Integration of E. coli aroG-pheA tandem genes into Corynebacterium glutamicum tyrA locus and its effect on L-phenylalanine biosynthesis

Table 1 Strains and plasmids

Strains or plasmids	Relevant characteristics	Sources or references
E.coli JM110	Strain for vectors construction	Our laboratory
C.glutamicum	Nxr, original strainmutagenesis	ATCC
C.glutamicum FP	FPr, from C.glutamicum ATCC13032	This work
C.glutamicum KM	C. glutamicum FP inserted with Km gene	This work
C.glutamicum GAK	C. glutamicum FP inserted with GAK	This work
pBluscript SK (pSK)	Apr, vector for gene cloning	Our laboratory
pPR	Apr, pBR322 derivative, suicidal vector	Our laboratory
pGA	Kmr, carrying PBF-aroG-pheA-T fragment	Ref.15
pT	pPR carrying tyrA gene	This work
pTK	pT carrying Km gene(tyrA::Km)	This work
pSKGAK	pSK carrying GAK fragment	This work
pTGAK	pT carrying GAK fragment (tyrA::GAK)	This work

FP^r: Resistance to p-fluorophenylalanine; Nx^r: Resistance to nalidixic acid; Ap^r: Resistance to ampicillin; Km^r: Resistance to kanamycin; ATCC: American Type Culture Collection. P_BF: Promoter from C.glutamicum; T: Terminator of gene32 in T₄ phage. GAK: P_BF-aroG-pheA-T-Km fragment.

Table 2 Relative activities of CM, PD and DS compared with the original strains

Strains	Enzymes
	CM	PD	DS
C. glutamicum	1.0	1.0	1.0
C. glutamicum FP	1.73	2.37	1.93
C. glutamicum KM	1.74	2.30	1.95
C. glutamicum GAK	2.90	3.36	3.00

The enzyme activity of C.glutamicum was used as the standard, i.e. the relative activity of 1. CM: Chorismate mutase; PD: Prephenate dehydratase; DS: 3-deoxy-D-arabinoheptulosonate-7-phosphate synthase.

Table 3 Yields of L-phenylalanine biosynthesis in flask fer-mentation (mean ± SD)

Strains	C.glutamicum	C.glutamicum FP	C.glutamicum KM	C.glutamicum GAK
Yields (g/L)	2.20 ± 0.15	3.12 ± 0.17	3.32 ± 0.12	3.76 ± 0.21
Relative yields	1	1.42	1.51	1.71

The phenylalanine yield of the original stain was used as the standard, i.e. the relative yield of 1.