An alpha-amylase inhibitor gene from Phaseolus coccineus encodes a protein with potential for control of coffee berry borer (Hypothenemus hampei).
Plant alpha-amylase inhibitors are proteins found in several plants, and play a key role in natural defenses. In this study, a gene encoding an alpha-amylase inhibitor, named alphaAI-Pc1, was isolated from cotyledons of Phaseolus coccineus. This inhibitor has an enhanced primary structure to P. vulgaris alpha-amylase inhibitors (alpha-AI1 and alpha-AI2). The alphaAI-Pc1 gene, constructed with the PHA-L phytohemaglutinin promoter, was introduced into tobacco plants, with its expression in regenerated (T0) and progeny (T1) transformant plants monitored by PCR amplification, enzyme-linked immunosorbent assay (ELISA) and immunoblot analysis, respectively. Seed protein extracts from selected transformants reacted positively with a polyclonal antibody raised against alphaAI-1, while no reaction was observed with untransformed tobacco plants. Immunological assays showed that the alphaAI-Pc1 gene product represented up to 0.05% of total soluble proteins in T0 plants seeds. Furthermore, recombinant alphaAI-Pc1 expressed in tobacco plants was able to inhibit 65% of digestive H. hampei alpha-amylases. The data herein suggest that the protein encoded by the alphaAI-Pc1 gene has potential to be introduced into coffee plants in order to increase their resistance to the coffee berry borer.
de Azevedo Pereira
Railene
R
EMBRAPA Recursos Genéticos e Biotecnologia, Brasília - DF 70770-900, Brazil.
Nogueira Batista
João Aguiar
JA
da Silva
Maria Cristina Mattar
MC
Brilhante de Oliveira Neto
Osmundo
O
Zangrando Figueira
Edson Luiz
EL
Valencia Jiménez
Arnubio
A
Grossi-de-Sa
Maria Fátima
MF
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003069:Coffee / Q000469:parasitology; D018548:Cotyledon / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000378:metabolism; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010572:Pest Control, Biological / Q000379:methods; D027805:Phaseolus / Q000737:chemistry / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism*; D030821:Plants, Genetically Modified; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics; D014026:Nicotiana / Q000235:genetics*; D048889:Weevils / Q000502:physiology; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
D003069:Coffee; D004791:Enzyme Inhibitors; D010940:Plant Proteins; D011994:Recombinant Proteins; C516708:alphaAI protein 1, Phaseolus coccineus; D000516:alpha-Amylases