Cloning and functional expression of the gene encoding an inhibitor against Aspergillus flavus alpha-amylase, a novel seed lectin from Lablab purpureus (Dolichos lablab).
Maize is one of the more important agricultural crops in the world and, under certain conditions, prone to attack from pathogenic fungi. One of these, Aspergillus flavus, produces toxic and carcinogenic metabolites, called aflatoxins, as byproducts of its infection of maize kernels. The alpha-amylase of A. flavus is known to promote aflatoxin production in the endosperm of these infected kernels, and a 36-kDa protein from the Lablab purpureus, denoted AILP, has been shown to inhibit alpha-amylase production and the growth of A. flavus. Here, we report the isolation of six full-length labAI genes encoding AILP and a detailed analysis of the activities of the encoded proteins. Each of the six labAI genes encoded sequences of 274 amino acids, with the deduced amino acid sequences showing approximately 95-99% identity. The sequences are similar to those of lectin members of a legume lectin-arcelin-alpha-amylase inhibitor family reported to function in plant resistance to insect pests. The labAI genes did not show any of the structures characteristic of conserved structures identified in alpha-amylase inhibitors to date. The recombinant proteins of labAI-1 and labAI-2 agglutinated human red blood cells and inhibited A. flavus alpha-amylase in a manner similar to that shown by AILP. These data indicate that labAI genes are a new class of lectin members in legume seeds and that their proteins have both lectin and alpha-amylase inhibitor activity. These results are a valuable contribution to our knowledge of plant-pathogen interactions and will be applicable for developing protocols aimed at controlling A. flavus infection.
Kim
Young-Hwa
YH
School of Food and Life Science, BPRC, Inje University, Obangdong, 607, Gyungnam, Korea.
Woloshuk
Charles P
CP
Cho
Eun Hee
EH
Bae
Jung Myung
JM
Song
Young-Sun
YS
Huh
Gyung Hye
GH
D000595:Amino Acid Sequence; D001231:Aspergillus flavus / Q000201:enzymology*; D015139:Blotting, Southern; D003001:Cloning, Molecular; D031294:Dolichos / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology; D004912:Erythrocytes / Q000187:drug effects; D018745:Genome, Plant / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D037121:Plant Lectins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology; D012639:Seeds / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
D004791:Enzyme Inhibitors; D037121:Plant Lectins; D011994:Recombinant Proteins; D000516:alpha-Amylases