77.3-77.7 kDa from mass spectra of 4 isoforms (Awade et al. 1994 [1358]).
Oligomeric Masses:
Monomer
Allergen epitopes:
Not reported.
Allergen stability: Process, chemical, enzymatic
Although ovotransferrin is highly cross-linked by disulphides, the apo form is not very stable to denaturants. Muralidhara & Hirose (2000) [1356] report that 0.88M and 1.3M guanidinium hydochloride or 1.51M and 2.61M urea gave midpoints of denaturation of a form with one disulphide reduced or native at pH 5.6.
Ovotransferrin is relatively stable to heating and DSC gave Tm of 82.5°C and 60.24°C for the diferric and apo forms (Lin et al. 1994 [1357]).
Lechevalier et al. (2003) [1259] describe the interaction of ovalbumin, ovotransferrin and lysozyme with an air water interface. Ovotransferrin was less stable than lysozyme. Contact with an interface led to an increase in surface hydrophobicity and modifications of its secondary structure characteristic of major conformational changes.
Nature of main cross-reacting proteins:
IgE cross reactivity might be expected with ovotransferrins from birds as the duck protein is 80% identical to that from chicken. No cross-reactivity is expected or reported to transferrins from liver.
Quirce et al. (2001) [1268] reported cross-reactivity between chicken serum albumin and ovotranferrin as ovotransferrin at high concentration partially inhibited IgE binding to chicken serum albumin in ELISA-inhibition assays. However, these proteins tend to co-purify.
Allergen properties & biological function:
Ovotransferrin comprises approximately 12-13 % of egg white protein and is an antimicrobial defence and iron binding protein. It functions partly by keeping iron inaccessible to microbes (cf. lactoferrin activity described by Singh et al, 2002 [1348]). Activity against microbial membranes has been described both as the intact protein and as derived peptides (Aguilera et al, 2003 [1347]; Ibrahim et al, 2000 [1349]). Anti-viral activity has also been described (Giansanti et al. 2005 [1350]) as well as a role in immunity in chickens (Xie et al, 2002 [1351]). Finally, ovotransferrin is incorporated into egg shell (Gautron et al. 2001 [1352]).
Allergen purification:
Mizutani et al. (2004) [1248] describe the expression and purification of ovotransferrin produced at a high level using a Pichia pastoris expression system. There was a single site of glycosylation which could be trimed by Endo-H treatment to give a mass of 75925 Da.
Ebbehoj et al. (1995) [1355] describe purification procedures for ovomucoid, ovotransferrin, ovalbumin, and lysozyme with less than 0.1% contaminating proteins as assessed by SDS-PAGE and crossed immunoelectrophoresis with polyclonal antibodies raised against an egg-white extract or the purified proteins.
Other biochemical information:
The mechanism of iron binding to proteins of the transferrin family and the related changes in 3-D structure have been extensively studied using ovotransferrin (Mizutani et al. 1999 [1250]; Kurokawa et al. 1999 [1249]) as well as to lactoferrin and serum transferrin. Two ferric irons are bound, one to the N-terminal and one to the C-terminal domain. Iron can bind to an apo-domain with an open iron binding site. However, the most stable structure of the domain when iron has bound is generated by a large conformational change which buries the ferric ion and surrounds it with protein ligands.
References (16)
Aguilera O, Quiros LM, Fierro JF.
Transferrins selectively cause ion efflux through bacterial and artificial membranes.
FEBS Lett. 548(1-3):5-10. 2003
PUBMED ID:
12885398
Awade AC, Moreau S, Molle D, Brule G, Maubois JL.
Two-step chromatographic procedure for the purification of hen egg white ovomucin, lysozyme, ovotransferrin and ovalbumin and characterization of purified proteins.
J Chromatogr A 677(2):279-288. 1994
PUBMED ID:
7921188
Gautron J, Hincke MT, Panheleux M, Garcia-Ruiz JM, Boldicke T, Nys Y.
Ovotransferrin is a matrix protein of the hen eggshell membranes and basal calcified layer.
Connect Tissue Res. 42(4):255-267. 2001
PUBMED ID:
11913770
Ibrahim HR, Sugimoto Y, Aoki T.
Ovotransferrin antimicrobial peptide (OTAP-92) kills bacteria through a membrane damage mechanism.
Biochim Biophys Acta 1523(2-3):196-205. 2000
PUBMED ID:
11042384
Kurokawa H, Dewan JC, Mikami B, Sacchettini JC, Hirose M.
Crystal structure of hen apo-ovotransferrin. Both lobes adopt an open conformation upon loss of iron.
J Biol Chem. 274(40):28445-28452. 1999
PUBMED ID:
10497206
Kurokawa H, Mikami B, Hirose M.
Crystal structure of diferric hen ovotransferrin at 2.4 A resolution.
J Mol Biol. 1995 Nov 24;254(2):196-207. 1995
PUBMED ID:
7490743
Lechevalier V, Croguennec T, Pezennec S, Guerin-Dubiard C, Pasco M, Nau F.
Ovalbumin, Ovotransferrin, Lysozyme: Three Model Proteins for Structural Modifications at the Air-Water Interface
J Agric Food Chem. 51(21):6354-61.
2003
PUBMED ID:
14518967
Lin LN, Mason AB, Woodworth RC, Brandts JF.
Calorimetric studies of serum transferrin and ovotransferrin. Estimates of domain interactions, and study of the kinetic complexities of ferric ion binding.
Biochemistry 33(7):1881-1888. 1994
PUBMED ID:
8110792
Mizutani K, Okamoto I, Fujita K, Yamamoto K, Hirose M.
Structural and functional characterization of ovotransferrin produced by Pichia pastoris.
Biosci Biotechnol Biochem. 68(2):376-383. 2004
PUBMED ID:
14981301
Mizutani K, Yamashita H, Kurokawa H, Mikami B, Hirose M.
Alternative structural state of transferrin. The crystallographic analysis of iron-loaded but domain-opened ovotransferrin N-lobe.
J Biol Chem. 274(15):10190-10194. 1999
PUBMED ID:
10187803
Muralidhara BK, Hirose M.
Structural and functional consequences of removal of the interdomain disulfide bridge from the isolated C-lobe of ovotransferrin.
Protein Sci. 9(8):1567-1575. 2000
PUBMED ID:
10975578
Quirce S, Maranon F, Umpierrez A, de las Heras M, Fernandez-Caldas E, Sastre J.
Chicken serum albumin (Gal d 5) is a partially heat-labile inhalant and food allergen implicated in the bird-egg syndrome.
Allergy 56(8):754-762.
2001
PUBMED ID:
11488669
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