Gluten – Not For All Gluttons!

  Shrestha Chowdhury       Aug 04, 2024
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Prelude

Have you heard people regularly complain about how miserable they feel having to avoid mouth-watering foods like pizza, cakes, parathas, or egg rolls? I’m sure you have. If you press them further, you might discover that these wheat-based products contain a substance called gluten. But what exactly is gluten, and why must some people avoid it at all costs?

The answer is a bit complex. Read on to learn more about gluten and how you can give some hope to your gluten-deprived friends, helping them rekindle their taste buds!

Definition, origin, and whereabouts

Gluten, in layman’s terms, is the rubbery mass left in wheat dough after the starch is washed away with water or a brine solution. Technically, it is a complex mixture of hundreds of different proteins found in wheat and similar grains like rye, barley, and oats. These grain storage proteins are specifically located in the endosperm of wheat kernels and play a crucial role in seed germination. The amount of gluten in wheat varies depending on its variety, the time of harvest, and the geographical location where it is grown.

Different parts of a wheat grain. Source

Discovery of glutens

Wheat gluten is one of the earliest scientifically studied proteins. The first study was conducted in 1745 by Jacopo Beccari, a chemistry professor at the University of Bologna. Before the 1970s, when gel electrophoresis was developed, a Connecticut-based plant protein chemist named Dr. T.B. Osborne successfully extracted proteins from various seeds using a series of solvents with varying polarity from 1886 to 1928. His technique, known as Osborne fractionation, is still in use today. According to this method, proteins are divided into four fractions: albumins (soluble in water), globulins (soluble in dilute saline), prolamins (soluble in 60–70% alcohol), and glutenins (insoluble in other solvents but extractable in alkali). Dr. Osborne coined the term ‘prolamins’ for a specific category of proteins due to their high proline content. He also identified various sub-classes of prolamins based on their cereal sources, including gliadin in wheat, hordein in barley, secalin in rye, and zein in maize.

Jacopo Beccari (left, source) and T.B Osborne (right, source).

Components of gluten: gliadins and glutenins

In popular terms, gliadin and glutenin primarily constitute gluten and are collectively known as prolamins. But what produces them inside a grain? The genes responsible for synthesizing gluten proteins in developing wheat grains are as follows: the Gli-1 and Gli-2 loci, which code for gliadin proteins, and the Glu-1 and Glu-3 loci, which code for glutenin polypeptides. To understand the development and synthesis of glutenin polypeptides, it’s essential to delve into the chemistry behind their formation.

At the molecular level, gliadins are monomeric proteins with molecular weights (MW) ranging from approximately 28,000 to 55,000, while glutenins are polymers, specifically of high molecular weight subunits, with weights ranging from around 67,000 to over 88,000.

Gliadins are broadly divided into three types: α\alpha/β−\beta-, γ−\gamma-, and ω−\omega-gliadins. The distribution of these types varies according to the wheat genotype and growing conditions, such as climate, soil, and fertilization. Among them, α\alpha/β−\beta- and γ−\gamma-gliadins are the major components compared to ω-gliadins.

Gliadin C29H41N7O9C_{29}H_{41}N_{7}O_{9} (source).

Upon closer inspection, gliadin proteins consist almost entirely of repetitive sequences rich in glutamine and proline (e.g., PQQPFPQQ). α\alpha/β−\beta- and γ−\gamma-gliadins have similar molecular weights and contain significantly lower amounts of glutamine and proline compared to ω-gliadins. On the other hand, glutenins are composed of both high molecular weight polymers and low molecular weight subunit polymers that are truncated from their higher counterparts. The low molecular weight glutenin subunits (LMW-GS) make up about 20% of the total gluten proteins. LMW-GS are similar to α\alpha/β−\beta- and γ−\gamma-gliadins in terms of molecular weight and amino acid composition.

As HMW-GS does not occur in flour and dough as monomers, it is generally assumed that they form interchain disulphide bonds. Fun fact, the largest polymers termed ‘glutenin macropolymer (GMP) make the greatest contribution to dough properties and their amount in wheat flour (E20–40 mg/g) is strongly correlated with dough strength and loaf volume.

Polymer chemistry in glutens

The study of gluten polymers is particularly relevant to bread preparation, a staple food across many cultures worldwide. Gluten proteins are highly hydrophobic and prefer to bond with lipids rather than water. However, in the context of dough, hydrated gliadins are much less elastic and cohesive compared to glutenins; they primarily contribute to viscosity and extensibility. In contrast, hydrated glutenins are both cohesive and elastic, providing dough with strength and elasticity. In simple terms, gluten functions as a bi-component adhesive, with gliadins acting as a solvent for the glutenins.

Formation of gluten upon mixing (source).

Glutenin subunits form large three-dimensional networks through hydrogen bonds and inter-chain disulfide bonds. These bonds interact with gliadins and other glutenin networks. During bread processing, the state of large glutenin polymers is influenced by three competitive redox reactions. First, the oxidation of free SH groups (thiol) in the peptide chains supports polymerization. Second, the presence of ‘terminators’ that halt polymerization. Finally, SH/SS interchange reactions between glutenins and thiol compounds, such as glutathione, can lead to depolymerization. The crucial role of disulfide bonds is highlighted by the effects of various additives: reducing agents weaken the dough, while thiol-blocking or oxidizing agents strengthen it.

On the other hand, oxygen is crucial for the formation of large glutenin polymers during dough mixing. Oxidizing agents such as potassium bromate, potassium iodate, and L-ascorbic acid produce effects similar to atmospheric oxygen. The baking process further alters the glutenin structure and functionality. For example, extractability in urea or SDS is significantly reduced, and most cysteine-containing α−\alpha-, β−\beta-, and γ−\gamma-gliadins become covalently bound to glutenin polymers after baking. Disulfide interchange reactions between gliadins and glutenins are also believed to contribute to these heat-induced changes.

In summary, gluten proteins are among the most complex protein networks in nature, due to their numerous components, varying sizes, and the variability introduced by genotype, growing conditions, and technological processes. They play a crucial role in determining the unique rheological properties of dough and the baking quality of wheat. Despite these advantages, the initial question remains unanswered: why are some people gluten intolerant? Which component of gluten proteins triggers adverse reactions in some individuals? We will explore this in the next part.

Harmful Effects of Gluten

It is indeed unfortunate that some people are unable to consume gluten-based products. Gluten contains certain toxic peptides that can lead to various health disorders, including celiac disease, wheat allergy, gluten sensitivity, gluten ataxia, and dermatitis herpetiformis. Some studies also suggest that gluten may exacerbate conditions such as schizophrenia, mood disorders, and psoriasis.

Flowchart showing various health disorders related to gluten consumption. Source

Among these conditions, celiac disease is particularly noteworthy. It is a lifelong autoimmune disorder where the ingestion of gluten causes severe inflammation of the intestinal walls, leading to the loss of villi—structures essential for nutrient absorption. This condition is commonly referred to as ‘leaky gut syndrome.’ As a result, affected individuals produce abnormal amounts of antibodies due to toxins leaking through the intestinal wall, which also leads to nutritional deficiencies. Symptoms can include altered bowel habits, nausea, and, in some cases, paranoid thinking. Celiac disease has a genetic component, with susceptibility linked to carrying the human leukocyte antigen (HLA)-DQ2 or DQ8. The only effective treatment is to adhere to a lifelong gluten-free diet to manage the disease and reduce symptoms.

Distinction between a normal gut and a leaky gut. Source

Gluten sensitivity presents symptoms similar to those of celiac disease but is associated with increased epithelial barrier function. This can be demonstrated using the lactulose-mannitol test. In this test, decreased epithelial barrier function leads to increased lactulose permeability across the small intestine, resulting in higher levels of lactulose in the urine of affected individuals.

Gluten ataxia is a progressive disorder where gluten affects the cerebellum of the brain. It is one of the extraintestinal manifestations of gluten-related disorders. In this condition, gluten can cause paraneoplastic cerebellar degeneration, impacting limb coordination, speech, and vision. Some cases have shown a reduction in cerebellum size following gluten consumption. Research suggests that antigliadin antibodies damage the central nervous system, as post-mortem examinations of affected patients reveal large numbers of T lymphocytes and B lymphocytes around the cerebellar cortex, along with patchy loss of Purkinje cells. Lifelong avoidance of gluten remains a key measure for managing this disorder.

Mapping of a-gliadin motifs. Those exerting cytotoxic activity are shown in red, immunomodulatory activity in yellow, zonulin release and gut permeating activity in blue, and CXCR3-dependent IL-8 release in celiac disease patients in dark green. Source

What makes bread so tasty? Any healthier alternatives?

Wheat has been a staple in human diets for over 10,000 years, primarily consumed in the form of bread. The unique viscoelastic properties of wheat dough contribute to the desirable texture and crunch of the finished product. However, the health risks associated with gluten have prompted many to seek alternatives. A gluten-free diet is a common solution, with foods considered gluten-free if they contain less than 20 ppm of gluten, according to the Food and Drug Administration in the United States.

There are several gluten-free alternatives available, often made from rice or corn starch along with fibers, hydrocolloids, and specific enzymes. To boost the nutritional content of gluten-free bread, whole grains like quinoa, sorghum, buckwheat, millet, and amaranth are added, as well as ingredients like carob germ flour, chickpea flour, and pea isolate.

Despite these efforts, gluten-free bread often lacks appeal due to its low volume, pale crust, bland flavor, crumbly texture, and high rate of staling. Addressing these issues requires understanding what gluten-free bread lacks compared to gluten-based bread. Wheat-based bread contains around 77 volatile aromatic compounds that contribute to its flavor and aroma. Scientists are working to replicate these compounds in gluten-free bread through the Maillard reaction of aromatic precursors to achieve similar taste and scent.

To improve texture, milk proteins are added to enhance dough elasticity, and chemically modified starches such as Hydroxypropyl Distarch Phosphate (HDP) and Acetylated Distarch Adipate (ADA) are used to increase the volume of gluten-free loaves. The addition of linseed mucilage can also enhance sensory acceptance.

Enzymatic treatments are another approach. Transglutaminase can substitute for hydrocolloids by modifying protein functionality and cross-linking, mimicking the elasticity of gluten-based doughs. Protease enzymes like bacillolysin, papain, and subtilisin can increase loaf volume by 30 to 60% and reduce crumb hardness by 10-30% compared to untreated bread.

Additionally, sourdough fermentation has garnered interest for its potential benefits in gluten-free breadmaking. The peptidase in sourdough helps detoxify wheat and rye proteins, enhances the extractability of bioactive compounds from flour, and improves the flavor profile of gluten-free bread.

Gluten-free bread. Source

Any silver lining? If at all?

Overall, gluten-free bread is more expensive compared to its gluten-containing counterparts. However, the market for gluten-free products is gradually expanding. Beyond human consumption, gluten also has applications in pet food and non-edible uses such as removing ink from waste paper, manufacturing pressure-sensitive medical bandages and adhesives, solidifying waste oils, and creating biodegradable resins. Gluten films are used to protect food from moisture and bacteria, demonstrating that it is not as detrimental as some media portray.

Conclusion

Though gluten was discovered about three hundred years ago, it has recently gained significant attention due to increasing awareness of gluten-related disorders. Fortunately, we no longer live in an era with no gluten-free options. Today, there are many available, though improvements in their taste and texture are still needed. The pursuit of gluten-free alternatives has opened up new business opportunities. So, encourage your friends who feel deprived of gluten—it’s not too far-fetched to imagine a future where new wheat cultivars are developed without the toxic polypeptides in gluten. Achieving this will require a deep understanding of protein folding, both experimentally and theoretically. After reading this article, will you take up the challenge to advance this field?

References

Footnotes

  1. When we refer to maida it means only the contents of the endosperm; atta refers to the contents of the whole wheat grain
  2. P: Proline; Q: Glutamine; F: Phenylalanine; Y: Tyrosine
  3. Each of both types has two different N and C-terminal domains. The N-terminal domain (40–50% of total proteins) consists mostly of repetitive sequences rich in glutamine, proline, phenylalanine, and tyrosine and is unique for each type. The repetitive units of α\alpha/β\beta -gliadins are dodecapeptides such as QPQPFPQQPYP which are usually repeated five times and modified by the substitution of single residues. The typical unit of γ−\gamma-gliadins is QPQQPFP, which is repeated up to 16 times and interspersed by additional residues. Within the C-terminal domains, α\alpha/β\beta - and γ−\gamma-gliadins are homologous. They present sequences that are non-repetitive, have less glutamine and proline than the N-terminal domain, and possess are more usual composition. Studies on the secondary structure have indicated that the N-terminal domains of α\alpha/β\beta - and γ−\gamma-gliadins are characterized by β−\beta-turn conformation, similar to ω-gliadins. The non-repetitive C-terminal domain contains considerable proportions of α−\alpha-helix and β−\beta-sheet structures.