Figure 2
From: The unusual properties of lactoferrin during its nascent phase
Reactivity of rLF cysteines toward DTNB and alkylating reagents. (A) TNBS¯ release after reaction of rLF (0.6 μM, 19.2 μM protein –SH) with sub-stoichiometric DTNB concentrations: 0.6 μM (ratio rLF:DTNB = 1:1, grey line); 1.2 μM (ratio rLF:DTNB = 1:2, green line); 1.8 μM (ratio rLF:DTNB = 1:3, dark cyan line); 2.4 μM (ratio rLF:DTNB = 1:4, blue line). The reactions were done in 0.2 M urea, 0.1 M sodium acetate buffer pH 5.0 at 25 °C. (B) Representative reaction scheme of rLF with sub-stoichiometric DTNB. (C) TNBS¯ release after reaction of rLF (0.6 μM, 19.2 μM protein –SH) with DTNB 9.6 μM (ratio rLF:DTNB = 1:16, red line). The reaction was done in 0.2 M urea, 0.1 M sodium acetate buffer pH 5.0 at 25 °C. (D) Second order kinetic constants of cysteines (krLF SH) normalized to the rate constants for the free GSH (kfree thiol). The number of protein cysteine per mole with a given reactivity is reported on the top of each column. Error bars represent the propagation of uncertainties for the quotients21. (E) Inhibition of the reactivity of cysteines in rLF (0.6 μM) toward DTNB (50 μM) by variable urea concentrations at pH 5.0 at 25 °C. The error bars represent the SD from three independent experiments.
