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A novel strategy for inhibition of α-amylases: Yellow meal worm α-amylase in complex with the Ragi bifunctional inhibitor at 2.5 Å resolution

Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule Hönggerberg, Zürich, Switzerland.
Structure (Impact Factor: 6.79). 08/1998; 6(7):911-21. DOI: 10.1016/S0969-2126(98)00092-6
Source: PubMed

ABSTRACT alpha-Amylases catalyze the hydrolysis of alpha-D-(1,4)-glucan linkages in starch and related compounds. There is a wide range of industrial and medical applications for these enzymes and their inhibitors. The Ragi bifunctional alpha-amylase/trypsin inhibitor (RBI) is the prototype of the cereal inhibitor superfamily and is the only member of this family that inhibits both trypsin and alpha-amylases. The mode of inhibition of alpha-amylases by these cereal inhibitors has so far been unknown.
The crystal structure of yellow meal worm alpha-amylase (TMA) in complex with RBI was determined at 2.5 A resolution. RBI almost completely fills the substrate-binding site of TMA. Specifically, the free N terminus and the first residue (Ser1) of RBI interact with all three acidic residues of the active site of TMA (Asp185, Glu222 and Asp287). The complex is further stabilized by extensive interactions between the enzyme and inhibitor. Although there is no significant structural reorientation in TMA upon inhibitor binding, the N-terminal segment of RBI, which is highly flexible in the free inhibitor, adopts a 3(10)-helical conformation in the complex. RBI's trypsin-binding loop is located opposite the alpha-amylase-binding site, allowing simultaneous binding of alpha-amylase and trypsin.
The binding of RBI to TMA constitutes a new inhibition mechanism for alpha-amylases and should be general for all alpha-amylase inhibitors of the cereal inhibitor superfamily. Because RBI inhibits two important digestive enzymes of animals, it constitutes an efficient plant defense protein and may be used to protect crop plants from predatory insects.

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    • "enzymes that are found in microorganisms, plants and animals. This enzyme catalyses the hydrolysis of a-D-(1,4)-glucan linkage in starch and related carbohydrates (Strobl et al. 1998). a-Amylase activity has been described from different species of several insect orders including Coleoptera, Hymenoptera, Diptera, Lepidoptera and Hemiptera (Baker and Woo 1985; Terra et al. 1988; Mendiola-Olaya et al. 2000; Oliveira-Neto et al. 2003; Kazzazi et al. 2005). "
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    • "α-Amylase is an important group of digestive enzymes (Franco et al. 2000; Bandani et al. 2009). These enzymes catalyse the hydrolysis of α-D-(1, 4)-glucan linkage in starch components, glycogen and other carbohydrates and are found in animals, plants and micro-organisms (Strobl et al. 1998). Different patterns of activity and its isoforms depends on the type of diet (Mendiola-Olaya et al. 2000; Kotkar et al. 2009). "
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    • "A comprehensive study (Strobl et al. 1998) on larval digestive !-amylase of Tenebrio molitor L. (Coleoptera: Tenebrionidae) showed that the enzyme has three domains. The central domain (domain A) is an (b/a)8-barrel that comprises the core of the molecule and includes catalytic amino acid residues, and domains B and C are almost opposite to each other, on each side of domain A (Strobl et al. 1998). Abundance and activity of insect !-amylases in the gut are dependent on food sources, so feeding on wool and plant tissues causes the lowest and highest amylolytic activity, respectively (Chapman 1998; Zibaee et al. 2008). "
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