Hydroxyproline and proline inhibit α-amylase from isolated barley aleurone layers

Abstract

Previously, we reported that 1 mM hydroxyproline appeared to inhibit the gibberellic acid-induced release of α-amylase from isolated Hordeum vulgare L. cv. Himalaya aleurone layers into an incubation medium. Here, we report our attempts to determine the mechanism(s) for this inhibition and whether this inhibition can be caused by other proline analogues. Both 1 mM hydroxyproline and proline inhibited extracellular a-amylase activity without affecting its intracellular activity. This suggested that neither hydroxyproline nor proline impaired the release of a-amylase. Lineweaver-Burk plots revealed that both hydroxyproline and proline uncompetitively inhibited α-amy-lase. Thus, the inhibition is probably an assay artifact resulting from the formation of an enzyme-substrate-hydroxyproline or -proline complex. Because azetidine-2-carboxylic acid, glutamic acid and pipecolic acid did not inhibit extracellular α-amylase activity, the uncompetitive inhibition of a-amylase must be unique to imino aicids as well as their precursors and derivatives which possess a five membered ring.