Crystal structure of α-amylase from Oryza sativa: molecular insights into enzyme activity and thermostability

Abstract

AmyI-1 is an α-amylase from Oryza sativa (rice) and plays a crucial role in degrading starch in various tissues and at various growth stages. This enzyme is a glycoprotein with an N-glycosylated carbohydrate chain, a unique characteristic among plant α-amylases. In this study, we report the first crystal structure of AmyI-1 at 2.2-Å resolution. The structure consists of a typical (β/α)₈-barrel, which is well-conserved among most α-amylases in the glycoside hydrolase family-13. Structural superimposition indicated small variations in the catalytic domain and carbohydrate-binding sites between AmyI-1 and barley α-amylases. By contrast, regions around the N-linked glycosylation sites displayed lower conservation of amino acid residues, including Asn-263, Asn-265, Thr-307, Asn-342, Pro-373, and Ala-374 in AmyI-1, which are not conserved in barley α-amylases, suggesting that these residues may contribute to the construction of the structure of glycosylated AmyI-1. These results increase the depths of our understanding of the biological functions of AmyI-1.

Graphical Abstract

Crystal structure of α-amylase AmyI-1 from rice.

Key words:

• α-amylase
• rice
• crystal structure
• thermostability
• glycoprotein

Funding

This work was supported by the Grants-in-Aid from the Japan Society for the Promotion of Science (to A.O. [No. 24780122] and to T.M. [No. 22380186]). We thank the beamline staff at the AR-NW12A station of the Photon Factory for their assistance with data collection. Synchrotron radiation experiments at this facility were performed as outlined in proposal number 2013G084.

Notes

Abbreviations: GH, glycoside hydrolase; AmyI-1, α-amylase isoform I-1 from Oryza sativa (rice); AMY1 and AMY2, α-amylases from Hordeum vulgare (barley); ER, endoplasmic reticulum; SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis; MALDI-TOF/MS, matrix assisted laser desorption/ionization time-of-flight mass spectrometry; ABS, absorbance; rmsd, root-mean-square deviations; PDB, Protein Data Bank; SBS, surface binding site.