n-Butanol production in Saccharomyces cerevisiae is limited by the availability of coenzyme A and cytosolic acetyl-CoA

Table 2 In vitro activities of various enzymes involved in n-butanol production pathways expressed in S. cerevisiae

Enzyme	Activity [U/mg] high copy vector	Activity [U/mg] low copy vector
Thiolase
^Sc ERG10	0.043 ± 0.009	0.10 ± 0.005
^Ca thlA	0.15 ± 0.04	4.96 ± 0.4
None	0.003 ± 0.0003	0.004 ± 0.002
Acetyl-CoA:malonyl-CoA acyltransferase
^Ss nphT7	NM	0.12 ± 0.009
None	NM	0.002 ± 0.001
3-Hydroxybutyryl-CoA DH
^Ca hbd	0.31 ± 0.04	1.47 ± 0.19
None	0.011 ± 0.004	0.063 ± 0.01
Crotonase
^Ca crt	3.5 ± 1.07	35.64 ± 3.2
None	0.17 ± 0.04	4.09 ± 2.2
Trans-2-enoyl-CoA reductase
^Eg ter	ND	0.002 ± 0.001
^Td ter	ND	0.0103 ± 0.002
None	ND	ND
Crotonyl-CoA reductase
^Sco ccr	NM	0.001 ± 0.00009
None	NM	ND
Butyraldehyde-DH
^Ca adhE2 (pVS1)	0.00027 ± 0.00002	0.0039 ± 0.003
^Ca adhE2 + ^Ec eutE (pVS6)	0.00091 ± 0.0004	0.06 ± 0.009
^Ec eutE	0.014 ± 0.0083	0.044 ± 0.004
None	ND	0.0022 ± 0.0009
Butanol-DH
^Ca adhE2 (pVS1)	0.07 ± 0.02	0.062 ± 0.015
^Ca adhE2 (pVS6)	0.053 ± 0.02	0.076 ± 0.005
^Ca bdhB	0.073 ± 0.03	0.044 ± 0.008
None	0.049 ± 0.01	0.04 ± 0.009

The mean values of three independent replicates are shown with standard deviations. One unit (U) is defined as the conversion of 1 µmol of substrate into the corresponding product per 1 min. The term “none” refers to the empty vector control, which means for the high copy variants (2 µ) pRS62K and for the low copy variants (CEN6/ARS4) pRS41K. Genes from Saccharomyces cerevisiae (Sc), Clostridium acetobutylicum (Ca), E.coli (Ec), Euglena gracilis (Eg), Streptomyces collinus (Sco), Streptomyces sp. CL190 (Ss), and Treponema denticola (Td) are indicated by prefixes in superscript
ND not detectable, NM not measured

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