Protein structures of common bean (Phaseolus vulgaris) alpha-amylase inhibitors.
Two nucleotide sequences for genes that encode alpha-amylase inhibitor 4 (alphaAI-4) from white kidney bean (WKB) cv. 858, designated gene alphaAI-4 (Accession No. ), and alpha-amylase inhibitor 5 (alphaAI-5) from black bean (BB), designated gene alphaAI-5 (Accession No. ), were determined. Genes alphaAI-4 and alphaAI-5 encode 244 amino acid prepro-alphaAI-4 and prepro-alphaAI-5 polypeptides that are 93 and 95% identical with alpha-amylase inhibitor l (alphaAI-l; Hoffman, L. M.; Ma, Y.; Barker, R. F. Nucleic Acids Res. 1982, 10, 7819-7828), 40 and 43% identical with red kidney bean lectin, and 52 and 55% identical with arcelin l of wild-type bean. The high degree of sequence similarity indicates the evolutionary relationship among these genes. PCR analysis of genomic DNA purified from six genotypes of Phaseolus vulgaris showed very similar band patterns in 2% agarose gel, another indication of the conserved size homology among these genes. Proteolytic processing sites were located between Asn77 and Ser78 for pro-alphaAI-4 and pro-alphaAI-5. A bend next to Asn77 in three-dimensional model structures of alphaAI-4 and alphaAI-5 proinhibitors indicates that the proteolytic cleavage is necessary to remove the conformational constraint for activation to the mature protein. Mature WKB alphaAI-4 was composed of four subunits (2alpha2beta) and had a molecular weight of 50000 determined by multiangle laser light scattering and 56714 determined by laser-assisted time-of-flight mass spectrometry.
Lee
Shih-Chieh
SC
AdvPharma, Inc., Tainen, Taiwan, Republic of China.
Gepts
Paul L
PL
Whitaker
John R
JR
D000595:Amino Acid Sequence; D001483:Base Sequence; D018744:DNA, Plant / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry; D036341:Intercellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D027805:Phaseolus / Q000737:chemistry / Q000378:metabolism*; D037121:Plant Lectins / Q000737:chemistry* / Q000235:genetics; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D020836:Protein Structure, Quaternary; D017386:Sequence Homology, Amino Acid; D000516:alpha-Amylases / Q000037:antagonists & inhibitors
D018744:DNA, Plant; D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; D037121:Plant Lectins; D010940:Plant Proteins; C061510:alpha-amylase inhibitor, Phaseolus vulgaris; C090486:arcelin 5 protein, Phaesolus vulgaris; D000516:alpha-Amylases