Figure 5.
Comparison of PL1-Labeling Profiles in Various Human Cell Lines
(A–C) (A) Heatmap illustrating the corresponding fold changes (log2PL1/PL) of PLP-DEs in the different cell lines. The map depicts only PLP-DEs that are significantly enriched in at least one cell line. In the case that a PLP-DE was not detected in samples of a certain cell line, the corresponding rectangle is colorless. Western blots against the PLP-DEs ALAS1, PLPBP, SHMT1, and SHMT2 (B), as well as vitamin B6 salvage enzymes PNPO and hPLK (C) revealed a similar expression of corresponding proteins in all cell lines studied.
(D) PLP-ome analysis using an anti-PL antibody raised against reduced cofactor in conjugation with BSA. Antibody specificity was validated upon incubation with hPLK (1), which does not covalently bind PLP, NaBH4-reduced forms of PLPBP (2), and SHMT1 (3), as well as the non-reduced PLPBP (4) and SHMT1 (5) counterparts (left). Western blot analysis of cofactor loading states of PLP-DEs in NaBH4-treated as well as not reduced proteomes (right).
See also Figures S3–S5 and Table S1.