Table 1 Stable enzyme set.

	Enzyme	Organism	Native/Mutant	Activity (units)^b	Half-life in 8% isobutanol (hours)
TmHex	Hexokinase^a	T. maritima	Native	14.5 ± 0.3	>>96
TmPgi	Glucose-6-phosphate isomerase^a	T. maritima	Native	29.6 ± 1.1	>>113
EcPfkB	Phosphofructokinase B	E. coli	Native	8.3 ± 0.1	55/123^d
TtFba	Fructose-1,6-bisphosphate aldolase^a	T. thermophilus	Codon-optimized	4.7 ± 0.10	>>117
TmTpi	Triosephosphate isomerase^a	T. maritima	Native	210 ± 0.7	>>100
TkGapN’	Glyceraldehyde-3-phosphate dehydrogenase (non-phosphorylating)^a	T. kodakarensis	Designed Codon-optimized	7.6 ± 0.8	>>117
AfGapDH	Glyceraldehyde-3-phosphate dehydrogenase (phosphorylating)^a	A. fulgidus	Native	10.9 ± 0.6	>>100
TmPgk	Phosphoglycerate kinase^a	T. maritima	Native	8.4 ± 0.5	>>100
Gs iPgm	Phosphoglycerate mutase (2,3-bisphosphoglycerate independent)^a	G. stearothermophilus	Native	49.4 ± 2.2	>>117
TtEno	Phosphoenolpyruvate hydratase^a	T. thermophilus	Codon-optimized	117 ± 5	>>117
TtPyk	Pyruvate kinase^a	T. thermophilus	Codon-optimized	23.8 ± 0.8	>>139
BsAlsS-P	Acetolactate synthase	B. subtilis	PROSS^c Codon-optimized	4.9 ± 0.2	>>115
GsIlvC	Ketol-acid reductoisomerase^a	G. stearothermophilus	Native	0.9 ± 0.1	>>123
SmIlvD	Dihydroxyacid dehydratase	S. mutans	Native	2.7 ± 0.2	125
KivD-S	Alpha-ketoisovalerate decarboxylase	L. lactis	PROSS^c/DE Codon-optimized	18.0 ± 1.6	123
EcYahK	Aldehyde reductase	E. coli	Native	4.5 ± 0.4	138

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