is chemically converted into Ambrox (Cheetham 1993). A fermentation process was
performed at Nippon Mining Co. for the production of a dicarboxylic acid which is
alpha-omega-alkanoic acid for macrocyclic musk molecules. Candida tropicalis has
been mutated to give high yields of α-ω-alkanoic acid from C10-C18 alkanes, 120 g
of product/L is produced on 20 m
3 scale (Cheetham 1999).
7.3.2.9 Synthetic Biology
Synthetic biology has become a new tool in the synthesis of aromatic molecules
because of the declining cost of DNA synthesis, rapid advances in bioinformatics
tools and expanding omics databases. Now in a heterologous microbial host it has
become possible to resolve single and multienzyme gaps in a heterologous microbial
host. Synthetic biology plays a revolutionary role in the creation of Saccharomyces
cerevisiae and E. coli as an aroma factory. It allows for the production of a
completely new set of microbial-derived flavours. In S. cerevisiae genome the
repetitive sequences were removed, LoxPsym sequences were introduced at the 5’
ends of all the genes in the yeast genome which are considered individually
non-essential. These are the sites which allow inducible homologous recombination
downstream of all non-essential genes which is mediated by the action of the sitespecific Cre recombinase. Rapid gene deletion, duplication or inversion is promoted
at these LoxPsym sites, the process is known as SCRaMbLE (Synthetic Chromosome Rearrangement and Modification by LoxPsym-mediated Evolution), which
allows for the rapid synthetic rearrangement and evolution of fermentation. Thus, a
large library of genomically divergent yeasts has also been created (Wyk et al. 2018).
7.3.2.10 Metabolic Engineering
The most recent approach in metabolic engineering involves tools and strategies
which employ engineering the microbial cells to follow a biosynthetic module. The
biochemical pathways involved in the production of these compounds have to be
understood. This further requires the identification of the genes and enzymes
involved in the synthesis of volatile compounds. The concept of metabolic engineering of aroma has also been previously applied to a variety of food items such as
fruits, vegetables and herbs (tomato, potato, etc.), milk products and alcoholic
beverages (Dudareva and Pichersky 2008).
Rational metabolic engineering and inverse metabolic engineering are the two
approaches used for production of bioflavours. Defined genetic manipulations are
made in genome to carry out a metabolic pathway of interest. Inverse metabolic
engineering strategy is carried; genes are knocked-out/knocked-in to get a desired
aromatic phenotype (Turanlı-Yıldız et al. 2017). The aromatic chemicals which are
derived from microorganisms are usually shikimate (SHK) and aromatic amino acids
like L-phenylalanine (L-PHE), L-tyrosine (L-TYR) and L-tryptophan (L-TRP).
These aromatic compounds can be categorized into intermediates and derivatives
of the shikimate (SHK) pathway and aromatic amino acids, e.g. L-phenylamine
(PHE), L-tyrosine (TYR), L-tryptophan (TRP) and their derivatives (Huccetogullari
et al. 2019).
7 Biotechnological Interventions for Production of Flavour and Fragrance. . .
147
performed at Nippon Mining Co. for the production of a dicarboxylic acid which is
alpha-omega-alkanoic acid for macrocyclic musk molecules. Candida tropicalis has
been mutated to give high yields of α-ω-alkanoic acid from C10-C18 alkanes, 120 g
of product/L is produced on 20 m
3 scale (Cheetham 1999).
7.3.2.9 Synthetic Biology
Synthetic biology has become a new tool in the synthesis of aromatic molecules
because of the declining cost of DNA synthesis, rapid advances in bioinformatics
tools and expanding omics databases. Now in a heterologous microbial host it has
become possible to resolve single and multienzyme gaps in a heterologous microbial
host. Synthetic biology plays a revolutionary role in the creation of Saccharomyces
cerevisiae and E. coli as an aroma factory. It allows for the production of a
completely new set of microbial-derived flavours. In S. cerevisiae genome the
repetitive sequences were removed, LoxPsym sequences were introduced at the 5’
ends of all the genes in the yeast genome which are considered individually
non-essential. These are the sites which allow inducible homologous recombination
downstream of all non-essential genes which is mediated by the action of the sitespecific Cre recombinase. Rapid gene deletion, duplication or inversion is promoted
at these LoxPsym sites, the process is known as SCRaMbLE (Synthetic Chromosome Rearrangement and Modification by LoxPsym-mediated Evolution), which
allows for the rapid synthetic rearrangement and evolution of fermentation. Thus, a
large library of genomically divergent yeasts has also been created (Wyk et al. 2018).
7.3.2.10 Metabolic Engineering
The most recent approach in metabolic engineering involves tools and strategies
which employ engineering the microbial cells to follow a biosynthetic module. The
biochemical pathways involved in the production of these compounds have to be
understood. This further requires the identification of the genes and enzymes
involved in the synthesis of volatile compounds. The concept of metabolic engineering of aroma has also been previously applied to a variety of food items such as
fruits, vegetables and herbs (tomato, potato, etc.), milk products and alcoholic
beverages (Dudareva and Pichersky 2008).
Rational metabolic engineering and inverse metabolic engineering are the two
approaches used for production of bioflavours. Defined genetic manipulations are
made in genome to carry out a metabolic pathway of interest. Inverse metabolic
engineering strategy is carried; genes are knocked-out/knocked-in to get a desired
aromatic phenotype (Turanlı-Yıldız et al. 2017). The aromatic chemicals which are
derived from microorganisms are usually shikimate (SHK) and aromatic amino acids
like L-phenylalanine (L-PHE), L-tyrosine (L-TYR) and L-tryptophan (L-TRP).
These aromatic compounds can be categorized into intermediates and derivatives
of the shikimate (SHK) pathway and aromatic amino acids, e.g. L-phenylamine
(PHE), L-tyrosine (TYR), L-tryptophan (TRP) and their derivatives (Huccetogullari
et al. 2019).
7 Biotechnological Interventions for Production of Flavour and Fragrance. . .
147