Synthesis of jojoba-like wax esters in metabolically engineered strains of Saccharomyces cerevisiae
Doktorsavhandling, 2018
Yeast has a long-standing tradition in human history as a production organism of choice. Besides being used for the production of fermented products like bread or beer, it has also been extensively explored for the production of proteins and chemicals. In the past, yeast research in this respect was often focused on the production of biofuels as an alternative to fossil fuels to enable the independence of crude oil. However, crude oil also functions as a source for a wide range of chemicals.
Some of these chemicals can be substituted by alternatives derived from plant oils, like jojoba oil. Jojoba seeds contains approximately 50% (w/w) oil, which consists mostly (up to 97%) of wax esters (WEs). Minor parts include phytosterols, triacylglycerols (TAGs) and fatty alcohols (FOHs). This makes jojoba exceptional, since plants usually accumulate TAGs as storage compounds. Jojoba oil can among other applications be used in cosmetic and personal care products as well as lubricants. Currently, around 4,000 tons/year of jojoba oil is produced, with an estimated demand of up to 200,000 tons/year. Because of this, oil produced from the jojoba plant will not be enough to meet the demand in the future, even if huge land areas in various parts of the world are planted.
Therefore, jojoba oil production in modified microorganisms represents a very promising approach. In this thesis, the yeast Saccharomyces cerevisiae was explored as a production organism for jojoba-like WEs. Jojoba-like WEs are naturally derived from the fatty acid (FA) metabolism of the plant, more specifically from very long-chain monounsaturated fatty acids (VLCMUFAs) with a carbon chain length of 20/22 (C20/C22). These VLCMUFAs can be converted to FOHs by the action of fatty acyl reductases (FARs). WEs are synthesized by wax synthases (WSs), which esterify an activated FA (fatty acyl-CoA) with a FOH molecule.
In this thesis, the synthesis of jojoba-like WEs in S. cerevisiae was established by making use of various enzymes derived from bacterial and plant sources as well as tuning S. cerevisiae FA metabolism towards the increased synthesis of VLCMUFAs. In this way a S. cerevisiae strain was created that produces 14.38 +/- 1.76 mg WEs/g CDW. Of these WEs, 39.2 mol% are jojoba-like diunsaturated C38:2-WEs to C42:2-WEs, with the most abundant ones being C42:2-WEs (18.3 mol% of all WE species). These are also the most abundant WEs in natural jojoba oil (46.8 mol% of all WE species).
Some of these chemicals can be substituted by alternatives derived from plant oils, like jojoba oil. Jojoba seeds contains approximately 50% (w/w) oil, which consists mostly (up to 97%) of wax esters (WEs). Minor parts include phytosterols, triacylglycerols (TAGs) and fatty alcohols (FOHs). This makes jojoba exceptional, since plants usually accumulate TAGs as storage compounds. Jojoba oil can among other applications be used in cosmetic and personal care products as well as lubricants. Currently, around 4,000 tons/year of jojoba oil is produced, with an estimated demand of up to 200,000 tons/year. Because of this, oil produced from the jojoba plant will not be enough to meet the demand in the future, even if huge land areas in various parts of the world are planted.
Therefore, jojoba oil production in modified microorganisms represents a very promising approach. In this thesis, the yeast Saccharomyces cerevisiae was explored as a production organism for jojoba-like WEs. Jojoba-like WEs are naturally derived from the fatty acid (FA) metabolism of the plant, more specifically from very long-chain monounsaturated fatty acids (VLCMUFAs) with a carbon chain length of 20/22 (C20/C22). These VLCMUFAs can be converted to FOHs by the action of fatty acyl reductases (FARs). WEs are synthesized by wax synthases (WSs), which esterify an activated FA (fatty acyl-CoA) with a FOH molecule.
In this thesis, the synthesis of jojoba-like WEs in S. cerevisiae was established by making use of various enzymes derived from bacterial and plant sources as well as tuning S. cerevisiae FA metabolism towards the increased synthesis of VLCMUFAs. In this way a S. cerevisiae strain was created that produces 14.38 +/- 1.76 mg WEs/g CDW. Of these WEs, 39.2 mol% are jojoba-like diunsaturated C38:2-WEs to C42:2-WEs, with the most abundant ones being C42:2-WEs (18.3 mol% of all WE species). These are also the most abundant WEs in natural jojoba oil (46.8 mol% of all WE species).
Saccharomyces cerevisiae
very long-chain monounsaturated fatty alcohols
very long-chain monounsaturated fatty acids
jojoba-like wax esters
KC-salen, Kemigården 4, Chalmers
Opponent: Prof. Paola Branduardi, Department of Biotechnology and Biosciences, University of Milano-Bicocca, Italy
Författare
Leonie Wenning
Systembiologi
Metabolic engineering of Saccharomyces cerevisiae for production of very long chain fatty acid-derived chemicals
Nature Communications,;Vol. 8(2017)p. Article number: 15587-
Artikel i vetenskaplig tidskrift
Establishing very long-chain fatty alcohol and wax ester biosynthesis in Saccharomyces cerevisiae
Biotechnology and Bioengineering,;Vol. 114(2017)p. 1025-1035
Artikel i vetenskaplig tidskrift
Production of jojoba seed oil in yeast
Jojoba oil, a natural oil derived from the seeds of the jojoba shrub (Simmondsia chinensis), can be used as a sustainable replacement for liquid paraffin, paraffinum liquidum, a highly refined form of mineral oil. Mineral oil is an inexpensive petrochemical byproduct made from petroleum and is commonly used in cosmetics and medicine. Its advantages are that it is colorless, odorless and that is has favorable water binding properties which make it a popular moisturizing ingredient. On the other hand, its disadvantage is that it forms an impenetrable film on the skin which can trap toxins and hinders normal skin respiration. Therefore, mineral oil can interfere with the natural moisturizing mechanism of the body. In contrast to that, natural moisturizers strengthen the skin’s natural lipid moisture barrier and protect it from the environment. When searching for alternatives to mineral oil for use in cosmetics and medicine, it is important to consider the natural composition of human sebum, a lipid-rich secretion produced by sebaceous glands to protect the skin. Human sebum is composed to 25% of wax monoesters, 41% triacylglycerols, 16% free fatty acids and 12% squalene. Based on this knowledge it is favorable to make use of natural sources that are rich in these constituents, e.g. the jojoba shrub, candelilla shrub or carnauba palm.
Jojoba is a perennial, woody shrub which is native to the semiarid regions of Southern Arizona, Southern California and Northwestern Mexico. Seeds of the jojoba plant contain approximately 50% (w/w) oil, which consists mostly (up to 97%) of wax monoesters and only to a minor extent of phytosterols, triacylglycerols and fatty alcohols. The current production of jojoba seed oil is ~ 4,000 tons/year with an estimated demand up to 200,000 tons/year. Since it will not be possible to meet this demand only with oil produced from the jojoba plant, jojoba oil production in modified (micro)organisms represents a very promising approach. Therefore, in this thesis the baker’s yeast Saccharomyces cerevisiae was investigated as a jojoba wax ester production platform. This yeast has been chosen, because it has already been extensively studied for the production of proteins and chemicals. The advantages of S. cerevisiae are that it is very efficient in fermenting sugars, shows a fast growth rate and is robust towards harsh industrial fermentation conditions. Another point making it an attractive choice is that it has a “generally recognized as safe” (GRAS) status. Moreover, since it is extensively used in industry, a broad knowledge about its metabolism has been acquired and a range of useful evolutionary and metabolic engineering tools have been developed. Naturally, S. cerevisiae mainly produces ethanol by fermenting sugars. By modifying its metabolism towards increased fatty acid synthesis, the precursors of wax esters, and introducing wax ester synthesizing enzymes from bacteria and plants, a yeast strain capable of producing jojoba-like wax esters was constructed.
Jojoba oil, a natural oil derived from the seeds of the jojoba shrub (Simmondsia chinensis), can be used as a sustainable replacement for liquid paraffin, paraffinum liquidum, a highly refined form of mineral oil. Mineral oil is an inexpensive petrochemical byproduct made from petroleum and is commonly used in cosmetics and medicine. Its advantages are that it is colorless, odorless and that is has favorable water binding properties which make it a popular moisturizing ingredient. On the other hand, its disadvantage is that it forms an impenetrable film on the skin which can trap toxins and hinders normal skin respiration. Therefore, mineral oil can interfere with the natural moisturizing mechanism of the body. In contrast to that, natural moisturizers strengthen the skin’s natural lipid moisture barrier and protect it from the environment. When searching for alternatives to mineral oil for use in cosmetics and medicine, it is important to consider the natural composition of human sebum, a lipid-rich secretion produced by sebaceous glands to protect the skin. Human sebum is composed to 25% of wax monoesters, 41% triacylglycerols, 16% free fatty acids and 12% squalene. Based on this knowledge it is favorable to make use of natural sources that are rich in these constituents, e.g. the jojoba shrub, candelilla shrub or carnauba palm.
Jojoba is a perennial, woody shrub which is native to the semiarid regions of Southern Arizona, Southern California and Northwestern Mexico. Seeds of the jojoba plant contain approximately 50% (w/w) oil, which consists mostly (up to 97%) of wax monoesters and only to a minor extent of phytosterols, triacylglycerols and fatty alcohols. The current production of jojoba seed oil is ~ 4,000 tons/year with an estimated demand up to 200,000 tons/year. Since it will not be possible to meet this demand only with oil produced from the jojoba plant, jojoba oil production in modified (micro)organisms represents a very promising approach. Therefore, in this thesis the baker’s yeast Saccharomyces cerevisiae was investigated as a jojoba wax ester production platform. This yeast has been chosen, because it has already been extensively studied for the production of proteins and chemicals. The advantages of S. cerevisiae are that it is very efficient in fermenting sugars, shows a fast growth rate and is robust towards harsh industrial fermentation conditions. Another point making it an attractive choice is that it has a “generally recognized as safe” (GRAS) status. Moreover, since it is extensively used in industry, a broad knowledge about its metabolism has been acquired and a range of useful evolutionary and metabolic engineering tools have been developed. Naturally, S. cerevisiae mainly produces ethanol by fermenting sugars. By modifying its metabolism towards increased fatty acid synthesis, the precursors of wax esters, and introducing wax ester synthesizing enzymes from bacteria and plants, a yeast strain capable of producing jojoba-like wax esters was constructed.
Yeast Cell Factories: Training Researchers to Apply Modern Post-Genomic Methods In Yeast Biotechnology (YEASTCELL)
Europeiska kommissionen (EU) (EC/FP7/606795), 2013-09-01 -- 2017-08-31.
Drivkrafter
Hållbar utveckling
Ämneskategorier
Industriell bioteknik
Biokemi och molekylärbiologi
Mikrobiologi
Infrastruktur
Chalmers infrastruktur för masspektrometri
Styrkeområden
Livsvetenskaper och teknik (2010-2018)
ISBN
978-91-7597-794-2
Doktorsavhandlingar vid Chalmers tekniska högskola. Ny serie: 4475
Utgivare
Chalmers
KC-salen, Kemigården 4, Chalmers
Opponent: Prof. Paola Branduardi, Department of Biotechnology and Biosciences, University of Milano-Bicocca, Italy