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Food Microbiology 24 (2007) 115–119

Chemistry of gluten proteins

Herbert Wieser
German Research Centre of Food Chemistry and Hans-Dieter-Belitz-Institute for Cereal Grain Research, D-85748 Garching, Germany
Available online 7 September 2006


Gluten proteins play a key role in determining the unique baking quality of wheat by conferring water absorption capacity, cohesivity,
viscosity and elasticity on dough. Gluten proteins can be divided into two main fractions according to their solubility in aqueous
alcohols: the soluble gliadins and the insoluble glutenins. Both fractions consist of numerous, partially closely related protein
components characterized by high glutamine and proline contents. Gliadins are mainly monomeric proteins with molecular weights
(MWs) around 28,000–55,000 and can be classified according to their different primary structures into the a/b-, g- and o-type. Disulphide
bonds are either absent or present as intrachain crosslinks. The glutenin fraction comprises aggregated proteins linked by interchain
disulphide bonds; they have a varying size ranging from about 500,000 to more than 10 million. After reduction of disulphide bonds, the
resulting glutenin subunits show a solubility in aqueous alcohols similar to gliadins. Based on primary structure, glutenin subunits have
been divided into the high-molecular-weight (HMW) subunits (MW ¼ 67,000–88,000) and low-molecular-weight (LMW) subunits
(MW ¼ 32,000–35,000). Each gluten protein type consists or two or three different structural domains; one of them contains unique
repetitive sequences rich in glutamine and proline. Native glutenins are composed of a backbone formed by HMW subunit polymers and
of LMW subunit polymers branched off from HMW subunits. Non-covalent bonds such as hydrogen bonds, ionic bonds and
hydrophobic bonds are important for the aggregation of gliadins and glutenins and implicate structure and physical properties of dough.
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Keywords: Gluten; Gliadins; Glutenins; Proline and glutamine

1. Gluten weights (MWs) of native proteins range from around

30,000 to more than 10 million. Traditionally, gluten
Gluten can be defined as the rubbery mass that remains proteins have been divided into roughly equal fractions
when wheat dough is washed to remove starch granules according to their solubility in alcohol–water solutions of
and water-soluble constituents. Depending on the thor- gluten (e.g. 60% ethanol): the solubles gliadins and the
oughness of washing, the dry solid contain 75–85% protein insoluble glutenins. Both fractions are important contri-
and 5–10% lipids; most of the remainder is starch and non- butors to the rheological properties of dough, but their
starch carbohydrates. In practice, the term ‘gluten’ refers to functions are divergent. Hydrated gliadins have little
the proteins, because they play a key role in determining elasticity and are less cohesive than glutenins; they
the unique baking quality of wheat by conferring water contribute mainly to the viscosity and extensibility of the
absorption capacity, cohesivity, viscosity and elasticity on dough system. In contrast, hydrated glutenins are both
dough. Gluten contains hundreds of protein components cohesive and elastic and are responsible for dough strength
which are present either as monomers or, linked by and elasticity. To simplify matters, gluten is a ‘two-
interchain disulphide bonds, as oligo- and polymers component glue’, in which gliadins can be understood as
(Wrigley and Bietz, 1988). They are unique in terms of a ‘plasticizer’ or ‘solvent’ for glutenins. A proper mixture of
their amino acid compositions, which are characterized by both fractions is essential to impart the viscoelastic
high contents of glutamine and proline and by low contents properties of dough and the quality of the end product.
of amino acids with charged side groups. The molecular Though cysteine belongs to the minor amino acids of
gluten proteins (E2%), it is extremely important for the
E-mail address: structure and functionality of gluten (Grosch and Wieser,

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116 H. Wieser / Food Microbiology 24 (2007) 115–119

1999; Wieser, 2003). Most cysteines are present in an into four different types: o5-, o1,2-, a/b- and g-gliadins
oxidized state and form either intrachain disulphide bonds (Table 1) (Wieser, 1996). Within each type, structural
within a protein or interchain disulphide bonds between differences are small due to substitution, deletion and
proteins. These bonds are the main target for most redox insertion of single amino acid residues. o-Gliadins are
reactions that occur during kernel maturation, milling, characterized by the highest contents of glutamine, proline
dough preparation and baking (Wieser, 2003). Additional and phenylalanine which together account for around 80%
covalent bonds formed during breakmaking are tyrosine– of the total composition. o5-Gliadins have higher MWs
tyrosine crosslinks between gluten proteins (Tilley et al., (E50,000) than o1,2-gliadins (E40,000). Most o-gliadins
2001) and tyrosine–dehydroferulic acid crosslinks between lack cysteine, so that there is no possibility of disulphide
gluten proteins and arabinoxylans (Piber and Koehler, crosslinks. These proteins consist almost entirely of
2005). The covalent structure of the gluten network is repetitive sequences rich in glutamine and proline (e.g.
superimposed by non-covalent bonds (hydrogen bonds, PQQPFPQQ). a/b- and g-gliadins have overlapping MWs
ionic bonds, hydrophobic bonds). Though this class of (E28,000–35,000) and proportions of glutamine and pro-
chemical bonds is less energetic than covalent bonds, they line much lower than those of o-gliadins (Table 1). They
are clearly implicated in gluten protein aggregation and differ significantly in the contents of a few amino acids, e.g.
dough structure (Wieser et al., 2006). Evidence for the tyrosine. Each of both types has two clearly different N-
presence of hydrogen bonds in gluten proteins are the and C-terminal domains. The N-terminal domain (40–50%
dough weakening effect of hydrogen bond breaking agents of total proteins) consists mostly of repetitive sequences
(e.g. urea) and the dough strengthening effect of heavy rich in glutamine, proline, phenylalanine and tyrosine and
water compared with that of ordinary water. The is unique for each type (sequence Sections I and II, Fig. 1).
importance of ionic bonds can be demonstrated by the The repetitive units of a/b-gliadins are dodecapeptides such
strengthening effect of NaCl or of bipolar ions such as as QPQPFPQQPYP which are usually repeated five times
amino acids or dicarboxylic acids. Hydrophobic bonds and modified by the substitution of single residues. The
contribute significantly to the stabilization of gluten typical unit of g-gliadins is QPQQPFP, which is repeated
structure. They are different from other bonds, because up to 16 times and interspersed by additional residues.
their energy increases with increasing temperature; this can Within the C-terminal domains, a/b- and g-gliadins are
provide additional stability during the baking process. homologous (sequence Sections III–V, Fig. 1). They
present sequences which are non-repetitive, have less
2. Gliadins glutamine and proline than the N-terminal domain and
possess are more usual composition. With a few exceptions,
Most gliadins are present as monomers; they were a/b-gliadins contain six and g-gliadins eight cysteines
initially classified into four groups on the basis of mobility located in the C-termional domain; they form three and
at low pH in gel electrophoresis (a-, b-, g-, o-gliadins in four homologous intrachain crosslinks, respectively
order of decreasing mobility). Later studies on amino acid (Grosch and Wieser, 1999) (Fig. 1). Studies on the
sequences, however, have shown that the electrophoretic secondary structure have indicated that the N-terminal
mobility does not always reflect the protein relationships domains of a/b- and g-gliadins are characterized by b-turn
and that a- and b-gliadins fall into one group (a/b-type). conformation, similar to o-gliadins (Tatham and Shewry,
Modern methods such as two-dimensional electrophoresis 1985). The non-repetitive C-terminal domain contains
or reversed-phase high-performance liquid chromatogra- considerable proportions of a-helix and b-sheet structures.
phy (RP-HPLC) allow the separation of the gliadin Though the distribution of total gliadins among the
fraction into more than hundred components. Based on different types is strongly dependent on wheat variety
the analysis of complete or partial amino acid sequences, (genotype) and growing conditions (soil, climate, fertiliza-
amino acid compositions and MWs, they can be grouped tion), it can be generalized that a/b- and g-gliadins are

Table 1
Characterisation of gluten protein types

Type MW  103 Proportionsa (%) Partial amino acid composition (%)

Gln Pro Phe Tyr Gly

o5-Gliadins 49–55 3–6 56 20 9 1 1

o1,2-Gliadins 39–44 4–7 44 26 8 1 1
a/b-Gliadins 28–35 28–33 37 16 4 3 2
g-Gliadins 31–35 23–31 35 17 5 1 3
x-HMW-GS 83–88 4–9 37 13 0 6 19
y-HMW-GS 67–74 3–4 36 11 0 5 18
LMW-GS 32–39 19–25 38 13 4 1 3
According to total gluten proteins.
H. Wieser / Food Microbiology 24 (2007) 115–119 117

(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.
After reduction of disulphide bonds, the resulting
glutenin subunits show a solubility in aqueous alcohols
similar to gliadins. The predominant protein type are
LMW glutenin subunits (LMW-GS), their proportion
amounts to E20% according to total gluten proteins
(Wieser and Kieffer, 2001). LMW-GS are related to
a/b- and g-gliadins in MW and amino acid composition
(Table 1). Similarly, they contain two different domains:
The N-terminal domain (sequence Section I, Fig. 1)
consists of glutamine- and proline-rich repetitive units
such as QQQPPFS and the C-terminal domain (sequence
Sections III–V, Fig. 1) is homologous to that of a/b- and g-
gliadins within Sections III and V. LMW-GS contain eight
cysteines (Grosch and Wieser, 1999; Wieser, 2003); six
residues are in positions homologous to a/b- and g-gliadins,
and therefore, are proposed to be linked by intrachain
disulphide bonds (Fig. 1). Two additional cysteine residues
unique to LMW-GS are located in Sections I and IV. They
are not able to form an intrachain bond, probably for steric
reasons. Consequently, interchain disulphide bonds with
cysteines of different gluten proteins are generated.
HMW glutenin subunits (HMW-GS) belong to the minor
components within the gluten protein family (E10%). Each
wheat variety contains three to five HMW-GS which can be
grouped into two different types, the x- and the y-type, with
MWs from 83,000–88,000 and 67,000–74,000, respectively
Fig. 1. Schematic architecture and disulphide structures of gluten proteins (Table 1). The nomenclature on single HMW-GS is based
(adapted from Grosch and Wieser, 1999).
on the coding genome (A, B, D), the type (x, y) and the
mobility of SDS-PAGE (nos. 1–12). HMW-GS consist of
major components, whereas the o-gliadins occur in much three structural domains (Fig. 1): a non-repetitive N-
lower proportions (Wieser and Kieffer, 2001) (Table 1). A terminal domain (A) comprising about 80–105 residues, a
minor portion of gliadins have an odd number of cysteines repetitive central domain (B) of about 480–700 residues and
due to point mutations and are linked together or to a C-terminal domain (C) of 42 residues (Shewry et al., 1992).
glutenins. They appear either in alcohol-soluble oligomers Domains A and C are characterized by the frequent
in the gliadin fraction or in the alcohol-insoluble glutenin occurrence of charged residues and by the presence of most
polymers. This form of gliadins is proposed to act as or all of cysteines. Domain B contains repetitive hexapep-
terminator of glutenin polymerization (see below). The tides (unit QQPGQG) as a backbone with inserted
oligomeric fraction has been called high-molecular-weight hexapeptides (e.g. YYPTSP) and tripeptides (e.g. QQP or
(HMW) gliadin, aggregated gliadin or ethanol-soluble QPG). The most important difference between the x- and
glutenin (Shewry et al., 1983; Huebner and Bietz, 1993). the y-type lies within the A and B domains. For example, the
It contains a/b-, g-gliadins and low-molecular-weight y-type has an insertion of 18 residues including two
(LMW) subunits linked by interchain disulphide bonds; neighbouring cysteines in domain A, and typical repetitive
the MWs range from around 100,000–500,000. units are less frequently repeated and more frequently
modified in domain B of the y-type. Because HMW-GS does
3. Glutenins not occur in flour and dough as monomers, it is generally
assumed that they form interchain disulphide bonds. The x-
The glutenin fraction comprises aggregated proteins type except subunit Dx5 has four cysteines, three in domain
linked by interchain disulphide bonds; they have varying A and one in domain C (Shewry and Tatham, 1997) (Fig. 1).
size ranging from about 500,000 to more than 10 million Two residues of domain A are linked by an intrachain, the
(Wieser et al., 2006). Thus, a part of glutenins belongs to other two by interchain disulphide bonds. Subunit Dx5 has
the largest proteins in nature. The MW distribution of an additional cysteine at the beginning of domain B and it
glutenins has been recognized as one of the main has been suggested that this might form another interchain
determinants of dough properties and baking performance. bond. The y-type has five cysteines in domain A and one in
The largest polymers termed ‘glutenin macropolymer’ each of domains B and C. At present, interchain bonds have
118 H. Wieser / Food Microbiology 24 (2007) 115–119

only be found for the adjacent cysteines of domain A which a/b- and g-gliadins present three and four intrachain
are connected in parallel with the corresponding residue of disulpide bonds, respectively, whereas polymeric LMW-
another y-type, and for cysteine of domain B which is linked and HMW-GS include both intra- and interchain bonds
to a cysteine of LMW-GS. (Fig. 1) (Shewry and Tatham, 1997). The disulphide
Various approaches have been combined to give details structure of native glutenins is not in a stable state, but
of the secondary structure of HMW-GS. Studies of the undergoes a continuous change from the maturing grain to
repetitive domain B indicated the presence of b-reverse the end product (e.g. bread) (Wieser, 2003; Wieser et al.,
turns (Shewry et al., 1992). These were predicted to be 2006). The MW distribution of glutenins has been
overlapping and form a loose spiral which was considered recognized as one of the main determinants of dough
to contribute decisively to gluten’s elasticity. The non- quality and is governed by the state of disulphide structure
repetitive domains A and C were proposed to have that depends on genetic factors (e.g. presence of subunit
globular structures containing a-helices. Numerous studies Dx5, ratio of HMW-GS to LMW-GS, amount of chain
demonstrated that dough properties are strongly influenced terminators), environmental factors (e.g. sulphur defi-
by the quantities of HMW-GS. Among HMW-GS the ciency, heat or water stress), and the redox state (e.g.
contribution of the x-type to dough properties has been presence of reducing or oxidizing agents) (Wieser et al.,
found to be more important than that of the y-type (Wieser 2006). Disulphide formation starts rapidly after synthesis
and Kieffer, 2001). Considering single subunits, the of proteins within the lumen of endoplasmatic reticulum as
presence of subunit Dx5 (which has an additional cysteine an integral part of protein folding. It has been proposed
for an interchain crosslink) and subunit Bx7 (which occurs that intrachain links form more rapidly than interchain
in the greatest amounts) has been proposed to be links (Kasarda, 1999). To participate in a growing polymer,
particularly important for dough quality and loaf volume proteins need at least two cysteines forming interchain
(Wieser and Zimmermann, 2000). disulphide bonds. HMW- and LMW-GS usually fulfil this
requirement; they act as ‘chain extender’. The only
4. Disulphide bonds crosslink found between HMW- and LMW-GS until now
is a disulphide bond between a cysteine residue in domain
Disulphide bonds play an important role in determining B of y-HMW-GS and a cysteine residue in the C-terminal
the structure and properties of gluten proteins. Monomeric domain of LMW-GS. The backbone of HMW-GS

Fig. 2. A model double unit for the interchain disulphide structures of LMW-GS () and HMW-GS (&) of gluten polymers (adapted from Wieser et al.,
H. Wieser / Food Microbiology 24 (2007) 115–119 119

polymers is established by end-to-end, probably head-to- future research. One of the priorities should be to get more
tail linkages. At least, four different cysteines lead to insight into changes of the disulphide structure beginning
branching of the backbone. LMW-GS form also linear from the synthesis of proteins in the growing plant and
polymers via cysteines of the N-terminal and C-terminal ending in the baked products. Another point should be a
domain. Terminators of polymerization were found to be better understanding of the relations between structure and
glutathione or gliadins with an odd number of cysteines. functionality of gluten proteins, particularly in combina-
With respect to quantitative data on the subunits tion with additives such as dough improvers. Heterologous
composition of glutenins (ratio of HMW-GS to LMW- expression and protein engineering could be a valuable aid
GS E1:2 and of x- to y-type E2.5:1) and the MW of in structure–function studies (Tamas and Shewry, 2006).
subunits, the basis molecular ‘double unit’ of glutenin
polymers might consist of 2 y-type HMW-GS, 4 x-type
HMW-GS and around 30 LMW-GS covalently linked by
interchain disulphide bonds (Wieser et al., 2006) (Fig. 2). References
The MW of this double unit amounts to about 1.5 million.
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The largest of the glutenins (GMP) might include more by ascorbic acid. J. Cereal Sci. 29, 1–16.
than ten of these double units in which the x/y-type ratio Huebner, F.R., Bietz, J.A., 1993. Improved chromatographic separation
and the HMW–/LMW-GS ratio increase. and characterization of ethanol-soluble wheat proteins. Cereal Chem.
The overriding importance of disulphide bonds can be 70, 506–511.
Kasarda, D.D., 1999. Glutenin polymers: the in vitro to in vivo transition.
demonstrated by the addition of reducing agents weaken-
Cereal Foods World 44, 566–571.
ing dough and of thiol-blocking or oxidizing agents Piber, M., Koehler, P., 2005. Identification of dehydro-ferulic acid-
strengthening dough (Wieser, 2003). During the whole tyrosine in rye and wheat: evidence for a covalent cross-link between
process of breadmaking, the state of large glutenin arabinoxylans and proteins. J. Agric. Food Chem. 53, 5276–5284.
polymers is characterized by three competitive redox Shewry, P.R., Tatham, A.S., 1997. Disulphide bonds in wheat gluten
reactions: (1) the oxidation of free SH groups which proteins. J. Cereal Sci. 25, 207–227.
Shewry, P.R., Miflin, B.J., Lew, E.J.-L., Kasarda, D.D., 1983. The
support polymerization; (2) the presence of ‘terminators’ preparation and characterization of an aggregated gliadin fraction
that stop polymerization and (3) SH/SS interchange from wheat. J. Exp. Bot. 34, 1403–1410.
reactions between glutenins and thiol compounds such as Shewry, P.R., Halford, N.G., Tatham, A.S., 1992. High molecular weight
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Gluten proteins are among the most complex protein
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