Purification and characterization of an insect alpha-amylase inhibitor/endochitinase from seeds of Job's Tears (Coix lachryma-jobi).
A protein inhibitor of locust gut alpha-amylase was purified from seeds of Job's Tears (Coix lachryma-jobi) using ammonium sulphate precipitation, affinity chromatography on Red Sepharose and reversed-phase HPLC. It consisted of two major isomers, each a dimer of two closely similar or identical subunits of Mr about 26,400, and associated by inter-chain disulphide bonds. These isomers also had closely similar amino acid compositions. The major isomer showed no inhibitory activity against amylases from other sources: human saliva, porcine pancreas, Bacillus subtilis, Aspergillus oryzae and barley malt. The manual DABITC/PITC method was used to determine about half of the amino acid sequence of the major isoform. This showed a high degree of homology with previously reported sequences of endochitinase enzymes from several species (tobacco, potato, barley, bean), and endochitinase activity was demonstrated by following the release of radioactivity from [3H]chitin. This novel combination of functions may be relevant to protection of the grain from insect feeding and fungal infection.
Ary
M B
MB
Biology Department, University of Durham, U.K.
Richardson
M
M
Shewry
P R
PR
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002688:Chitinases / Q000302:isolation & purification* / Q000494:pharmacology; D002523:Edible Grain / Q000032:analysis; D006110:Grasshoppers / Q000201:enzymology*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000302:isolation & purification; D012639:Seeds / Q000032:analysis*; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
D000596:Amino Acids; D010940:Plant Proteins; D000516:alpha-Amylases; D002688:Chitinases