20240305 Gray Figure 3 data

FIG 3 Copper, zinc, and nickel inhibit RclA activity in vitro. Reactions (500 µl) contained 10 mM HEPES-KOH buffer (pH 7.4), 160 µM NADH, 100 µM HOSCN, and the indicated concentrations of metal salts, and were started by addition of 10 nM RclA. I quantified NADH consumption at A340 for 1 min at 20°C and calculated specific activities as µmol NADH consumed min-1 mg-1 RclA (n=3-4 experimental replicates; error bars of 1 standard deviation). Asterisks indicate activities significantly different from that of RclA with no metals added in a given set of experiments (one-way ANOVA with Dunnett’s multiple comparisons test; ** = P<0.01. *** = P<0.001, **** = P<0.0001). X symbols indicate that RclA precipitated at 100 µM CuCl2.

图3 铜、锌与镍在体外抑制RclA活性。反应体系总体积为500 μL，包含10 mM HEPES-KOH缓冲液（pH 7.4）、160 μM 烟酰胺腺嘌呤二核苷酸（NADH）、100 μM HOSCN以及不同浓度的金属盐，通过添加10 nM RclA启动反应。本实验于20℃下在340 nm波长处监测1分钟内NADH的消耗量，并以每毫克RclA每分钟消耗的NADH微摩尔数（μmol NADH·min⁻¹·mg⁻¹）计算比活性；实验重复次数为3至4次，误差棒代表1倍标准差。星号代表在对应实验分组中，该活性与未添加金属的RclA活性存在显著差异（采用单因素方差分析（one-way ANOVA）结合Dunnett多重比较检验（Dunnett’s multiple comparisons test）；**表示P<0.01，***表示P<0.001，****表示P<0.0001）。X符号代表当氯化铜浓度为100 μM时，RclA发生沉淀。