Extended Data Fig. 3. Hem25p function is required for CoQ biosynthesis.

a, Relative PPHB abundances in hem25Δ yeast carrying WT and mutant HEM25^FLAG constructs under the control of the endogenous HEM25 promoter. Levels are relative to WT yeast carrying the empty expression vector (*p = 0.0011, **p = 0.0006, ***p = 0.0116, ****p = 0.0009 HEM25^FLAG vs mutants or empty vector, mean ± SD, n = 3 biologically independent samples, two-sided Student’s t-test). b, c, Immunoblots of whole cell lysates (b) or isolated mitochondria (c) from WT, coq6Δ, or hem25Δ cells expressing various EV and Hem25p-FLAG constructs under the control of the endogenous HEM25promoter. d, e, Relative CoQ (d) and PPHB (e) abundances in hem25Δ yeast carrying WT and mutant HEM25^FLAG constructs under the control of the constitutive GPD promoter. Levels are relative to WT yeast carrying the empty expression vector (*p = 0.0006, **p = 0.0005, ***p = 0.0009, ****p = 0.0019, ^#p = 0.0012, ^##p = 0.0014, ^###p = 0.0012, ^####p = 0.0012 HEM25^FLAG vs mutants or empty vector, mean ± SD, n = 3 biologically independent samples, two-sided Student’s t-test). f, g, Immunoblots of whole cell lysates (f) or isolated mitochondria (g) from WT, coq6Δ, or hem25Δ cells expressing various EV and Hem25p-FLAG constructs under the control of the constitutive GPD promoter. h, Time course of 50 μM [1-¹⁴C]-IPP uptake by E. coli cells expressing WT MBP-Hem25p, mutant MBP-Hem25p, or the empty vector (*p = 0.0494, **p = 0.0023, ***p = 0.0136, ****p = 0.0063 empty vector vs WT, mean ± SD, n = 3 biologically independent samples, two-sided Student’s t-test). i, Immunoblot for WT and mutant MBP-Hem25p in E. coli membranes preparations. j, k Normalized abundance of unlabeled OPP (j) and CoQ₈ (k) in E. coli cells expressing the EV, WT MBP-Hem25p, or the R181P MBP-Hem25p mutant (mean ± SD, n = 3 biologically independent samples). Source numerical data and unprocessed blots are available in source data.