Extended Data Fig. 6: LDHB has a small effect on the total LDH activity in H1299 cells. Related to Fig. 6.
a. The IC50 of OAA against human LDHB determined using purified recombinant human LDHB and analysis of 13C1 lactate derived from 13C1 pyruvate. The IC50 was identified as 1,021 ± 86 μM. b. Western blot of H1299 lysate and increasing concentrations of purified recombinant LDHB. Corresponding standard curve constructed from the average pixel density of LDHB purified recombinant protein band and used to determine the amount of LDHB protein in H1299 lysate. c. LDH activity, determined using a fluorescence based LDH activity assay using diluted lysates from H1299 vector and LDHB knockdown cells. The corresponding western blots of the H1299 lysates used in the above activity assay with antibodies against LDHB and β-actin. Data is represented as the mean and error bars represent the standard deviation from n = 4 for (a), n = 3 for (c) of independent replicates in vitro. Standard curve in (b) is a representative experiment of 3 independent replicates in vitro. Western blots in (b) (c) are representative experiments of 3 biologically independent replicates. P values were determined by a two-tailed Student’s t-test.
