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The following individuals and organisations from 13 countries contributed to the PROTALL project during its lifetime:
- Dr Karin Hoffmann-Sommergruber
- Dr Heimo Breiteneder
- Professor Dietrich Kraft
- Dr Christoff Ebner
- Professor Otto Scheiner
Divisions of Immunopathology, Applied
Experimental Pathology and General and Experimental Pathology, University of Vienna.(WGs1
and 3)
Three working groups will be involved,
Biotechnology and Biochemical Diagnostics (headed by Dr Breiteneder), Allergy Research
(headed by Dr. Ebner) and Molecular Immunopathology (headed by Dr. Valenta). The research
of these groups is focussed on the production of recombinant tree pollen and food
allergens, their immunological and molecular characterisation, together with investigating
the functional aspects of plant allergens. Research is also focussed on clinically
observed allergic cross-reactions to pollen and food allergens, together with animal
models for Type I allergic reactions.
Department of Food Chemistry, Graz University
of Technology. (WG2)
The research team has an interest in allergic
reactions to food proteins, and the effects of processing on allergenic activity.
Contributions will involve investigating the effects of hydrolysis of purified allergens
supplied by other partners, on cytokine secretion of human cell lines obtained from cord
blood. There is also an interest in using immunological methods for following changes in
allergens after processing and their detection on foods. Expertise in the assembly of
databanks will also be brought to the project as members of the team have been involved in
establishing one covering the availability of foods free from allergens or intolerant
substances (eFID-databank).
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- Dr Lien-Ahh Tran
- Dr James Astwood
International Life Sciences Institute (ILSI)
European Branch, Allergy Task Force, Brussels. (WGs2 and 3)
International Life Sciences Institute (ILSI) as a nonprofit
worldwide foundation established in 1978 to advance the understanding of scientific issues
relating to nutrition, food safety, toxicology, risk assessment and the environment. It is affiliated to
the World Health Organisation as a non-governmental organisation and has specialised
consultative status with the Food and Agriculture Organisation of the United Nations.
ILSI Europe was created to focus on the specific
needs defined by the Institute's European partners. Its main goals are to:
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foster scientific advances by promoting
collaboration among scientific experts in industry, academia, and national and
international regulatory bodies
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provide coherent scientific answers to issues of
common concern for the well-being of the general public
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support an active publication programme for the
dissemination of scientific information to the broadest possible audience including the
scientific community, international organisations and regulatory agencies
James Astwood from Monsanto is contributing directly to the Protall
workprogramme. This is being done
through sharing information between the Allergy Task Force and the Protall consortium.
The ILSI Task force also form an important link for dissemination of information
to the
European food industry.
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Institute of Food Safety and Toxicology,
Danish Food and Veterinary Administration, Soeborg. (WG1)
IFT will contribute to the project in a number
of ways. An oral infant rat model for assessing sensitisation to allergens in the young is
being developed. This will be used to study sensitisation to allergens supplied by other
partners. The rats' response will be determined using, for example, serum IgE and IgG
levels, together with histopathological examination of intestine and related lymphoid
tissue. Finally IFT will contribute to the discussion on important allergenic foods that
might be included in a database. IFT is already involved in the EU network on natural
toxicants, NETTOX, which does not include information on allergens at present.
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Nutrition and Allergy related to Mucosal
Immunology (NAMI) Group, Department of Paediatrics, Turku University. (WG1)
The NAMI group will contribute to the project
by evaluating the immunological activity of purified allergenic proteins, supplied by
other partners, using children with documented food allergies. The effects of the plant
food proteins on gut mucosal barrier function will also be assessed, together with an
immunological evaluation of the gut humoral response together with cytokine generation,
both in vivo and in vitro.
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- Dr Marie-Francoise Gautier
INRA Unitede Biochemie et Biologie Moleculaire
des Cereales, Montpellier. (WG3)
For many years the UBBMC has been studying the
genes encoding low molecular weight cysteine-rich proteins from wheat. Certain of these,
the CM proteins, have been found to be involved in determining the surface state of cooked
pasta. Furthermore the lipid-transfer proteins (LTPs) and puroindolines have a high
affinity for lipid and are thought to play a role in determining bread-making quality. In
addition to their relevance in determining the technological properties of food, the LTPs
and puroindolines have anti-fungal activities, whilst the CM proteins are a -amylase
inhibitors. It is a long term goal to manipulate these proteins in order to improve the
technological properties of wheats by genetic engineering to modify their properties
and/or by genetic transformation. The contribution to the project will be to produce
purified wheat cysteine-rich proteins for assessment of their allergenic potential and to
produce modified (by site directed mutagenesis) or hybrid proteins, in order to identify
sequences important for allergenicity. Transgenic plants superexpressing cysteine-rich
proteins will also be analysed when available.
INRA Laboratoire de Biochemie et Technologie
des Proteines, Nantes. (WG2)
LBTP is composed of different units among
including one involved in the purification of proteins at laboratory and pilot scale
(S.BEROT, J.P.COMPOINT), immunochemistry (T.GABORIT), biochemistry and biophysics
(D.MARION, J.PDOULIEZ, G.COMPOINT). In this concerted action we will perform large scale
purification of some plant lipid binding proteins including puroindolines, lipid transfer
proteins and thionins for exchange with other partners. We will also use various
monoclonal and polyclonal antibody preparations to these proteins to probe them during
food processing. We will also investigate their lipid binding properties as necessary, as
well as providing other partners with different lipoprotein complexes.
SPI-BIO, Paris. (WG2)
The company has great experience in the
exploitation of molecular methods (both genetic and immunological) and providing them in a
suitable format for use by researchers. Thus using purified proteins, and possible
antibody preparations available from other partners, assays maybe developed for
determining the presence of allergens in foods, or for monitoring the alterations in
allergen structure resulting from processes, such as cooking. In certain instances
antibody preparations maybe developed to purified allergens to facilitate assay
development. The assays would also be applicable to clinical studies for monitoring the
uptake of dietary allergens into the circulation.
Laboratoire d'Immuno-Allergie
Alimentaire, Laboratoire Associe INRA-CEA, Paris. (WG1)
Development of methods for the evaluation of
allergic hazards due to food or technology additives and natural or processed food
proteins also constitutes an important element of the research projects on toxicological
aspects of food safety at INRA. The team has been much involved in the study of protein
technology and digestion of food proteins for many years. Techniques for controlled and
modeled hydrolysis, artificial digestion, preparation and identification of pure peptides
form part of the Institute's expertise. The team also has a great deal of experience
in mapping the allergenic epitopes of food allergens. This expertise will be brought to
bear using the purified protein allergens supplied by other partners.
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Allergy Unit, P & A Kyriakou
Chideren's Hospital, University of Athens. (WG1)
The Allergy Unit is a paediatric allergy
referral centre which sees in excess of 60000 patients a year. This will ensure that
sufficient patients with food allergy can be identified to undertake the work proposed.
The Immunology Laboratory is part of the University Research Laboratory and is equipped to
support the in vitro part of the study which will be to investigate T-cell mediated
immune responses. The tasks will be incorporated in the current research programme dealing
with the immune response to food allergens, in an effort to advance and develop reliable
diagnostic procedures in food allergy.
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Blackrock Clinic, Blackrock, Co. Dublin,
Ireland
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Faculty of Agriculture and Environmental
Quality Services, The Hebrew University of Jerusalem, Rehovot. (WG3)
The teams interests are focussed around the
genetic basis of yield physiology in grain legume crops such as chickpea. Attempts are
being made to combine day-length insensitivity with high biomass production. Such a
combination will allow the productivity of crops to be increased in arid zones without the
need for supplementary irrigation. Another research objective is to understand the genetic
basis of nutritional characters (eg. calcium or carotenoid contents) and their
relationship with yield parameters and fungal resistance. This expertise will bring the
aspects of crop physiology and genotype x environment interactions to bear on the
allergenicity of proteins which may be important in conferring fungal resistance to
plants.
The Institue of Biochemsitry, Nutrition and
Food Science, The Hebrew University of Jerusalem, Rehovot. (WG1)
The clinical practice of the team leader at
the hospital is primarily concerned with treating patients with gastrointestinal and
nutritional problems among which food allergy is a major component. Patients are also
treated who have inflammatory bowel disease and other gastrointestinal abnormalities
associated with digestion, absorption and allergic reactions to foods. The contribution to
the project would centre around these patients and investigating their reactivity (humoral
responses) in vitro using allergens supplied by other partners. A human nutrition
unit for obtaining blood and tissue samples is available at the two participating medical
centres. Reactivity to allergens will be determined using a range of biochemical methods.
Technicon, Haifa. (WG2)
The interests of the research team are
focussed around the effects of processing and food component interactions on the ability
of food allergens to stimulate the immune system. Plant foods will be subjected to various
different types of treatment, including heat, homogenisation (shear), acid and alkali.
Complexes of allergens with lipid, carbohydrate, as well as protein-protein aggregates
which may have formed through cross-linking or as a consequence of protein unfolding
during processing, will be isolated. The allergenic potential of these samples will be
determined by using animal models following IgG and IgE levels, histological analysis of
the intestinal mucosa and related lymph nodes, together with alterations in mast call
protease II.
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- Professor Vincenzo Buoncore
Diprtimento di Agrobiologia e Agrochimica,
Universita della Tuscia, Viterbo. (WG3)
The group is actively involved in the
isolation and characterisation of proteins and genes related to pathogenesis, also known
as PR proteins. These have been isolated from wheat and include the group 4 (wheatwins)
and group 6 (WTI, a trypsin inhibitor) which are strong inhibitors of phytopathogenic
fungi. A number of allergens from birch and other tree pollens are homologous to PR
proteins. As PR proteins are target transgenes and subjects for mutagenesis to create
forms with enhanced activity, their potential allergenicity is an issue of importance.
Class 4 and 6 PR proteins will therefore be isolated for use by other partners in
assessing their allergenicity. The primary structure of the proteins will be determined
and compared with known allergens. PR proteins of other classes with potential allergenic
features will be identified.
- Professor Elide Pastorello
Regional Reference Centre for Allergic
Disease, Osperdale Maggiore di Milano. (WG1)
The research of the team is focussed on the so
called ‘oral allergy syndrome' (OAS). which is generally observed in response to
fruits and vegetables. Such reactions are also commonly associated with sensitisation to
pollens, particularly from mugwort and birch. Research is underway to identify and
characterise new food allergens from vegetables using sera from patients with OAS for
fruits or vegetables. Especial attention will be paid to patients with allergy to tomato
as the allergens in this fruit have yet to be identified. This work will also make use of
relevant proteins available from other partners.
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Department of Gastroenterology, University
Hospital, Maastricht. (WG1)
The research team has a great interest in the
role of food allergy in the appearance of abdominal complaints, especially in conjunction
with functional abdominal complaints or chronic inflammatory bowel disease. Because of the
drawbacks of IgE RAST testing in adults with food allergy, a project has started in which
colonic intramucosal allergen injection is evaluated. Furthermore, the department has
broad scientific experience in the determination of small bowel permeability. A novel
method to determine colonic permeability will be evaluated soon. The department will
contribute to the project by contributing to a serum and mucosal specimen bank from
patients with known (multiple) food allergies. This will be used to evaluate plant
allergens from other partners in clinical studies ( possibly by intramucosal injection) or
in vitro using cell lines to determine cell permeability. The department may
contribute to in vitro and in vivo studies to elaborate whether a change in small
bowel/colonic permeability has a predictive value for, or may serve as a sensitive
parameter of the process of sensitisation to a food allergen.
RIKILT-DLO, Wageningen. (WG1)
RIKILT-DLO has a high level of expertise in
the area of food safety evaluation of foods, including novel foods, including those
originating from GMOs. Contributions will therefore focus around these areas, using
existing testing methods to assess the allergenic potential of purified proteins from
other partners in the project.
Stichting Consument en Biotechnologie, Den
Haag. (WG2)
The Consumer and Biotechnology foundation has
many years experience in consumer related issues regarding genetically modified foods. One
of the main items has been the safety of genetically modified foods for human consumption,
including the issue of food allergy in relation to genetic engineering. It is also of
interest to the organisation to translate new scientific knowledge into regulation aimed
at consumer protection and consumer information. The contribution to the project will
mainly focus on the consumers interest in scientific research and dissemination of the
results of the project to an audience of consumer organisations and their members.
Agrotechnological Research Instutue (ATO-DLO),
Wageningen. (WG2)
As a consequence of better preservation of
sensory and health-related quality attributes of food products, there is a general
tendency to apply less stringent processing techniques in the manufacture of foods. Such
milder processing methods will, however, also affect the allergenic properties of food
products. The feasibility of a combined biochemical and physical approach to processing
can be evaluated by characterising the effects of protease treatment and mild physical
processing on the structure of epitopes in proteinaceous food allergens. Once such
epitopes have been identified proteases of suitable specificity will be selected to
irreversibly modify the epitope structure. The effects of a limited number of physical
treatments, such as time-temperature combinations, on epitope structure will also be
assessed. Epitope integrity will be assessed by following the interaction between modified
epitopes and suitable antibody preparations using immunochemical assays.
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Instituto de Estructura de la Materia, CSIC,
Madrid. (WG3)
The main objective of the research undertaken
by the group is to understand protein folding and stability. Such studies require a sound
background in methods of structure determination. The group has contributed some original
methodology in that area and the laboratory is a national facility for protein structure
determination by NMR, determining structures of proteins supplied by research groups all
over Spain. Some of the recently assigned structures have been for plant proteins, one of
them, napinIb being a 2S albumin seed protein, which is highly homologous to the major
allergen of mustard, sinIa. This expertise will be brought to bear on the relevant tasks
of the project, especially the identification of common structures of allergens, and the
application of that knowledge to the prediction of allergenic determinants in novel
proteins.
- Professor Rosalia Rodriguez
- Dr Rafael Monsalve
Departmento de Biochimiqua y Biologia
Moleculair, Universidad Complutense, Madrid. (WG3)
The research of the team is focussed on the
detection, isolation and analysis of allergens from different plant sources, including
mustard seeds and olive tree pollen. More recently recombinant allergens have been
produced to study the cross-reactivity of allergens from different plants. The team will
contribute purified natural and recombinant allergens, together with chimeric proteins for
use in the structural, immunological and biological analysis of allergens. These
preparations will also be available for clinical studies and the analysis of the
cross-reactive nature of many plant allergens. The thermal and proteolytic stability of
allergens, their detection on seeds, fruits and vegetables will also be covered.
- Dr Gabriel Salcedo
- Dr Maria Rosa Sanchez-Monge
Department de Biotechnologia Unidad de
Bioquimica, Universidad Politecnica Madrid. (WG3)
Purified wheat, barley and rye allergens
associated with baker's asthma will be analysed with respect to the biochemical
properties of the allergens, and clinical data relating to their allergenic properties.
This will include a family of a -amylase inhibitors, of which there are around 15 members,
and an endosperm specific peroxidase. The role of complex asparagine-linked glycans play
in conferring the allergenic properties of certain plant proteins, especially in relation
to heat-stable epitopes, will be investigated. In addition putative
‘panallergens' implicated in the ‘latex-fruit' syndrome will be
characterised at a biochemical level. The biological activity of the isolated proteins,
both in vivo and in vitro will be assayed.
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- Dr Vaidrius Navikas
- Gran Akesson
SIK, Lund. (WG2)
SIK has an ongoing research programme into the
effects of process induced changes in protein structure on immunological molecular
recognition events. This covers unfolding and exposure of hydrophobic domains. There is
also an interest in applying immunological methods to monitoring structural changes in
allergens during food processing, with particular emphasis on the detection of allergens
in food. Thus contributions to the project will focus on characterising alterations to
food allergens during processing and interactions with food components, using biochemical
and immunological methods.
Pharmacia and Upjohn Diagnostics, Uppsala.
(WG1)
Pharmacia & Upjohn Diagnostics has been
involved in research, development, and marketing of immunoassay systems for the
measurement of allergen specific IgE antibodies. The current allergen research programme
includes immunochemical characterisation of allergen sources, allergen molecular biology,
studies on pure allergenic proteins produced by recombinant techniques, together with
clinical studies on the properties and performance of allergens and allergen reagents.
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DuPont Cereals Innovation Centre, Cambridge.
(WG2)
DuPont Cereals Innovation Centre is a
biotechnology company involved in the production of food, medical and industrial products.
Among the activities which are undertaken at the research centre is the development of new
functional protein products. It is of interest to investigate the allergenicity of these
protein products and to understand how processing effects allergenicity. Protein samples
from different plant sources, produced by different processes, can be supplied to other
partners for evaluation.
- Rene Crevel
- Dr Paul Davis
Unilever Research, Sharnbrook. (WGs1 and 2)
The SEAC Toxicology laboratory has extensive
expertise in using in vitro and in vivo toxicological studies, including
those for assessing the allergenicity of substances. In addition to having projects
involved in validating procedures for determining the immunotoxic potential of substances,
studies have also been undertaken to investigate the effects of high fat diets on the
immune response. The methodology in place for these projects will be used to assess the
allergenic potential of protein preparations supplied by other partners in the project.
Zeneca-Central Toxicology
Laboratory, Macclesfield, Cheshire. (WG3)
- Dr Clare Mills
- Dr Annette Fillery-Travis
Institute of Food Research, Norwich. (WG2)
The expertise of the team is focussed on
investigating the role of lipids and proteins at interfaces found extensively in food
systems, such as foams and emulsions. Thus the surface active properties of proteins
provided by other partners, and their interactions with lipids will be determined using a
range of interfacial methods. This research interest has also been used to investigate the
effect of surface active components (such as bile salts) and dietary lipids on the
bioavailability of vitamin A from carrots. Model systems that mimic the pH and composition
of the gut have been developed and these will be used to ascertain the effect of lipids
and bile components on the uptake of allergens by the gut.
- Professor Peter Shewry
- Dr Arthur Tatham
Long Ashton Research Station, Bristol.
(WG3)
The research team has extensive experience in
the purification and characterisation of plant proteins. The contribution to the project
will be based on this expertise. Thus in relation to sub-task 1.2 information will be
supplied on allergens from wheat and related cereal grains, the 2S albumins of
dicotyledonous seeds, including walnut, Brazil nut, castor bean, cottonseed, oriental
mustard and yellow mustard, together with the storage globulins of soyabean and peanut.
With respect to task 2 work will focus on the 2S albumins, using structural prediction,
modelling and biophysical (spectroscopic) studies to compare their structures and
stabilities. Non-allergenic 2S albumins, notably from sunflower, will be used in addition
to the previously characterised allergens listed under sub-task 1.2. On the basis of data
from these tasks the utilisation of allergens and related proteins (e.g. non-allergenic
methionine-rich 2S albumins) in transgenic plants.
With regards supply of proteins (sub-task 4.1)
proteins prepared will include prolamins (from wheat), 2S albumins (from sunflowers and
other species, depending on the availability of samples from other participants) and lipid
binding proteins (PLTPs) from sunflower. The conformation of related allergenic and
non-allergenic proteins prepared under sub-task 4.1 will be compared using spectroscopic
methods.
- Professor John Warner
- Kate Grimshaw
Department of Child Health, University of
Southampton. (WG1)
The research group has extensive expertise in
the area of allergy, and has ongoing research projects to investigate the role certain
allergens play in the development of severe allergic reactions to certain foods such as
peanuts. Clinical cases of peanut allergy have been enrolled in studies and a clinical
suite for investigations involving patients is available. These resources, together with
laboratories for in vitro tests involving cell culture and purification and
characterisation of proteins, will be used to assess the allergenic potential of proteins
supplied by other partners in the project.
Anaphylaxis Campaign, PO Box 149, Fleet,
Hampshire GU13 9XU
The Anaphylaxis Campaign is a registered charity which spreads
awareness and educates the public about life-threatening food allergies. It
produces a wide range of information sheets and videos, primarily targeted at
patients but also at the food industry, health professionals, educational
establishments, Government agencies and other bodies. There is also a strong
campaigning role, particularly in the field of food labelling. The Campaign is
guided by some of the leading allergy experts in the UK, including Prof John
Warner. With 6,000 members by the summer of 2000, the Campaign feels it has a
wealth of experiences from which to draw, and is in a good position to represent
the patient's interest.
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Dr Bryan Hanley
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Dr Heather Hird
Central Science Laboratory, York
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