Functional Study of Rice Starch Synthase I (SSI) by Using Double Mutant with Lowered Activities of SSI and Isoamylase1

Abstract

The functions of the major isozymes related to the starch biosynthesis in cereal endosperm have been resolved based on the researches using the mutant lines. For further understanding of the relationships between isozymes, the analyses of double mutant lines must be very efficient. In this study, double mutant rice line (ss1L/isa1) between leaky starch synthase I mutant line (ss1L) and isoamylase 1 mutant line (isa1) was generated and structures of the stored α-glucans were characterized. Cross-section of the parent isa1 and ss1L/isa1 seeds was negative for iodine staining and more than 90% of the α-glucans was soluble in water. Chain-length distributions of the glucans were similar to each other between the two lines, and the differential plot obtained by subtraction of the chain-length distribution of isa1 from that of ss1L/isa1 was nearly identical to the corresponding differential plot of ss1L vs. wild type, indicating effect of SSI is simply additive on chain-length distribution of the product regardless of isoamylase1 activity. Analyses of β-amylase limit dextrins revealed slight, statistically significant difference in internal chain-length distributions of the two isa1 mutant lines in contrast to the previous result on isa1 mutant lines with or without exogenous indica-type SSIIa. This study demonstrated that the primary role of SSI in the isa1 background is outer-chain elongation with a little effect on branch-position distribution, which is the same role as the case in normal amylopectin biosynthesis in the presence of isoamylase 1. Such chain-elongation properties differ among the three major SS isozymes.