5	150	56	45
CarAa
Crude extract	200	1,700	300	5.5	100	1
DEAE	11	440	20	21	26	4
HQ/L (1)b	12	330	3.8	86	20	16
HQ/L (2)b	1.7	310	2.3	130	19	24
Sephacryl S-200	1.5	190	1.7	110	12	21

^aFor the NADH:cytochrome c oxidoreductase activity of His-tagged CarAd, 1 U of activity was defined as the amount of the enzyme required to reduce 1 μmol of electron acceptor per min at 30°C. For CARDO system activities to determine CarAa and His-tagged CarAc activities, 1 U of activity was defined as the amount of the enzyme required to degrade 1 nmol of CAR per min at 30°C.

^bThe HQ/L column was used twice with different buffer systems. HQ/L (1) was performed with an NaCl gradient in Tris-HCl buffer, and HQ/L (2) was performed with an Na₂SO₄ gradient in Tris-H₂SO₄ buffer. Both buffers contained 10% glycerol.