Are Transgenic Proteins Allergenic?

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5 Jan 2005 14:36:35 -0000

 

Are Transgenic Proteins Allergenic?

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ISIS Press Release 05/01/05

 

Are Transgenic Proteins Allergenic?

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Some two-thirds of all transgenic proteins have similarities to known

allergens. Should we worry? Drs. Mae-Wan Ho, Arpad Pusztai, Susan

Bardocz and Prof. Joe Cummins tell us why we should

 

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Similarities to known allergens

 

A report published in 2002 [1] should raise concerns over the safety of

the foreign proteins incorporated into GM crops that are commercially

approved.

 

Researchers at the Institute of Food Safety in Wageningen, The

Netherlands, screened transgenic proteins in GM food crops for the

presence of

short amino acid sequences identical to those in known allergens, and

to find out if these are involved in binding IgE, the class of

antibodies produced in allergic reactions.

 

They screened 33 transgenic proteins for continuous runs of at least 6

amino acids identical to known allergenic proteins. Twenty-two of the

transgenic proteins showed positive results in runs of 6 or 7 amino

acids; these include all the Bt toxins (Cry proteins), the CP4 EPSPS and

GOX conferring glyphosate tolerance, the coat protein of the papaya

ringspot virus, and even marker proteins such as GUS.

 

But on account of the limited data available, only a small number could

be identified as linear epitopes (sites) that might bind to IgE

antibodies. Although most identical stretches may be " false

positives " , the

researchers said the results " warrant further clinical testing for

potential allergenicity " .

 

How reliable are current tests for potential allergenicity?

 

Potential allergenicity is one major aspect of safety assessment of GM

crops. As many new proteins are introduced into GM food crops, it is

important to find reliable methods of assessing their potential to cause

allergic reactions, when eaten as food, through contact, or by inhaling

(as pollen, for example).

 

One of the first steps in assessing if a protein is potentially

allergenic is to compare its amino acid sequence with those of known

allergenic proteins stored in computer databases, using available

computer

algorithms.

 

When such comparisons are made, identities of continuous runs of 8 or

more amino acids are considered " immunologically relevant " . But shorter

stretches can also be relevant according to existing findings; for

example, small sequences of four and six amino acids can be recognized

and

bound by IgE antibodies from allergic patients [2].

 

Apart from these continuous or linear epitopes, discontinuous epitopes

may also be present, consisting of amino acids in different parts of

the polypeptide chain that end up next to one-another when the

polypeptide chain is folded up in its three-dimensional conformation.

Thus,

overall amino-acid similarity with an allergenic protein, i.e., 35%

identity

within a run of 80 amino acids, might be suspect. At the moment, it is

difficult to predict which amino acids may form discontinuous epitopes,

as we need to know the three-dimensional structure of the protein.

 

In addition to the linear and conformational peptide epitopes, glycans

(carbohydrate chains linked to the protein) have also been shown to be

major IgE binding sites in allergenic glycoproteins.

 

In a follow-up study published September 2004 [3], a new webtool was

used to predict potential allergenicity of proteins and peptides

according to the current recommendations of the FAO/WHO Expert

Consultation, as

outlined in the Codex Alimentarius [4, 5]. The Codex Alimentarius

Commission was created by the United Nations FAO (Food and Agriculture

Organization) and WHO (World Health Organization) to set international

food

standards.

 

The amino acid sequence of a protein is compared with all known

allergenic proteins retrieved from the protein databases to identify

stretches

of 80 amino acids with more than 35% similarity, or small identical

runs of at least 6 amino acids.

 

The ability of the procedure to predict allergens is evaluated by

screening sets of known allergens and non-allergens. Apart from making

correct predictions, both methods generated " false positive " and " false

negative " hits. The number of false negatives decreases when a larger

database of allergen sequences is used, whereas the number of false

positives grows with the size of the database.

 

" False negatives " , " false positives " and the need for precaution

 

The researchers point out that the number of false positives may be

overestimated, because some of the `non-allergens' used are related to

and

display similarities with their allergenic counterparts.

 

But that's precisely why we need to take any positive hits seriously.

In fact, at least 5 of the 12 protein sequences used as `non-allergens'

were reported to react with other classes of antibodies, IgG and IgM,

and are hence immunogenic, if not allergenic.

 

Another caveat, pointed out by the researchers, is that a protein

belonging to a completely new group of allergens is likely to generate

false

negative results. This would apply to the majority of transgenic

proteins that have never been part of our food chain.

 

As advised in the earlier publication, and also by the FAO/WHO, the

outcomes should therefore be combined with other methods of assessing

allergenicity, such as digestibility and binding of antisera from

allergic

patients, and possibly animal exposure tests. But that too, leaves a

lot to be desired.

 

It is very difficult to assess the allergenicity of GM foods when the

gene transferred into the plant is from an organism whose allergenic

potential is unknown. Moreover, it is also possible that as a result of

the gene transfer or insertion of the transgenic DNA, a new allergen is

developed, or the expression of a minor allergen is elevated in the GM

crop. The gene product can also have an allergenic adjuvant (helper)

effect on a food component previously of low allergenic potential; or

conversely, some component in the GM food may have an adjuvant effect on

the allergenicity of the transgene product.

 

Unfortunately, while there are good animal models for

nutritional/toxicological testing, no satisfactory animal models have

so far been

developed to test for allergenicity [6]. For the time being, only

indirect

methods are available for assessing the allergenic potential of GM foods

derived from sources of unknown allergenicity. The screening tests

described above are a useful preliminary step.

 

If the result is positive, then in vitro tests for IgE reaction need to

be performed, especially as most epitopes are discontinuous. The

absence of a positive in vitro reaction does not guarantee that the

transgenic protein is not an allergen. In a decision-tree type of

indirect

approach, the next step is to consider the molecular size, glycosylation,

stability, solubility and isoelecgtric point of the transgenic protein

compared with known allergens [7]. Unfortunately, in most studies

to-date, the all-important ability of the transgenic protein to resist

breakdown in the gut is investigated in an in vitro simulated

gastric/intestinal system [8, 9]; and this is fundamentally flawed.

The results are

therefore at best misleading and at worst erroneous. Reliance on the

concept that most allergens are abundant proteins is probably also

misleading because for example, Gadc1, the major allergen in codfish,

is not a

predominant protein [10].

 

In the absence of new and reliable methods for allergenicity testing,

particularly the lack of good animal models, it is at present almost

impossible to definitely establish

whether a new GM crop is allergenic or not in advance of its release

into the human/animal food/feed chain.

 

In our view, with foods consumed by millions, any positive results

should be assumed to be significant until fuller testing can definitively

rule it out as a false positive. In North America and elsewhere, GM

foods are not labelled and this may have led to the spread of

allergens not

identified as having originating with the GM foods that may in fact be

the case.

 

 

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General Enquiries sam

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