Table 1.

Catalytic activities of Trx Rs

Substrate	Specific activities of TrxR enzymes, μmol per min per mg
	Control TrxR1	(B)CAM-TrxR1	SeCys498 → Cys498 TrxR1
DTNB	42	0.6	2.7
TrxS2	13	0.07	1.5
Oxidized insulin, TrxS2	15	Not detectable	Not detectable

DTNB reduction was monitored spectrophotometrically at 412 nm in a 1-ml reaction mixture containing 100 mM potassium phosphate (pH 7.4), 2 mM EDTA, 1.5 mM DTNB, 0.2 mg of BSA, 0.2 mM NADPH, and 100–500 ng of native, 1–5 μg of (B)CAM-modified, or 1–5 μg of SeCys → Cys mutant TrxR1. NADPH-dependent reduction of TrxS₂ was monitored spectrophotometrically at 340 nm in reaction mixtures containing 120 μM TrxS₂, 50 mM potassium phosphate (pH 7.0), 1 mM EDTA, and 0.2 mM NADPH. Activities of TrxR were also measured by coupling the NADPH-dependent reduction of Trx and oxidized insulin in reaction mixtures containing 7.5 μM TrxS₂, 80 μM oxidized insulin, 50 mM potassium phosphate (pH 7.0), 1 mM EDTA, 0.2 mM NADPH, and 1 μg of TrxR1.