Fig. 5.

DTT-dependent reversibility of inhibition of purified GAPDH protein. GAPDH from chicken muscle was incubated for 30 min with indicated compounds in Tris buffer (pH 7.4). A: Sper/NO (400 μM), decomposed Sper/NO (400 μM), and nitrite (NO₂⁻, 600 μM). Decomposed Sper/NO was obtained by incubating Sper/NO at neutral pH overnight. Nitrite concentration matched the nitrite concentration that was measured in the decomposed Sper/NO sample. B: CysNO (400 μM) and Sper/NO (400 μM). For the sample that was incubated with CysNO and Sper/NO, 30-min incubation with CysNO was followed by 30-min incubation with Sper/NO. C: Sper/NO (400 μM) and GSH (800 μM). For the sample that was incubated with Sper/NO and GSH, both compounds were added at the same time. The GAPDH activity was determined following the consumption of NADH at 340 nm in the presence and absence of DTT. For the assay with DTT, samples were pretreated with DTT for 30 min. Data represent means ± SE (n = 3 experiments).