Limits...
Inhibition of Coenzyme Qs Accumulation in Engineered Escherichia coli by High Concentration of Farnesyl Diphosphate.

Samoudi M, Omid Yeganeh N, Shahbani Zahiri H, Shariati P, Hajhosseini R - Avicenna J Med Biotechnol (2015 Jul-Sep)

Bottom Line: Over-expression of ispA under the control of stronger trc promoter, however, led to a severe decrease in CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains, as reflected by reductions from 629±40 to 30±13 and 564±28 to 80±14 μg/g Dried Cell Weight (DCW), respectively.The results showed high level of FPP reduces endogenous CoQ 8 production as well and that CoQs are produced in a complimentary manner, as the increase in production of one decreases the production of the other.The reduction in CoQ 10 production can be a result of Dds inhibition by high FPP concentration.

View Article: PubMed Central - PubMed

Affiliation: Institute of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.

ABSTRACT

Background: Coenzyme Q 10 (CoQ 10 ) is an isoprenoid component used widely in nutraceutical industries. Farnesyl diphosphate synthase (FPPS) is a responsible enzyme for biosynthesis of farnesyl diphosphate (FPP), a key precursor for CoQs production. This research involved investigating the effect of FPPS over-expression on CoQs production in engineered CoQ 10 -producing Escherichia coli (E. coli).

Methods: Two CoQ 10 -producing strains, as referred to E. coli Ba and E. coli Br, were transformed by the encoding gene for FPPS (ispA) under the control of either the trc or P BAD promoters.

Results: Over-expression of ispA under the control of P BAD promoter led to a relative increase in CoQ 10 production only in recombinant E. coli Br although induction by arabinose resulted in partial reduction of CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains. Over-expression of ispA under the control of stronger trc promoter, however, led to a severe decrease in CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains, as reflected by reductions from 629±40 to 30±13 and 564±28 to 80±14 μg/g Dried Cell Weight (DCW), respectively. The results showed high level of FPP reduces endogenous CoQ 8 production as well and that CoQs are produced in a complimentary manner, as the increase in production of one decreases the production of the other.

Conclusion: The reduction in CoQ 10 production can be a result of Dds inhibition by high FPP concentration. Therefore, more effort is needed to verify the role of intermediate metabolite concentration and to optimize production of CoQ 10 .

No MeSH data available.


Related in: MedlinePlus

HPLC chromatogram for CQs identification in E. coli Ba. The recombinant E. coli produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4508334&req=5

Figure 2: HPLC chromatogram for CQs identification in E. coli Ba. The recombinant E. coli produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8.

Mentions: As mentioned in our previous work 24, two type of dds genes, rsdds and atdds, were inserted into the pBr1MCS2 plasmid, and resulted in the formation of plasmids pBrsdds and pBatdds. The recent plasmids were transformed into E. coli DH5α, and designated as E. coli Br and E. coli Ba, respectively. The recombinant strains were grown in 2YTGH medium under the same experimental conditions. Their CoQs content was extracted and determined quantitatively by detection of the corresponding peaks in the HPLC chromatogram. The expression of rsdds or atdds in the transformed E. coli Ba and E. coli Br led to the production of CoQ10together with CoQ8. On average, E. coli Ba had CoQ10content of 629±40 μg/g DCW while E. coli Br had 564±28 μg/g DCW under the described experimental conditions. E. coli naturally has an Ods encoding gene. So, the CoQ8content was also measured in CoQ10-producing strains. Expression of dds led to a significant decrease in CoQ8production by both E. coli Ba (from 990±32 to 360±23 μg/g DCW) and E. coli Br (from 990±32 to 246±24 μg/g DCW), when compared to that in E. coli DH5α 24. An example of HPLC chromatogram for CQs identification in E. coli Ba is shown in figure 2. The recombinant E. coli strain produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8. The CoQ9might be produced as a result of immature release of a fraction of the growing polyprenyl chains from the Dds 17.


Inhibition of Coenzyme Qs Accumulation in Engineered Escherichia coli by High Concentration of Farnesyl Diphosphate.

Samoudi M, Omid Yeganeh N, Shahbani Zahiri H, Shariati P, Hajhosseini R - Avicenna J Med Biotechnol (2015 Jul-Sep)

HPLC chromatogram for CQs identification in E. coli Ba. The recombinant E. coli produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4508334&req=5

Figure 2: HPLC chromatogram for CQs identification in E. coli Ba. The recombinant E. coli produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8.
Mentions: As mentioned in our previous work 24, two type of dds genes, rsdds and atdds, were inserted into the pBr1MCS2 plasmid, and resulted in the formation of plasmids pBrsdds and pBatdds. The recent plasmids were transformed into E. coli DH5α, and designated as E. coli Br and E. coli Ba, respectively. The recombinant strains were grown in 2YTGH medium under the same experimental conditions. Their CoQs content was extracted and determined quantitatively by detection of the corresponding peaks in the HPLC chromatogram. The expression of rsdds or atdds in the transformed E. coli Ba and E. coli Br led to the production of CoQ10together with CoQ8. On average, E. coli Ba had CoQ10content of 629±40 μg/g DCW while E. coli Br had 564±28 μg/g DCW under the described experimental conditions. E. coli naturally has an Ods encoding gene. So, the CoQ8content was also measured in CoQ10-producing strains. Expression of dds led to a significant decrease in CoQ8production by both E. coli Ba (from 990±32 to 360±23 μg/g DCW) and E. coli Br (from 990±32 to 246±24 μg/g DCW), when compared to that in E. coli DH5α 24. An example of HPLC chromatogram for CQs identification in E. coli Ba is shown in figure 2. The recombinant E. coli strain produced CoQ10and slight amounts of CoQ9in addition to naturally occurring CoQ8. The CoQ9might be produced as a result of immature release of a fraction of the growing polyprenyl chains from the Dds 17.

Bottom Line: Over-expression of ispA under the control of stronger trc promoter, however, led to a severe decrease in CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains, as reflected by reductions from 629±40 to 30±13 and 564±28 to 80±14 μg/g Dried Cell Weight (DCW), respectively.The results showed high level of FPP reduces endogenous CoQ 8 production as well and that CoQs are produced in a complimentary manner, as the increase in production of one decreases the production of the other.The reduction in CoQ 10 production can be a result of Dds inhibition by high FPP concentration.

View Article: PubMed Central - PubMed

Affiliation: Institute of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.

ABSTRACT

Background: Coenzyme Q 10 (CoQ 10 ) is an isoprenoid component used widely in nutraceutical industries. Farnesyl diphosphate synthase (FPPS) is a responsible enzyme for biosynthesis of farnesyl diphosphate (FPP), a key precursor for CoQs production. This research involved investigating the effect of FPPS over-expression on CoQs production in engineered CoQ 10 -producing Escherichia coli (E. coli).

Methods: Two CoQ 10 -producing strains, as referred to E. coli Ba and E. coli Br, were transformed by the encoding gene for FPPS (ispA) under the control of either the trc or P BAD promoters.

Results: Over-expression of ispA under the control of P BAD promoter led to a relative increase in CoQ 10 production only in recombinant E. coli Br although induction by arabinose resulted in partial reduction of CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains. Over-expression of ispA under the control of stronger trc promoter, however, led to a severe decrease in CoQ 10 production in both recombinant E. coli Ba and E. coli Br strains, as reflected by reductions from 629±40 to 30±13 and 564±28 to 80±14 μg/g Dried Cell Weight (DCW), respectively. The results showed high level of FPP reduces endogenous CoQ 8 production as well and that CoQs are produced in a complimentary manner, as the increase in production of one decreases the production of the other.

Conclusion: The reduction in CoQ 10 production can be a result of Dds inhibition by high FPP concentration. Therefore, more effort is needed to verify the role of intermediate metabolite concentration and to optimize production of CoQ 10 .

No MeSH data available.


Related in: MedlinePlus