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Bioremediation for Sustainable Environmental Cleanup

a high copy number and a broad host range that carried efe from P. syringae. The recombinant’s

maximum ethylene production rate was 26 times greater than that of the parental P. syringae. The

oxygen and carbon sources could be improved to increase yield even further (Wang et al. 2010).

8.4.2 Medium Chain Length PHA

A bioadaptive functional polymer is medium-chain-length polyhydroxyalkanoate (mcl-PHA). It

is found in fluorescent Pseudomonas species that share the same rRNA homology as P. putida,

P. oleovorans and P. aeruginosa (Kim et al. 2006). Mcl biosynthesis is controlled by the phaC1

and phaC2 genes (Rehm and Steinbüchel 1999, Steinbüchel and Hein 2001). The phaC1 gene was

overexpressed, resulting in a 2.86-fold increase in mcl biosynthesis. It produced larger, higher

molecular weight and less crystalline mcl-PHA granules, which aided in the purification process.

Overexpression of phaC2 had no effect.

The engineered P. putida KCTC1639 overexpressing the phaC1 gene encoding PHA synthase

I was grown in a pH-stat fed-batch system, resulting in increased mcl-PHA concentration and

content of 8.91 g L^–1 and 70.5%, respectively (Kim et al. 2006). Under nitrogen-limited conditions,

P. menodocina NK-01 produces mcl-PHA and alginate oligosaccharide from glucose. This strain’s

production was improved by using a counter-selective maker (Wang et al. 2015). Ferulic acid is

a phenolic lignocellulose component. In P. putida, ferulic acid is converted into mcl-PHA via a

non-beta-oxidative pathway. This conversion rate is extremely low. To overcome the limitations

of conventional CRISPR-Cas9n—Red genome editing, a type II CRISPR-Cas9n—Red genome

editing strategy was used (in bacteria with no or less NHEJ system, the cell viability goes down,

mutation efficiencies of different targets are highly variable). The expression of nine genes was

altered in order to improve the ferulic acid to PHA conversion (Volke et al. 2020).

8.4.3 Short Chain Length PHA

Thermoplastic polyhydroxyalkanoate (PHA) with a short chain length (scl) can be utilized as an

alternative to plastics derived from petroleum (Gahlawat and Soni 2017). It has application in the

field of medical as well as packaging (Kamravamanesh et al. 2018). PHAs that have shorter chain

lengths are made up of 3-hydroxybutylate, also known as 3HB (Lemoigne 1926). The melting

point of the polymer can be lowered by adding poly(3-hydroxybutyrate-co-3-hydroxyvalate) and

4-hydroxyvalate to scl-PHA. Additionally, the polymer’s toughness can be increased, making it a

very valuable material (Gahlawat and Soni 2017, Yang et al. 2012, Schmack et al. 1998). Levulinic

acid, which is used in the production of 3 Hydroxyvalate and 4 Hydroxyvalate monomers, is

not made from petroleum-based precursors; rather, it is produced from sugars generated from

biomass, specifically glucose and xylose (Koller et al. 2017, Novackova et al. 2019). In P. putida,

the breakdown of LA is controlled by a seven-gene operon called lva (lvaABCDEFG). P. putida

strain SP01 was developed by removing the native PhaC1, PhaC2 and PhaZ genes from P. putida

strain EM42. This was done with the intention of inhibiting the synthesis of mcl-PHA and halting

the degradation of scl-PHA. It was not necessary to apply an external inducer because the IvaA

promoter is already stimulated by LA. The PHA synthase (PhaEC) of Thiococcus pfennigii, which

has a broad substrate specificity, was only expressed in the LA catabolic pathway of P. putida in

order to create scl-PHA from two intermediates (4HV-CoA and 3HV-CoA). This route is responsible

for the degradation of fatty acids (Cha et al. 2020). P. putida has been genetically modified to

produce a greater amount of PHA. The gcd gene, which encodes dehydrogenase, has been removed

in order to divert flux towards products that are more desirable (Tiso et al. 2016).

8.4.4 2-methylcitric Acid and others

2-methylcitric acid (2-MC) has the potential to inhibit fast cancer cells. It can also be used in synthetic

materials or pharmaceutical products as an emulsifier or polymer plasticizer (Ewering et al. 2006).