Biomolecular Engineering Solutions for Renewable Specialty Chemicals. Группа авторов

gene (SDH). This leads to accumulation of succinic acid giving 43 g/l succinic acid from crude glycerol (Gao et al., 2016a). Later acetyl‐CoA hydrolase (Ylach) gene was deleted from Y. lipolytica to inhibit aceitic acid production and giving a yield of 23.8 g/l succinic acid (Yu et al., 2018). Ylach‐deficient strain with overexpression of phosphoenolpyruvate carboxykinase (ScPCK) and endogenous succinyl‐CoA synthase beta subunit (YlSCS2) improved succinic acid titer by 4.3‐fold (Cui et al., 2017). In a recent study, in spite of deleting SDH gene, SDH promoter was truncated with expressing ScPCK from A. succinogenes. It produced 7.8 ± 0.0 g/l succinic acid with a yield of 0.105 g/g glucose (Babaei et al., 2019). S. cerevisiae’s SDH have four subunits SDH1, SDH2, SDH3, and SDH4 (Kubo et al., 2000). For SDH to be inactivated SDH1 and SDH2 is to be disrupted. This leads to increase in succinic acid synthesis by further adding malic transporter from Schizosaccharomyces pombe (Ito et al., 2014).

      1.3.2.4 Fumaric Acid

      Fumaric acid (C₄H₄O₄) is an intermediate of TCA cycle. It is white crystal solid acid with no odour and is used in plastic industry and in manufacture of sizing resins in paper industry. Being nontoxic in nature it is used in corn tortillas, wheat, sour dough, fruit juice, rye breads, refrigerated biscuit dough, nutraceutical drinks, gelatin desserts, pie fillings gelling aids, wine, and as supplement in animal feed (Khan et al., 2017). In 2014, the global market size of fumaric acid was 245.4 kilo tons and expanding at a compound annual growth rate of 6.1% from 2015 to 2022 (Fumaric acid market analysis by application, https://www.grandviewresearch.com/industry‐analysis/fumaric‐acid‐market).

      It was first isolated by a plant Fumaria. After that many microorganisms’ strains such as Rhizopus, Mucor, Cunninghamella, and Circinella are used for the production of fumaric acid, and among them Rhizopus nigricans, Rhiozopus arrhizus, R. oryzae, and Rhiozopus formosa give higher yield (Singh et al., 2017). Yu et al. irradiated A. oryzae by femtosecond laser twice, and the mutant strains were selected for fumaric acid production (Yu et al., 2012). There was an increase in yield of 36.6% in comparison to the wild‐type strain. Expressing exogenous gene phosphoenolpyruvate carboxylase and overexpressing endogenous gene pyruvate carboxylase increase the carbon flux to oxaloacetate and hence fumarate (Zhang et al., 2012). Fumarase overexpression leads to decrease in fumaric acid as fumarase does hydration of fumarate to malate (Zhang and Yang, 2012). Site‐saturation engineering of proline 474 in pyruvate carboxylase from R. oryzae was done to improve fumaric acid production in S. cerevisiae (Xu et al., 2012). Ethanol yield was lower down by deleting fumarate hydratase gene. The generated strain gives a yield of 314.5 ± 3.8 mg/l.

      Similarly, E. coli can also be engineered by overexpressing and deleting the abovementioned genes. The iclR gene encoding a repressor of glyoxylate shunt operon was deleted to direct carbon flux to glyoxylate shunt (Song et al., 2013). All three fumarase genes were also deleted enhancing fumaric acid production to 1.45 g/l. in a strain with all three fumarase genes deleted with the overexpression of phosphoenolpyruvate carboxylase gene gives yield of 3.65 g/l fumaric acid (Li et al., 2014).

      1.3.3 Therapeutic Proteins

      Proteins are the building block of our body and are essential for proper functioning of body. Any defect in a particular protein can lead to a disease such as hemophilia (lack of various blood‐clotting factors), dwarfism (lack of growth hormone), and diabetes (less or no insulin production) (Vajo et al., 2001; Takeda et al., 2010). These diseases caused by the deficiencies of the proteins can be treated by regular administration of them. These are synthesized by the other organisms, isolated, purified, and then given to patients. Proteins synthesis for human use is completely different form the enzymes used for chemical industry as for human’s quality constraints are strict and detailed clinical trials are required. More therapeutic proteins are under clinical trial than in the market.

      Out of the many recombinant therapeutic proteins human growth hormones (hGH) is at the top. Its market was estimated to be $2850.70 million in 2018 and is anticipated to reach $5653.60 million by 2026 observing a CAGR of 8.2% over the estimated period. Two of the most common hGh currently in the market are Accretropin™ and Sermorelin (Reh and Geffner, 2010). Accretropin™ (recombinant human growth hormone (r‐hGH); somatropin) is a protein, which is manufactured by rDT. The production process includes fermentation of E. coli that gives a protein product containing 192 amino acids. Later process includes removal of the N‐terminal amino acid, methionine that yields a product which is physio‐chemically and chemically similar to pituitary‐derived hGH, comprising of 191 amino acids. These are arranged in a single polypeptide chain. In general terms, Accretropin vitalize linear growth in those patients whose body lacks sufficient production of the endogenous growth hormones. Other than that, it also shows substantial effect in process such as tissue growth, metabolism of protein, carbohydrate, and lipids along with minerals. The drug got approved by FDA on 23 January 2018. It is manufactured by Cangene Corporation, Canada.

      Similarly, Sermorelin or growth hormone releasing factor 1‐29 NH₂‐acetate is classified as a member of the growth hormone‐releasing hormone analogue class drug. It is made up of 29 amino acids out of the 44 that originate in its natural state (Walker, 2006). It can notably increase the release of the growth hormone in the body. It also aims to enhance growth hormone serum concentrations and insulin‐like growth factor 1 (IGF‐1). The bodies of those children, which does not produce enough growth hormone, can be provided with Sermorelin to assist with the increase in amount of growth hormone processed by the pituitary gland. Other than that, it is also commonly prescribed for the treatment of adult growth hormone deficiency. It is also getting popular with those doctors who use them partly in their medical weight loss plans. The drug was first developed during early 1980s and was approved by the FDA by 1997 for sale via prescription.

Another fine example is IGF‐1 (Laron, 2001). They can be related to the family of insulin‐related peptides, which comprise of relaxin and various other peptides, which are extracted from lower invertebrates. Being a small peptide and comprising of 70 amino acids and with a molecular weight of 7469 Da, it binds to the insulin receptor with a relatively low affinity. Along with IGF‐2, they were first found in 1957 by Salmon and Daughday. The IGF‐1 plays major role in proliferation and function of just about every cell, tissue, and organ in the body. Their action mechanism conciliates through IGF/IGFR binding, the activation of kinase, and signaling via AKT pathway.

1.4 Photosynthetic Production of Biofuels

      The increase in energy demand globally is affecting environment due to global warming and depletion of nonrenewable energy sources. Conventional fossil fuel sources such as petrol, diesel, natural gas, and coal, which were considered to be the primary sources earlier are getting exhausted due to extensive uses (Kumar et al., 2017). A 37% surge in fuel demand by 2040 has been estimated due to the rapid increase in demand (Joshi et al., 2017). Besides this the petroleum‐based fuel upon burning releases greenhouse gases. This requires a renewable substitute of petroleum‐based fuels. Biofuels are the renewable energy source produced by photosynthetic organisms utilizing Earth’s biggest fuel source, the Sun as the carbon source. Harvesting solar energy via photosynthesis is one of nature’s noteworthy achievements that could also be a solution for the future worldwide economy. Earlier biofuels were produced from plants known as the first generation of biofuels. They are mainly generated from wheat, barley, corn, oilseed, sunflower, etc. the plants, which are rich in carbohydrates and oils. Biofuels produced by them compete with agriculture croplands leading to crisis in food production for human beings (Surriya et al., 2015). Additionally, harvesting of plants take full season and then processing the complex sugars from plants to simpler sugars that can be used by microorganisms limits the production of biofuel (Voloshin et al., 2019). Second generation of biofuels solves the problem of utilizing croplands as they are based on agricultural waste, forest dregs, waste wood residues, and organic waste materials. Biofuels from algae are considered to be third‐generation biofuels. Microalgae