(A) Top line, SDS-PAGE and Western blot analysis of purified PykF and its acetylated variants from BL21 (DE3) cells. The acetylated variants with mutation of a single lysine site in PykF to glutamine. Lane 1, wild-type PykF; Lane 2, PykF-K13Q; Lane 3, PykF-K19Q; Lane 4, PykF-K52Q; Lane 5, PykF-K59Q; Lane 6, PykF-K68Q; Lane 7, PykF-145Q; Lane 8, PykF-K317Q; Lane 9, PykF-K319Q; Lane 10, PykF-K340Q; Lane 11, PykF-K368Q; Lane 12, PykF-K382Q. The same amount of protein was loaded. Anti-AcK: anti-acetyl lysine antibody. Bottom line, pyruvate kinase activity of PykF and its acetylated variants. Pyruvate kinase activity of the wild-type PykF was set as 1. Mean values and standard deviations were calculated based on three replicates (n = 3). (B) Top line, SDS-PAGE and Western blot analysis of purified PykF and its deacetylated variants from BL21 (DE3) cells. The deacetylated variants with mutation of a single lysine site in PykF to arginine. Lane 1, wild-type PykF; Lane 2, PykF-K13R; Lane 3, PykF-K19R; Lane 4, PykF-K52R; Lane 5, PykF-K59R; Lane 6, PykF-K68R; Lane 7, PykF-145R; Lane 8, PykF-K317R; Lane 9, PykF-K319R; Lane 10, PykF-K340R; Lane 11, PykF-K368R; Lane 12, PykF-K382R. The same amount of protein was loaded. Anti-AcK, anti-acetyl lysine antibody. Bottom line, pyruvate kinase activity of PykF and its deacetylated variants. Pyruvate kinase activity of the wild-type PykF was set as 1. Mean values and standard deviations were calculated based on three replicates (n = 3). Two-tailed P-values were determined by the t-test, and the significance level is 0.05. *p < 0.05.
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