Multiple molecular forms of the gibberellin-induced α-amylase from the aleurone layers of barley seeds

Abstract

A class of plant growth regulators, gibberellins, induce the synthesis of α-amylase (1,4-α- d-glucan glucanohydrolase, EC 3.2.1.1) in the aleurone layers of barley ( Hordeum vulgare L. var. Himalaya) seeds. The purified α-amylase is composed of multiple isozymic forms with indistinguishable molecular weights, but different net charges. These α-amylase isozymes separate on isoelectric focusing gels into two groups, each containing multiple species. One group has an apparent isoelectric point (p I) of approximately 5.8 (the high p I group). The other group's p I values are around 4.5 (the low p I group). On some gels a small amount of protein focuses between the high and low p I isozymes. These proteins comigrate with the low p I isozymes upon reelectrophoresis. The synthesis of these two groups is temporally regulated. The high p I group is the dominant set of isozymes secreted from embryoless half seeds during the first two days of gibberellin administration. After four days, however, the major isozymes are those of the low p I group. This shift in isozyme pattern is due to a shift in their relative rates of synthesis. Peptide analysis of these two groups of isozymes with Staphylococcus aureus V8 protease and cyanogen bromide shows amino acid sequence differences. However, members within the same group have similar peptide patterns. Both groups of isozymes are synthesized in vitro in a wheat germ extract primed with poly(A) + RNA isolated from gibberellin-treated aleurone layers. This indicates that the synthesis of the two groups of α-amylase isozymes is probably directed by two or more different populations of mature mRNA. A model that explains these observations and the available genetic information is that barley aleurone α-amylase isozymes are encoded by at least two sets of structural genes.