Fig. 2. The deletion of COQ9 in a yeast coq5-5 point mutant leads to the accumulation of 3-hexaprenyl-4-aminophenol (4-AP) and the disappearance of demethyl-demethoxy-Q₆ (DDMQ₆), but imino-demethyl-demethoxy-Q₆ (IDDMQ₆) is still present.

Yeast coq5-5 point mutants with COQ8 over-expressed (coq5-5:mcCOQ8), without COQ8 over-expressed (coq5-5), or with the COQ9 gene deleted and COQ8 over-expressed (coq5-5 coq9Δ:mcCOQ8), were cultured in SD complete or SD−Ura with 50 μg/ml ¹³C₆-pABA and 2 μl ethanol/ml medium at 0.5 A_600nm/ml and collected after 6 hours. Q₄ (145.4 pmol) was added prior to extraction to serve as an internal standard. Lipid extracts prepared from the cell pellets were analyzed by RP-HPLC-MS/MS. Multiple reaction monitoring (MRM) detected precursor-to-product ion transitions 552.4/158.0 (¹³C₆ –IDDMQ₆), 553.4/159.0 (¹³C₆-DDMQ₆), and 524.4/128.0 (¹³C₆-4-AP). Just the oxidized forms of IDDMQ₆ and DDMQ₆ were detected, while only the reduced form of 4-AP was present. ¹³C₆-DDMQ₆ accumulates in the coq5-5 and coq5-5:mcCOQ8 yeast mutants (B), and ¹³C₆-IDDMQ₆ is readily detected (A). ¹³C₆-IDDMQ₆ and ¹³C₆-4-AP accumulate in coq5-5 coq9Δ:mcCOQ8 (A and C), but ¹³C₆-DDMQ₆ is not detected (B). In all panels, the blue traces designate the Q-intermediate signals in coq5-5 and green traces designate the Q-intermediate signals in coq5-5:mcCOQ8, and the red traces indicate the Q-intermediate signals in coq5-5 coq9Δ:mcCOQ8.