TABLE 3.

Apparent kinetic constants of cellobiose dehydrogenase from S. rolfsii for several electron acceptors^a

Electron acceptor	Wavelength monitored (nm)	Extinction coefficient (mM−1 · cm−1)	pH optimum	Stoichiometryb	CDH kinetic constants			CBQ relative activities
					Km (μM)	kcat (s−1)	kcat/Km (mM−1 · −1)
DCIP	520	6.8	3.2–4.8c	1	15	30	2,000	0.95
1,4-Benzoquinone	290	2.24	3.0–4.2	1	25	30	1,200	n.d.
2,6-Dimethyl-1,4-benzoquinone	330	0.37	4.0–5.0	1	550	15	27	0.3
2,6-Dimethoxy-1,4-benzoquinone	395	0.58	3.5–4.0	1	n.d.	5e	n.d.	0.1
3,5-Di-tert-butyl-1,2-benzoquinone	420	1.4d	3.0–4.0	1	53	23	434	0.5
Methylene blue	620	34.3	3.1–3.6	1	4.4	18	4,100	0.04
Methylene green	654	38.1	3.4–3.8	1	4.3	18	4,190	0.15
Meldola's blue	640	5.6	3.0–3.8	1	31	24	800	0.15
ABTS cation radicalf	436g	29.3	4.0	2	0.4	27	67,500	n.d.
Cyt c (horse heart)	550	19.6h	3.5	2	0.3	34	113,000	0
K3Fe(CN)6	420	0.98	2.7–3.4	2	20	37	1,850	1

^aConstants were determined at 25°C using 2 mM cellobiose as the substrate and at the pH optimum of the respective acceptor. Protein was determined using the BCA method. n.d., not determined. 

^bMolecules of electron acceptor reduced per molecule of cellobiose oxidized. 

^cFor both CDH and CBQ. 

^dDatum is from Roy et al. (43). 

^eMaximum activity. 

^fThe cation radical was prepared by oxidizing ABTS with laccase, the enzyme was removed by ultrafiltration, and the radical was quantified with the help of the extinction coefficient. 

^gDatum is from Niku-Paavola et al. (40). 

^hDatum is from Canevascini et al. (11).