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Table 1 Some representative examples of limonene production in engineered microbial hosts

From: Engineering the oleaginous yeast Yarrowia lipolytica to produce limonene from waste cooking oil

Host

Product

Titer

Yield

Productivity

Strategy

References

E. coli

l-Limonene

430 mg/L

–

5.97 mg/L/h

A plasmid with a truncated GPP synthase and limonene synthase gene was co-expressed with a plasmid increasing the MK and PMK levels (MBI-f operon) and replacing the HMGS and HMGR from S. cerevisiae (MevT operon) with those from S. aureus (MTSA operon) using different IPTG levels

[28]

l-Limonene

2700 mg/Lorg

–

60 mg/Lorg/h

(1) Truncated LS gene and truncated GPPS2 gene with all the genes of the MVA pathway, that convert endogenous acetyl-CoA into IPP and DMAPP from S. cerevisiae was co-expressed. (2) A two-liquid phase fed-batch fermentation with glycerol as carbon source were carried out in 3.1 L stirred-tank reactors

[35]

l-Limonene

75.12 mg/L

–

3.13 mg/L/h

(1) Juice from sugar beets was utilized as a novel feedstock. (2) A synthetic DXP operon (dxs-ipphp-gpps) encoding three rate-limiting enzymes from the MEP pathway was co-expressed with limonene synthases gene

[75]

S. cerevisiae

d-Limonene and l-limonene

0.12 mg/L d-limonene and 0.49 mg/L l-limonene

–

1.67 µg/L/h d-limonene and 6.8 µg/L/h l–limonene

The truncated limonene synthases with Cytochrome P450 reductase (CPR1) (Gal1 promoter) were co-expressed, and a setup was built which allowed trapping of exhaust volatiles during growth

[33]

Y. lipolytica

d-Limonene and l-limonene

11.705 mg/L d-limonene and 11.088 mg/L l-limonene

0.443 mg/g DCW for d-limonene and 0.385 mg/g DCW for l-limonene

0.0542 mg/L/h d-limonene and 0.0513 mg/L/h l-limonene

(1) The heterologous d-limonene synthase from Citrus limon and l-limonene synthase from Mentha spicata with ten homologous genes involved in the MVA pathway were overexpressed, respectively. (2) Fermentation conditions of engineered strains were optimized. (3) Fed-batch fermentation of engineered strains was used. (4) WCO was used as carbon resource

This work

d-Limonene

23.56 mg/L

1.36 mg/g DCW

0.33 mg/L/h

The truncated genes tLS and tNDPS1 with HMG1 and ERG12 were co-expressed

[72]

Anabaena sp. PCC 7120

l-Limonene

–

–

172.7 µg/L/48 h

A synthetic DXP operon (dxs-ipphp-gpps) encoding three rate-limiting enzymes from the MEP pathway was co-expressed with limonene synthases gene

[76]

Synechococcus sp. PCC 7002

l-Limonene

4 mg/L

–

0.042 mg/L/h

The (−)-4S-limonene synthase gene was expressed

[30]

Synechocystis sp. PCC 6803

d-Limonene

0.564 mg/L

–

2.33 µg/L/h

Introducing an additional copy of three genes (dxs, crtE and ipi) under the control of a strong trc promoter

[77]