Universidade Estadual de Campinas

Relationship between zein and coixin --Laura M. M. Ottoboni, Adilson Leite, Maria Luiza N. Targon, Marcio J. DaSilva and Paulo Arruda The prolamins from seeds of maize and Job's tears (Coix lachryma-jobi L.) were extracted with 55% isopropanol containing 2% of 2-mercaptoethanol and subjected to SDS-PAGE on gradient gels. The prolamins of Coix were subdivided into five bands namely, C1 - 27kDa, C2 - 25kDa, C3 - 22kDa, C4 - 17.5kDa, and C5 - 15kDa (Fig. 1A, lane 1). These coixin bands are in the same molecular weight range as zeins which were subdivided into the well-known molecular weight classes of 27kDa (Z1, gamma zein), 22kDa (Z2, alpha zein), 19kDa (Z3, alpha zein) and 15kDa (Z4, Z5, Z6, Z7, beta zein) (Fig. 1A, lane 2).

Figure 1. SDS-PAGE and Western blots of zein and coixin extracted from endosperms of maize var. Maya and Coix lachryma-jobi var. Adlay. (A) SDS-PAGE of coixin (lane 1) and zein (lane 2). (B) Western blot of coixin (lane 1) and zein (lane 2) of replicate gel from (A) using C2 antiserum. (C) Western blot of coixin (lane 1) and zein (lane 2) of replicate gel from (A) using C3 antiserum.

Polyclonal antibodies raised against coixin C2 and C3 were used to study the immunological properties of coixin and zein. Two replicate gels containing the same samples as in Fig. 1A were transferred to nitrocellulose sheets and blotted separately with C2 and C3 antiserum. C2 antiserum recognised both C1 and C2 coixins (Fig. 1B, lane 1), but did not cross-react with any zein band (Fig. 1B, lane 2). C3 antibodies reacted with the C3 coixin and cross-reacted with a 40.5kDa protein present in the Coix extract (Fig. 1C, lane 1). Surprisingly the C3 antibodies cross-reacted strongly with gamma zein and also with proteins of 40.5 and 50.0kDa present in the maize extract (Fig. 1C, lane 2).

The coixins and zeins were extracted with 55% isopropanol with and without 2-mercaptoethanol and analysed by SDS-PAGE. Both C3 and gamma zein presented similar behaviour as they were not extracted in the absence of the reducing agent. A further similarity in the properties of the gamma zein and C3 is that antibodies raised against C3 protein cross-reacted with high molecular weight polypeptides (Fig. 1C). The nature of these high molecular weight proteins is unknown. They could be an aggregation of C3 or gamma zein subunits, or genuine high molecular weight proteins with gamma zein and C3 homology.

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