Characterization of Maize Elongation Factor 1A and Its Relationship to Protein Quality in the Endosperm

Abstract

The protein synthesis elongation factor I A (eEF1 A) is a multifunctional protein in eukaryotic cells. In maize (Zea mays 1.) endosperm eEFlA co-localizes with actin around protein bodies, and its accumulation is highly correlated with the protein-bound lysine (Lys) content. We purified eEFlA from maize kernels by ammonium sulfate precipitation, ion-exchange, and chromatofocusing. The identity of the purified protein was confirmed by microsequencing of an endoproteinase glutamic acid-C fragment and by its ability to bundle actin. Using purified eEFlA as a standard, we found that this protein contributes 0.4% of the total protein in W64A+ endosperm and approximately 1 O/O of the protein in W64Ao2. Because eEFlA contains 10% Lys, it accounts for 2.2% of the total Lys in W64A+ and 2.3% of the Lys in W64Ao2. However, its concentration predicts 90% of the Lys found in endosperm proteins of both genotypes, indicating that eEFlA is a key component of the group of proteins that determines the nutritional quality of the grain. This notion is further supported by the fact that in froury2, another high-lys mutant, the content of eEFlA increases with the dosage of the f/oury2 gene. These data provide the biochemical basis for further investigation of the relationship between eEFl A content and the nutritional quality of cereals.

Moonlighting Functions of Polypeptide Elongation Factor 1: From Actin Bundling to Zinc Finger Protein R1-Associated Nuclear Localization

“…However, a mutant called``opaque'' has a higher lysine content, which correlates well with the EF-1a content. 172,173) The expression of several EF-1a genes out of 10 to 15 EF-1a genes is increased in the endosperm of the opaque2 mutant. The lysine content of zein itself is not changed in the mutant, but the ratio of some lysine-rich proteins in seeds to zein is increased.…”

Cytoskeletal Proteins Are Coordinately Increased in Maize Genotypes with High Levels of eEF1A

“…eEF1A. For example, the levels of a-zeins, pyruvate orthophosphate dikinase, and acetolactate synthase were decreased in o2 mutants compared to wild type (Damerval and Le Guilloux, 1998;Hunter et al, 2002), while that of eEF1A was increased (Habben et al, 1993;Sun et al, 1997). For some enzymes involved in carbohydrate metabolism, such as glyceraldehyde-3-phosphate dehydrogenase and sorbitol dehydrogenase, earlier studies showed these proteins are increased in o2 mutants (Damerval and Le Guilloux, 1998).…”

“…Other Lys-rich proteins that exist in association with this cytoskeletal network may be increased as the consequence of a greater protein body surface area (Sun et al, 1997). Oh51Ao2 has more a-zein and also appears to have a larger number of small protein bodies (larger RER surface area) than Oh545o2 (Wang et al, 2001), which could result in a more extensive ERassociated cytoskeletal network.…”

“…However, the biological basis of this relationship is unclear. eEF1A contains 11% Lys, but this protein only accounts for 2.3% of the Lys in W64Ao2 endosperm (Sun et al, 1997). This suggests a stoichiometric relationship exists between eEF1A and the other major proteins contributing to the Lys content of the grain.…”

“…Biochemical characterization of eEF1A from maize endosperm revealed this protein binds F-actin (Sun et al, 1997;Lopez-Valenzuela et al, 2003) and immunolocalization showed eEF1A is associated with an F-actin network that surrounds the rough endoplasmic reticulum (RER) at sites where protein bodies are forming (Clore et al, 1996). Wang et al (2001) found a quantitative trait locus linked with eEF1A content that mapped with a cluster of 22-kD a-zein genes.…”