Insect pests and pathogens (fungi, bacteria and viruses) are responsible for severe crop losses. Insects feed directly on the plant tissues, while the pathogens lead to damage or death of the plant. Plants have evolved a certain degree of resistance through the production of defence compounds, which may be aproteic, e.g. antibiotics, alkaloids, terpenes, cyanogenic glucosides or proteic, e.g. chitinases, β-1,3-glucanases, lectins, arcelins, vicilins, systemins and enzyme inhibitors. The enzyme inhibitors impede digestion through their action on insect gut digestive α-amylases and proteinases, which play a key role in the digestion of plant starch and proteins. The natural defences of crop plants may be improved through the use of transgenic technology. Current research in the area focuses particularly on weevils as these are highly dependent on starch for their energy supply. Six different α-amylase inhibitor classes, lectin-like, knottin-like, cereal-type, Kunitz-like, γ-purothionin-like and thaumatin-like could be used in pest control. These classes of inhibitors show remarkable structural variety leading to different modes of inhibition and different specificity profiles against diverse α-amylases. Specificity of inhibition is an important issue as the introduced inhibitor must not adversely affect the plant's own α-amylases, nor the nutritional value of the crop. Of particular interest are some bifunctional inhibitors with additional favourable properties, such as proteinase inhibitory activity or chitinase activity. The area has benefited from the recent determination of many structures of α-amylases, inhibitors and complexes. These structures highlight the remarkable variety in structural modes of α-amylase inhibition. The continuing discovery of new classes of α-amylase inhibitor ensures that exciting discoveries remain to be made. In this review, we summarize existing knowledge of insect α-amylases, plant α-amylase inhibitors and their interaction. Positive results recently obtained for transgenic plants and future prospects in the area are reviewed.

Plant alpha-amylase inhibitors and their interaction with insect alpha-amylases.

Potent α-glucosidase inhibitors purified from the red alga Grateloupia elliptica

Protein inhibitors of enzymes, which have potentially deleterious effects in animal nutrition and possible defensive functions against insect and microbial pests, are of widespread occurrence in plants1,2. Because cereals provide about 55% of direct human protein intake their inhibitors have received much attention3. The amino-acid sequences of α-amylase inhibitors in wheat4–6, and ragi (Eleusine coracana)7; proteinase inhibitors from maize8 and barley9,10; and bifunctional proteinase/amylase inhibitors in ragi11, barley12,13 and wheat14 show varied evolutionary relationships. Here we describe the sequence of a maize protein which is a potent in vitro inhibitor of bovine trypsin and the α-amylase from Tribolium castaneum beetles. The protein has little similarity to any of the other enzyme inhibitors, but is highly similar to the intensely sweet protein thaumatin15 and to a protein induced in tobacco following infection by tobacco mosaic virus (TMV)16. This unexpected finding suggests a possible function for these proteins.

A possible function for thaumatin and a TMV-induced protein suggested by homology to a maize inhibitor

Proteinaceous α-amylase inhibitors

Publisher Summary This chapter discusses that the family Leguminosae includes approximately 600 genera, with about 13,000 species. In legume seeds, the cotyledons form the bulk of the seed and synthesize most of the proteins. Research on identification and quantification of the storage proteins in beans (Phaseolus) goes back several decades. These investigators referred to three proteins in Phaseolus (phaseolin, phaselin, and conphaseolin). The chapter discusses that bean proteins were first studied by Osborne. He found that the extractable proteins were mostly globulins, a fact which was later confirmed and emphasized by other investigators, who found three globulins in Phaseolus seeds. The biological value of bean proteins in general is low as compared to most other food proteins. According to Boyd, hemagglutinating activity was first described by Stillmark in 1888 in extracts of castor bean (Ricinus communis) and the first plant lectin found to be blood group specific was from lima bean (Phaseolus lunatus). Proteins that inhibit a-amylases from animals and insects have been reported in a variety of plants. Common beans (Phaseolus vulgaris) are normally harvested with about 20% moisture in the seeds and dried to about 10% water content before storage or consumption. The chapter discusses that common beans contain from 18% to 35% protein, however, their biological value is much lower than expected on a protein and amino acid composition basis.

Physical, chemical, and nutritional properties of common bean (Phaseolus) proteins.

α-Amylase inhibitor is presented in maize seeds. It is a protein as indicated by precipitation with ammonium sulfate and trichloroacetic acid, denaturation by heat, digestion with proteases and by dye-staining.



It was purified to homogeneity by ammonium sulfate precipitation and Sephadex G-75 gel filtration. It had an apparent molecular weight of 29,600 and did not contain any carbohydrate.



Its properties differed from those of previously reported α-amylase inhibitors, since it was active against α-amylase of maize, produced during germination as well as against Bacillus subtilis α-amylase.



It was also active against α-amylase from the insects Tribolium castaneum, Sitophilus zeamais and Rhyzopertha dominica, but it was inactive against α-amylase from human saliva, hog pancreas, Aspergillus oryzae, wheat, rye, barley, triticale, and sorghum.



It was stable for 5 min at 96°C at pH 7. Maximal inhibition required at least 10 min of preincubation with the enzyme at pH 6.8 and 257deg;C. Polyacrylamide gel electrophoresis gave three protein bands, but only one was obtained in S.D.S. and mercaptoethanol.

F. A. Iturbe-Chiñas

Papers

PURIFICATION AND CHARACTERIZATION OF AN α‐AMYLASE INHIBITOR FROM MAISE (ZEA MAIZE)