Three Masters of Science graduates were overheard talking about the impossible burger— if you haven’t heard about it, it is a vegan-meat substitute produced by Impossible Foods Inc. which palatably mimics an actual ground-beef burger. The one was telling the others, that the burger was not only delicious and filing, but that it smelled and tasted like real beef. When questioned as to what gave it the beef-like sensation, she answered heme, but was unsure of the source of the heme. This was puzzling because most people are familiar with heme being a component of animal blood, and the idea that heme could be vegan or synthetically produced from amino acids in a lab seemed dubious. 

However, as it turns out, heme is ubiquitous to many living organisms, and can be found in  bacteria, fungi, protozoa, higher plants, and animals. In fact, the heme used by Impossible Foods Inc. is derived from a plant heme known as leghemoglobin, that is sourced from soybeans. The leghemoglobin is not directly extracted from the soybean, as extracting leghemoglobin from plant roots is tedious and not cost effective due to low yield. Therefore, to increase productivity, the leghemoglobin is synthesized through genetic engineering.

Leghemoglobin & Heme

Hemoglobin, colloquially referred to as heme, is an iron binding protein that is involved in oxygen transport in living organisms.  In vertebrae, heme is produced in mammals, and dietarily acquired through meat, poultry, and fish. In plants there are two types of heme: symbiotic and non-symbiotic. Symbiotic  plant hemoglobin is produced in the root nodules of nitrogen-fixing plants symbiotically infected with rhizobium bacteria — hence the name. Symbiotic plant heme can be found in legumes and non-legumes and responsible for oxygen diffusion. Non-symbiotic heme is related to symbiotic heme, but more widespread, being found in rice and barley, in addition to soybean. Unlike symbiotic heme, non- symbiotic heme production is associated with metabolically activity or stress, and is thought to be capable of reversible oxygen binding.

Leghemoglobin is the heme found in the root nodules of leguminous plants, such as soybeans (Glycine max). Leghemoglobin is a soluble protein that is structurally and functionally similar to mammalian myoglobins and hemoglobins; having a high affinity for binding oxygen. There are several isoproteins (similar forms) of Leghemoglobin and all are responsible for the transport, respiration, or metabolism of oxygen in plant tissue.

Laboratory synthesized Leghemoglobin

The heme used in the production of vegan-beef is a recombinant protein genetically engineered through inserting the leghemoglobin c2 gene from the soybean plant  into a yeast host, pichia pastoris. The genetically modified new strain of p. pastoris is capable of expressing and secreting leghemoglobin.  Recombinant leghemoglobin, of  greater than 65% purity, can be grown in large commercial batches allowing for the production of large amounts of the protein. However, some of the proteins from the yeast host are also expressed in the recombinant leghemoglobin, and this has caused some concern over allergenicity and safety. Genetically modified food substances may pose a risk of eliciting allergy through cross-reactivity to similar proteins or through the insertion of a gene alter the amount of  allergen naturally produced by the host species. In other words, someone allergic to soybean, may have an allergy to the recombinant leghemoglobin because its has a gene sourced from soybean, or someone who is allergic to pichia yeast may be allergic to the recombinant leghemoglobin because it contains some of the proteins from the yeast host.

 Safety of  Yeast Host

Yeast are capable of correctly expressing biologically active proteins in a process known as recombinant protein production.Various yeast strains are incorporated in recombinant protein technology to produce resources that are hard to procure from their natural source, and have employed by the pharmaceutical, cosmetic, food, and chemical industries since the early 1980’s.

Pichia pastoris is an obligate aerobic yeast, meaning it requires oxygen for respiration and metabolic survival. It is in the family Saccharomycetaceae; the same family as the yeasts saccharomyces cerevisiae and brettanomyces,  which are commonly used in food and beverage production ( bread and beer). However, unlike S. cerevisiae, p. pastoris is crabtree negative yeast, meaning it does not ferment carbohydrates or produce ethanol, but can use methanol as its carbon source. As a result, p. pastoris can produce a higher yield of recombinant leghemoglobin, and is therefore favored in various industries as the host for the production of various recombinant proteins.

Since it is similar to other yeasts historically used in the food industry and readily digested by pepsin, a component of gastric fluid, the likelihood of p. pistoria proteins causing allergy (at the concentrations present in recombinant leghemoglobin) is low.

Overall Safety of Recombinant Leghemoglobin

Likewise, recombinant leghemoglobin is readily digested in the stomach, has comparable bioavailability as  leghemoglobin and mammalian heme, and is not likely to pose a significant risk of allergenicity. Leghemoglobin itself, is routinely consumed through eating of various vegetables, and though derived from soybean, the leghemoglobin found in soybean plants is present in the roots and not the bean. Therefore,  allergy to to the bean or legume of the soy plant does not necessarily confer allergy to the heme protein symbiotically produced in its roots. Furthermore recombinant proteins are used in the production of insulin, collagen, human serum albumin, hepatitis B vaccines, and much more, without any significant risk of harm of humans. Overall, adding leghemoglobin to vegan-meat not only imparts, beef-like taste and aroma, but contributes a good source of bioavailable iron, which vegans tend to be deficient in.

References

1. Liu, S. (2019). A better burger. Chemical Engineering Progress, 115(1), 24.
2. Arredondo-Peter, R., Hargrove, M. S., Moran, J. F., Sarath, G., & Klucas, R. V. (1998). Plant hemoglobins. Plant Physiology, 118(4), 1121.
3. Vieira Gomes, A. M., Talita, S. C., Lucas, S. C., Frederico Mendonça Bahia, & Nádia, S. P. (2018). Comparison of yeasts as hosts for recombinant protein production. Microorganisms, 6(2)