Kinetics of the inhibition of Fusarium serine proteinases by barley (Hordeum vulgare L.) inhibitors

Anja I. Pekkarinen (Corresponding Author), Colin Longstaff, Berne L. Jones

Research output: Contribution to journalArticleScientificpeer-review

26 Citations (Scopus)

Abstract

Fungal infections of barley and wheat cause devastating losses of these food crops. The endogenous proteinase inhibitors produced by plant seeds probably defend the plants from pathogens by inhibiting the degradation of their proteins by the pathogen proteases. We have studied the interactions of barley grain inhibitors with the subtilisin-like and trypsinlike proteinases of Fusarium culmorum. The inhibition kinetics of three inhibitor proteins, chymotrypsin/subtilisin inhibitor 2 (CI-2), barley α-amylase/subtilisin inhibitor (BASI), and Bowman−Birk trypsin inhibitor (BBBI), have been studied in detail for the first time using fungal enzymes. The kinetic studies were performed at physiological pH values to mimic in vivo conditions. Numerical approaches to kinetic analyses were used to calculate the inhibition constants, because the data analyses were complicated by some inhibitor turnover and the instability of enzymes and substrates. All were slow, tight-binding inhibitors that followed either a two-step mechanism (CI-2 and BASI) or a single-step mechanism (BBBI) under the conditions investigated. The overall Ki values derived were approximately 50 pM, 1 nM, and 0.1 nM for CI-2, BASI, and BBBI, respectively. The main difference between the CI-2 and the BASI inhibitions was accounted for by the stabilities of their final complexes and the rate constants for their second dissociation steps (9 × 10-6/s and 3 × 10-4/s, respectively).
Understanding the inhibition mechanisms will be valuable in designing improved strategies for increasing the resistance of the grains to fungal infections.
Original languageEnglish
Pages (from-to)2736-2742
JournalJournal of Agricultural and Food Chemistry
Volume55
Issue number7
DOIs
Publication statusPublished - 2007
MoE publication typeA1 Journal article-refereed

Keywords

  • Barley
  • Fusarium
  • proteinase
  • tight-binding inhibitors

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