Novel proteinase inhibitors in seeds of sunflower (Helianthus annuus L.): Polymorphism, inheritance and properties

A highly sensitive gelatin overlay procedure was used to identify inhibitors of serine proteinases and of the cysteine proteinase ficin in seeds and leaves of sunflower. One major and two minor groups of trypsin inhibitors were identified in seeds, the former having a high pI (≃10) and also inhibiting chymotrypsin. Three groups of trypsin/subtilisin inhibitors were also present in seeds, together with three inhibitors of ficin. All groups showed polymorphism between lines of Helianthus annuus, while the trypsin and trypsin/subtilisin inhibitors also varied between wild species of Helianthus, with no apparent relationship to growth type (annual or perennial), genome constitution or ploidy level. Genetic analysis showed that the major trypsin inhibitor and three groups of trypsin/subtilisin inhibitors are each controlled by single Mendelian loci, with the three loci for trypsin/subtilisin inhibitors showing recombination values of 0.23-0.40. Purification by RP-HPLC allowed the Mr of two trypsin inhibitors to be determined by SDS-PAGE to be about 1,500 and 2,500, while the three trypsin/subtilisin inhibitors varied in Mr from about 1,500 to 6,000.