Structure of the bifunctional inhibitor of trypsin and α-amylase from ragi seeds at 2.9 Å resolution

The crystal structure of a bifunctional inhibitor of α-amylase and trypsin from the seeds of ragi (Indian finger millet, Eleusine coracana Gaertneri) has been determined by an X-­ray diffraction method. The inhibitor consists of 122 amino acids with five disulfide bridges and belongs to the plant α-amylase/trypsin-inhibitor family. This is the first crystal structure determination of a member of this family. The protein, purified from the seeds of ragi, has a molecular mass of 13300 Da with a pI of 10.3. Crystals were grown by a microdialysis method using ammonium sulfate as precipitant. The improved purification protocol and the modified crystallization conditions enabled reproducible growth of the crystals. The cell parameters are a = 41.2, b = 47.4, c = 55.9 Å. The intensity data were collected to 2.9 Å resolution, and the crystal structure was determined using the molecular-replacement method. The structure was refined using the X-­PLOR and CCP4 program packages to a conventional R factor of 21%. The structure contains four α-helices between residues 19–29, 37–51, 56–65 and 90–95, and two short antiparallel β-strands between residues 67–70 and 73–75.