This Small Business Innovation Phase I Research Grant proposal aims to develop aUV temperature and pH-stable edible blue C-phycocyanin (CPC) pigment derived from bio- engineered cyanobacterium Arthrospira platensis (a.k.a. Spirulina) for the industrial ingredientmarketplace. Specifically this pigment will be initially used in the food industry for dyeingbeverages confectionary products and baked goods but has the future potential to be appliedto cosmetic textile ink paint medical therapeutic and diagnostic industries.Objectives: 1) Use metabolic engineering to create a strain of Arthrospira platensis thatproduces a stable blue CPC pigment.2) Demonstrate proof of concept by expressing purifying and testing the stability of the A.platensis pigment at high temperature (>47C) low pH < 3 and prolonged UV light exposure.
Keywords: Arthrospira platensis metabolic engineering CRISPR extremophile cyanobacteriaindustrial biotechnology plant protein food ingredients colorant stabilityIntellectual Merit: There is limited information available in the scientific literature about thegenetic engineering of Arthrospira platensis (spirulina). The organism is highly-resistance toconventional metabolic engineering techniques with high endonuclease activity thereforenovel methods of genetic engineering need to be attempted to create a platform forproduction of industrial biotechnological ingredients primarily ingredients for the foodindustry. Particularly of interest is the pigment C-phycocyanin (CPC) which is a rare biobasedblue pigment. In order to increase the stability of this pigment so that it can be used inconventional food production processes novel CRISPR-based engineering techniques will beapplied to enhance the resistance to high temperature low pH and increased UV exposure. Thesame techniques used to develop CPC mutants can be used to expand the stability of othermolecules overall color palette of chromoproteins and expand the usage of biobasedreplacements for petrochemical-derived dyes for any current colors.Broader Impacts: As a company Spira is focused on supplying CPC from A. platensis as naturalblue color additive that since its FDA approval in August 2013 has been used in the food (i.e.confectionary dairy) and beverage (i.e. health drinks) industries. Other commercial applicationsthat benefit from a stable acidothermo CPC are worth noting: Phycobilisomes containing CPCare also finding use as naturally engineered solar light concentrators to "sensitize" metal andsemiconductor surfaces for organic photoelectron-chemical (PEC) photovoltaic cells. CPC is alsoused in cosmetics and textiles as a colorant. Small quantities of reagent-grade CPC are used asfluorescent tags in diagnostic cell labeling. As a potent anti-oxidant CPC can non-enzymaticallyneutralize and scavenge free-radicals of reactive oxygen species (ROS) causing oxidative stress.CPC anti-microbial activity for the in vitro growth of fungi viruses and bacteria enables it tocontribute to natural (i.e.non-artificial) food preservation and improvement of food shelf life.CPC is also an immune stimulant and anti-inflammatory agent capable of modulatingmacrophage function. CPC has been shown to have anti-cancer properties as well via itsinhibition of tumor cell proliferation and/or inducement of tumor cell apoptosis. As an anti- obesity agent CPC inhibits pancreatic lipase activity and has been shown to decrease serumcholesterol. As a neuro-protector CPC interacts with -synuclein to reduce formation of fibrilformation in neurodegenerative diseases. In summary CPC is used a colorant in food textileand cosmetic industries and has potential to be more extensively used as a therapeutic via itsantioxidant anti-inflammatory anti-microbial anti-obesity antihepatoxicity and anti-cancerproperties among others.
