ovomucoid (Gal d 1)

4.2 Properties of Ovomucoid

4.2.1 Molecularbiological Properties

Ovomucoid (OM) References

Allergen Nomenclature Gal d 1 (1) King et al. 1994

Molecular Mass Mr 28.0 kDa (1, 3)
26.0-27.7 kDa in mass spectrometry (2)
25-31 kDa unresolved distribution in mass spectrometry (3) (1) Holen & Elsayed 1990
(2) Haginaka et al. 1995
(3) Besler et al. 1997

Isoelectric Point pI 4.4 - 4.6
2 isoforms in IEF/PAGE (1)
(1) Holen & Elsayed 1990

Amino Acid Sequence
SWISS-PROT: P01005
186 residues (1) (1) Kato et al. 1987

cDNA Sequence
EMBL: J00902
5.6 kb (1) Sequence (2) (1) Lai et al. 1979
(2) Catterall et al. 1980

mRNA Sequence
821 nucleotides (1) Sequence (2) (1) Lai et al. 1979
(2) Catterall et al. 1980

recombinant OM
expression in Escherichia coli:
domain I (OM 1-68) (Mr 7500) (3)
domain III (OM 131-186) (1, 2) (1) Hinck et al. 1993
(2) Kojima et al. 1994
(3) DeKoster & Robertson 1997

3D-Structure
X-ray studies of enzyme complex with domain III from turkey OM (1)
NMR studies of domain III from turkey OM (3, 4)
NMR studies of OM-glycopeptide (2)
NMR studies of a penta-antennary N-glycan (5) (1) Bode et al. 1986
(2) Davis et al. 1994
(3) Krezel et al. 1994
(4) Hoogstraten et al. 1995
(5) Rutherford et al. 1995

Posttranslational Modifications
Disulfide bonds
9 disulfide bonds: 5-44, 22-41, 30-62, 70-109, 87-106, 95-127, 138-168, 146-165, 154-186 (2, 6)
Glycosylation of OM:
carbohydrate content: 22-29% of whole Mr (1)
carbohydrate composition: 14-16% GlcNAc, 6.5-8.5% Man, 0.5-4.0% Gal and, 0.04-2.2% NeuNAc (1)
5 N-glycosylation sites: 10, 53, 69, 75 and partly 175 (3, 7)
covalent multiantennary structures of glycans (sequential exoglucosidase digest, mass spectrometry) (4, 5, 8)
hydrazinolysis of sialyl-oligosaccharides and sialydase digestion (6)
(1) Beeley 1971
(2) Beeley 1976a
(3) Beeley 1976b
(4) Yamashita et al. 1982
(5) Yamashita et al. 1983
(6) Yamashita et al. 1984
(7) Kato et al. 1987
(8) Yet et al. 1988

Genetic Variants
deletion of Val-134-Ser-135 (1) (1) Kato et al. 1987

Biological Function
serin protease inhibitor, kazal family of protease inhibitors (1)
3 tandem domains (1):
domain I (OM 1-68)
domain II (OM 65-130)
domain III (OM 131-186)
active sites: 24-25, 89-90, 148-149 (1) (1) Kato et al. 1987

Stability
lower trypsin-inhibitory activity and heat denaturation stability of chemically deglycosylated OM (1)
glycosylated first domain has increased thermal stability in comparison with recombinant domain I (2) (1) Gu et al. 1989
(2) DeKoster & Robertson 1997

Ovomucoid (OM)	References
*Allergen Nomenclature* Gal d 1	(1) King et al. 1994
*Molecular Mass* Mr 28.0 kDa (1, 3) 26.0-27.7 kDa in mass spectrometry (2) 25-31 kDa unresolved distribution in mass spectrometry (3)	(1) Holen & Elsayed 1990 (2) Haginaka et al. 1995 (3) Besler et al. 1997
*Isoelectric Point* pI 4.4 - 4.6 2 isoforms in IEF/PAGE (1)	(1) Holen & Elsayed 1990
*Amino Acid Sequence* SWISS-PROT: P01005 186 residues (1)	(1) Kato et al. 1987
*cDNA Sequence* EMBL: J00902 5.6 kb (1) Sequence (2)	(1) Lai et al. 1979 (2) Catterall et al. 1980
*mRNA Sequence* 821 nucleotides (1) Sequence (2)	(1) Lai et al. 1979 (2) Catterall et al. 1980
*recombinant OM* expression in Escherichia coli: domain I (OM 1-68) (Mr 7500) (3) domain III (OM 131-186) (1, 2)	(1) Hinck et al. 1993 (2) Kojima et al. 1994 (3) DeKoster & Robertson 1997
*3D-Structure* X-ray studies of enzyme complex with domain III from turkey OM (1) NMR studies of domain III from turkey OM (3, 4) NMR studies of OM-glycopeptide (2) NMR studies of a penta-antennary N-glycan (5)	(1) Bode et al. 1986 (2) Davis et al. 1994 (3) Krezel et al. 1994 (4) Hoogstraten et al. 1995 (5) Rutherford et al. 1995
*Posttranslational Modifications* Disulfide bonds 9 disulfide bonds: 5-44, 22-41, 30-62, 70-109, 87-106, 95-127, 138-168, 146-165, 154-186 (2, 6) Glycosylation of OM: carbohydrate content: 22-29% of whole Mr (1) carbohydrate composition: 14-16% GlcNAc, 6.5-8.5% Man, 0.5-4.0% Gal and, 0.04-2.2% NeuNAc (1) 5 N-glycosylation sites: 10, 53, 69, 75 and partly 175 (3, 7) covalent multiantennary structures of glycans (sequential exoglucosidase digest, mass spectrometry) (4, 5, 8) hydrazinolysis of sialyl-oligosaccharides and sialydase digestion (6)	(1) Beeley 1971 (2) Beeley 1976a (3) Beeley 1976b (4) Yamashita et al. 1982 (5) Yamashita et al. 1983 (6) Yamashita et al. 1984 (7) Kato et al. 1987 (8) Yet et al. 1988
*Genetic Variants* deletion of Val-134-Ser-135 (1)	(1) Kato et al. 1987
*Biological Function* serin protease inhibitor, kazal family of protease inhibitors (1) 3 tandem domains (1): domain I (OM 1-68) domain II (OM 65-130) domain III (OM 131-186) active sites: 24-25, 89-90, 148-149 (1)	(1) Kato et al. 1987
*Stability* lower trypsin-inhibitory activity and heat denaturation stability of chemically deglycosylated OM (1) glycosylated first domain has increased thermal stability in comparison with recombinant domain I (2)	(1) Gu et al. 1989 (2) DeKoster & Robertson 1997

4.2.2 Allergenic Properties

Ovomucoid (OM) References

Immunoglobulines
IgE-binding studies of OM (1)
median percentage of serum IgE:
33% against domain I (OM 1-68)
48% against domain II (OM 65-130)
28% against domain III (OM 131-186)
in 45 egg allergic children with atopic dermatitis
(total OM-specific serum IgE = 100%) (RAST) (2) (1) see 4.1 Sensitization to Egg White Allergens
(2) Cooke & Sampson 1997

B-Cell Epitopes
IgE binding sites located on:
OM 1-20 (synthetic peptide) (b) (3)
OM 49-56 (synthetic peptide) (b) (3)
OM 85-96 (synthetic peptide) (b) (3)
OM 90-121 (trypsin digest) (a) (4)
OM 115-122 (synthetic peptide) (b) (3)
OM 134-186 (pepsin digest) (a) (4)
OM 175-186 (synthetic peptide) (b) (3)
Carbohydrate epitopes:
specific serum IgE against N-glycosylated domain III (Asn-175) higher than against non-glycosylated domain III in egg allergic patients (1)
no difference in IgE-binding of OM and deglycosylated OM (a, c) (2)
(a) SDS-PAGE / immunoblot
(b) dot / immunoblot
(c) EAST / RAST inhibition
(1) Matsuda et al. 1985
(2) Besler et al. 1997
(3) Cooke & Sampson 1997
(4) Besler et al. 1999

T-Cell Epitopes
T-Cell Proliferation with:
domain I (OM 1-68)
domain II (OM 65-130) (predominant)
domain III (OM 131-186) (predominant)
in 4 egg allergic patients (1)
domain I (OM 1-68) in 28% '
domain II (OM 65-130)   in 55%
domain III (OM 131-186) in 64%
of 33 (' 29) egg allergic children with atopic dermatitis (3)
whole OM (2)
(1) Eigenmann et al. 1996
(2) Holen & Elsayed 1996
(3) Cooke & Sampson 1997

PBMC Stimulation / Cytokines
PBMC stimulation with OM:
expression of IL-5 in 4 egg allergic patients (1) (1) Eigenmann et al. 1996

Alteration of Allergenicity
cyanogen bromide cleavage:
no change in IgE- and IgG-binding in 6 OM allergic patients (direct ELISA) (1)
deglycosylation: see carbohydrate epitopes
performic acid oxidation:
no change in IgE- (max. inhibition 93%) and decreased IgG-binding (max. inhibition 39%) in 7 egg allergic patients (EAST inhibition) (2)
increased PBMC proliferation in 11 egg allergic patients (2)
reduction and alkylation:
IgE-binding lost in 4/6 and retained in 2/6 OM allergic patients, respectively (1)
IgG-binding lost in 4/6 and retained in 2/6 OM allergic patients, respectively (1)
decreased IgE-(max. inhibition 72%) and decreased IgG-binding (max. inhibition 82%) in 7 egg allergic patients (EAST inhibition) (2)
increased PBMC proliferation in 11 egg allergic patients (2)
(1) Djurtoft et al. 1991
(2) Cooke & Sampson 1997

Ovomucoid (OM)	References
*Immunoglobulines* IgE-binding studies of OM (1) median percentage of serum IgE: 33% against domain I (OM 1-68) 48% against domain II (OM 65-130) 28% against domain III (OM 131-186) in 45 egg allergic children with atopic dermatitis (total OM-specific serum IgE = 100%) (RAST) (2)	(1) see 4.1 Sensitization to Egg White Allergens (2) Cooke & Sampson 1997
*B-Cell Epitopes* IgE binding sites located on: OM 1-20 (synthetic peptide) (b) (3) OM 49-56 (synthetic peptide) (b) (3) OM 85-96 (synthetic peptide) (b) (3) OM 90-121 (trypsin digest) (a) (4) OM 115-122 (synthetic peptide) (b) (3) OM 134-186 (pepsin digest) (a) (4) OM 175-186 (synthetic peptide) (b) (3) Carbohydrate epitopes: specific serum IgE against N-glycosylated domain III (Asn-175) higher than against non-glycosylated domain III in egg allergic patients (1) no difference in IgE-binding of OM and deglycosylated OM (a, c) (2) (a) SDS-PAGE / immunoblot (b) dot / immunoblot (c) EAST / RAST inhibition	(1) Matsuda et al. 1985 (2) Besler et al. 1997 (3) Cooke & Sampson 1997 (4) Besler et al. 1999
*T-Cell Epitopes* T-Cell Proliferation with: domain I (OM 1-68) domain II (OM 65-130) (predominant) domain III (OM 131-186) (predominant) in 4 egg allergic patients (1) domain I (OM 1-68) in 28% ' domain II (OM 65-130) in 55% domain III (OM 131-186) in 64% of 33 (' 29) egg allergic children with atopic dermatitis (3) whole OM (2)	(1) Eigenmann et al. 1996 (2) Holen & Elsayed 1996 (3) Cooke & Sampson 1997
*PBMC Stimulation / Cytokines* PBMC stimulation with OM: expression of IL-5 in 4 egg allergic patients (1)	(1) Eigenmann et al. 1996
*Alteration of Allergenicity* cyanogen bromide cleavage: no change in IgE- and IgG-binding in 6 OM allergic patients (direct ELISA) (1) deglycosylation: see carbohydrate epitopes performic acid oxidation: no change in IgE- (max. inhibition 93%) and decreased IgG-binding (max. inhibition 39%) in 7 egg allergic patients (EAST inhibition) (2) increased PBMC proliferation in 11 egg allergic patients (2) reduction and alkylation: IgE-binding lost in 4/6 and retained in 2/6 OM allergic patients, respectively (1) IgG-binding lost in 4/6 and retained in 2/6 OM allergic patients, respectively (1) decreased IgE-(max. inhibition 72%) and decreased IgG-binding (max. inhibition 82%) in 7 egg allergic patients (EAST inhibition) (2) increased PBMC proliferation in 11 egg allergic patients (2)	(1) Djurtoft et al. 1991 (2) Cooke & Sampson 1997