Improvement of L-phenylalanine production from glycerol by recombinant Escherichia coli strains: The role of extra copies of glpK, glpX, and tktA genes

Table 3 Enzyme activities of fructose-1,6-bisphosphatase II and glycerol kinase in the crude extract

Strain	FBPase activity [U mg crude extract ⁻¹ ]	Glycerol kinase activity [U mg crude extract ⁻¹ ]
		0 mM FBP	10 mM FBP
FUS4	0.53 ± 0.21	-	-
FUS4.6	0.97 ± 0.24	-	-
DH5α pJF119	0.42 ± 0.22	1.5 ± 0.3	0.2 ± 0.1
DH5α pKUS13	3.05 ± 0.62	-	-
DH5α pKUS3	-	52.7 ± 4.1	11.2 ± 0.9
DH5α pKUS3-1	-	64.4 ± 3.6	43.3 ± 3.8

Cells were grown and induced with IPTG as described in Materials and methods. FBPase and glycerol kinase activity was measured in cell-free extracts. DH5α/pKUS13 (carrying gene glpX) was compared to the control, DH5α/pJF119EH (empty vector). DH5α/pKUS3 (carrying gene glpK) and DH5α/pKUS3-1 (carrying gene glpK ^G232D) were also compared to the control, DH5α/pJF119EH. After chromosomal integration of glpX FBPase activity was measured in the cell-free extract of the resulting strain FUS4.6 and as control in the parent strain, FUS4. FBPase activity was measured by quantifying phosphate release using a malachite green/ammonium molybdate solution and glycerol kinase activity was measured using a pyruvate kinase and lactate dehydrogenase coupled assay as described in Materials and methods [50].
The values represent the mean value and the standard deviation.

ISSN: 1475-2859