Designing Rose-scented Saccharomyces cerevisiae; a Customized Yeast to Enhance the Aroma of Baked Goods

Design Brief

Julia Ashley, Marcella Bangkong, Nancy Jiang, Noah Luch,Vivien Marmerstein, Xinyi Tian, Yumeng Wu,

BioBuilder Club, Western Reserve Academy, Hudson, Ohio, USA

Reviewed on 11 May 2019; Accepted on 17 June 2019; Published on 28 October 2019

With help from the 2019 BioTreks Production Team.

AbstractVideoPDFCopyright

With this synthetic biology project, we aim to transform Saccharomyces cerevisiae, or bread yeast, using the genes responsible for the rose aroma in order to give bread the rose scent. Chefs often use roses in dessert making due to its worldwide recognition as a symbol of love and romance. Different types of rose dishes, including rose crepe cake, rose mousse, and rose pound cake, are popular all around the world. The objective of this paper is to produce yeast that can inherently give bread an elegant rose odor during fermentation as a replacement to the conventional long process used by chefs. We discovered the following genes which account for the production of chemicals responsible for the rose odor: BBa_K1507006, a gene which has been successfully transformed into Escherichia coli, and BBa_K2281005 which has been successfully transformed into yeast. The latter group of researchers reported that the yeast proved to be appropriate for the production of the chemical which accounts for the rose scent. Future data will determine if the rose aroma successfully remains in the yeast and allows it to hold the scent during fermentation and produce rose-scented bread.

Keywords: Citronellol, rose scent, Saccharomyces cerevisiae, bread

Authors are listed in alphabetical order. Beth Pethel, Robert Aguilar, and William Beeson mentored the group. Please direct all correspondence to pethelb@wra.net.

This is an Open Access article, which was copyrighted by the authors and published by BioTreks in 2019. It is distributed under the terms of the Creative Commons Attribution License, which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited.

Background 

Rose is not only famous as a symbol of love and romance but is often used in various dessert making. A few desserts such as Rose crepe cake, rose mousse, and rose pound cake are popular all around the world. However, currently rose-flavored compounds such as rose water, perfume, and oil are very difficult to use for beginner chefs since the flavor will vaporize during baking stage in the oven. Moreover, nowadays customers have a natural tendency against foods with too many additives, and rose flavored bread without additive would appear healthy and natural alternative to all the different additives. The objective of this paper is to produce yeast that can give bread an elegant rose odor during fermentation thus allowing everyone to easily make rose flavored baked goods. Saccharomyces cerevisiae is a strain of yeast most commonly used for bread making. We chose this specific strain because it is commonly used among professional chefs, home cooks, and everyone in between. This transformation of this yeast will make it widely accessible for all.

Systems Level

It is proposed to transform the rose aroma gene (Wang 2017) into yeast. Using BBa_K2281005, we will transform S. cerevisiae, a species of yeast specifically used for bread making. This transformation would test the ability to produce citronellol (a chemical which is found in rose oil and has the rose aroma) when the gene coding for its production is transformed into yeast. Figure 1. depicts the complete pathway of the production of citronellol in a transformed Saccharomyces cerevisiae cell.

Figure 1. The diagram depicts the pathway of glucose to citronellol, the product known for producing the classic rose scent. The process begins with glucose and undergoes three major steps to produce the end product citronellol. Through the Mevalonate (MVA) pathway and DXP reductoisomerase (part of the MEP pathway), which are two essential metabolic pathways, DMAPP and IPP are synthesized and transformed into GPP. The enzyme geraniol synthase (GES) is used to produce geraniol from GPP by removing the two phosphate groups. Finally, with the Old Yellow Enzyme (OYE), an allylic alcohol double bond from Geraniol was reduced, and the final product, citronellol is produced (Yuan et al. 2011).

The LiAc transformation method will be used to transform the aroma gene into the yeast. Competent yeast are prepared and suspended in a LiAc solution with the plasmid DNA to be transformed. We plan to select the transformants using the URA3 complementation method as shown in Figure 2. 

Figure 2. The selection method for transformed S. cerevisiae relies on the URA3 method. The mutated cells used in the process lack a URA3 gene while the plasmid DNA has a copy of URA3 gene as well as the URA3 promoter. S. cerevisiae with a URA3 deletion is transformed with a plasmid containing a URA3 gene and promoter. Transformants are selected by growing the transformed yeast on a specific media without uracil. The genes of interest will be inserted into the plasmid and cloned into S. cerevisiae using this method.

Device Level

The transformed S. cerevisiae will be used in bread-making because it gives the bread a pleasant rose odor by producing citronellol in a safe and environmental-friendly way. Using the selection process described in Figure 2, we would insert the plasmid which accounts for the genes that produce citronellol into the S. cerevisiae cells. This will allow the cells to produce citronellol.

As seen in Figure 1, the process starts with glucose and takes three major steps to produce the end product, Citronellol. Through the Mevalonate pathway (also known as the MVA pathway) and DXP reductoisomerase (part of the MEP pathway), which are two essential metabolic pathways, dimethylallyl pyrophosphate (DMAPP) and isopentenyl pyrophosphate (IPP) are synthesized and converted into geranyl pyrophosphate (GPP).  The enzyme geraniol synthase (GES) is used to produce geraniol from GPP by removing the two phosphate groups. Finally, with the Old Yellow Enzyme (OYE), an allylic alcohol double bond from Geraniol reduces, resulting in the final product, citronellol (Yuan, Chen, Lu, Liu, & Zeng, 2011).

Parts Level

We will be using S. cerevisiae cells and need to either order competent cells or make them in a lab for transformations. Also, a transformation kit for S. cerevisiae would be useful in the process. One of such devices is available from Sigma Aldrich (Yeast Transformation Protocols). On the iGEM Parts Registry, we located the gene that will be used for our transformations and experiment. This is the BBa_K2281005 gene which will be inserted into the yeast in order to produce citronellol to create the pleasant rose scent. 

Safety

Saccharomyces cerevisiae is widely used in the bakery industries, therefore is not harmful to people or the environment.  

Citronellol, the main compound that produces the rose scent, is widely regarded as safe to use. Companies use it for the production of a variety of products, including perfumes and essential oils. Although slight irritability has appeared in certain patch tests, this only occurs when the citronellol is in high concentration and is applied directly to the skin for an extended period of time (HSDB 2015).

Discussions

Our project is the production of an eco-friendly and safe yeast that produces a rose odor when making bread. It will be useful to everyone from master chefs to amateur cooks all around the globe, as it will allow them to easily make rose scented pastries.

Acknowledgements

The Western Reserve Academy BioBuilder Team would like to acknowledge Dr. Pethel, Dr. Beeson, and Dr. Aguilar for their help and support throughout this process. We would also like to thank the MIT BioBuilder team for their help in the process.

This project was accomplished through participation in the BioBuilderClub, an after-school program organized by BioBuilder Educational Foundation.  BioBuilderClub engages high school teams around the world to combine engineering approaches and scientific know-how to design/build/test their own project ideas using synthetic biology. 

References 

HSDB. Citronellol [Internet]. 2015 Dec 23 [cited 2019 August 17]. Available from: http://bit.ly/30bSar8

Hsu, W. [Internet]. Part:BBa_K1507006. 2017, Oct 7 [cited 2019 May 02].  Available from: Registry of Standard Biological Parts website: http://bit.ly/2KPNVeb

Merk. Yeast transformation protocols [Internet]. [cited 2019 May 02] Available from: http://bit.ly/2Miihct 

Tian-Tian Yuan, Qian-Qian Chen, Pei-Ji Zhao, Ying Zeng,corresponding author Xiao-Zhu Liu, Shan Ludoi.  Identification of enzymes responsible for the reduction of geraniol to citronellol. Nat Prod Bioprospect. 2011. 1(3): 108-111 10.1007/s13659-011-0032-6. Available from: https://bit.ly/2OtAJzj

Wang, Y. (2017, October 26). Part:BBa_K2281005. Retrieved May 2, 2019, from Registry of Standard Biological Parts website: http://bit.ly/33CrrpF