Table 2.

Mitochondrial respiration and ATP synthesis/hydrolysis.

Strain	Respiration Rates nmol O/min/mg				ATP Synthesis Rate nmol Pi/min/mg		ATPase Activity µmol Pi/min/mg			P/O
	NADH	NADH + ADP	NADH + CCCP	Asc/TMPD + CCCP	-oligo	+oligo	-oligo	+oligo	Inhib %
WT	329 ± 3	619 ± 99	1548 ± 113	2865 ± 325	1531 ± 168	151 ± 30	4.7 ± 0.2	0.5 ± 0.1	88	1.21 ± 0.18
aL242P	96 ± 8	79 ± 11	242 ± 48	440 ± 26	142 ± 43	96 ± 34	3.3 ± 0.4	3.2 ± 0.2	4	0.89 ± 0.20
aL242S	192 ± 21	353 ± 23	793 ± 60	1443 ± 264	866 ± 170	170 ± 31	4.2 ± 0.1	0.6 ± 0.2	85	1.24 ± 0.19
aL242T(CTA)	220 ± 13	410 ± 22	949 ± 47	1704 ± 75	1056 ± 90	164 ± 32	4.2 ± 0.5	0.5 ± 0.2	88	1.25 ± 0.17
aL242T(ACA)	224 ± 28	462 ± 84	1133 ± 127	2101 ± 316	1066 ± 89	160 ± 18	4.0 ± 0.2	0.5 ± 0.1	87	1.06 ± 0.17

Mitochondria were isolated from wild type MR6 strain and mutant strains aL242P, aL242S and aL242T (with either the CTA or ACA threonine codon) grown for 5-6 generations in rich galactose medium (YPGalA) at 28 °C. Reaction mixes for assays contained 0.15 mg/mL protein, 4 mM NADH, 150 µM ADP (for respiration assays), 1 mM ADP (for ATP synthesis assays), 12.5 mM ascorbate (Asc), 1.4 mM TMPD, 4 µM CCCP, 4 µg/mL oligomycin (oligo). The values reported are averages of triplicate assays ± standard deviation. Respiration and ATP synthesis activities were detected on freshly isolated and osmotically protected mitochondria in the respiratory buffer (see Section 4, Materials and Methods). The ATPase assays were performed from mitochondrial samples that had been frozen at −80 °C in the absence of osmotic protection, in the ATPase buffer (see Materials and Methods).