Ovotransferrin is a redox-dependent autoprocessing protein incorporating four consensus self-cleaving motifs flanking the two kringles.
Embryos of avian eggs and mammals are highly sensitive to oxidative stress and hence maintaining a steady reducing environment during the embryonic development is known to confer protection. Although information is completely lacking, proteins of avian egg albumin which have been suggested to play various biological functions, are the major targets for such reducing state during embryogenesis. In this study, we found that ovotransferrin (OTf), the second major protein in egg albumin, undergoes autocleavage at distinct sites upon reduction with thiol-reducing agent or thioredoxin-reducing system. Mass spectral and microsequencing analysis indicated that OTf is able to cleave itself through the unique chemical reactivity of four tripeptides motifs, HTT (residues 209-211), HST (residues 542-544) and two CHT (residues 115-117 and 454-456). Intriguingly, these self-cleavage sites were uniquely located upstream and downstream of the two disulfide kringle domains (residues 115-211 and 454-544) of OTf. These reduction-scissile sequences, His/Cys-X-Thr, are evolutionary conserved self-cleavage motifs found in several autoprocessing proteins including hedgehog proteins. Interestingly, reduction of other two members of transferrin family induced autocleavage patterns, similar to that of OTf, in bovine lactoferrin (bLf) while human lactoferrin (hLf) showed much less self-cleaving activity. This finding is the first to describe that transferrins are a new subset in the class of proteins able to carry out autoprocessing, providing insight into this unusual biochemical process that appears to be a molecular switch involved in triggering a yet unidentified function(s) of OTf as well as bLf.
Ibrahim
Hisham Radwan
HR
Department of Biochemistry and Biotechnology, Faculty of Agriculture, Kagoshima University, Kagoshima 890-0065, Japan. hishamri@chem.agri.kagoshima-u.ac.jp
Haraguchi
Taku
T
Aoki
Takayoshi
T
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002645:Chickens; D003207:Conalbumin / Q000737:chemistry*; D016384:Consensus Sequence; D004229:Dithiothreitol / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D018082:Kringles; D007781:Lactoferrin; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction
D003207:Conalbumin; D007781:Lactoferrin; D004229:Dithiothreitol