Cloning and expression of rapeseed procruciferin in Escherichia coli and crystallization of the purified recombinant protein.
Two rapeseed cruciferin cDNAs (cru2/3a and cru2/3b) were cloned and sequenced. A comparison of their DNA and protein sequences with other cruciferins, indicated cru2/3b to be a novel clone and, among them, an inherent and highly conserved sequence of twelve amino acids was identified. Procruciferin 2/3a and 2/3b were expressed in Eschericha coli, and procruciferin 2/3a was obtained in a soluble form. The expressed procruciferin 2/3a has a trimeric structure and formed crystals although the quality was not good, suggesting that this expression system is useful for protein engineering of procruciferin 2/3a.
Tandang
Mary Rose G
MR
Laboratory of Food Quality Design and Development, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
Adachi
Motoyasu
M
Utsumi
Shigeru
S
D000485:Allergens; D000595:Amino Acid Sequence; D052179:Antigens, Plant; D029741:Brassica rapa / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular / Q000379:methods*; D003460:Crystallization / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000648:ultrastructure; D055314:Seed Storage Proteins; D012639:Seeds / Q000235:genetics / Q000378:metabolism; D017385:Sequence Homology
D000485:Allergens; D052179:Antigens, Plant; D010940:Plant Proteins; D011994:Recombinant Proteins; D055314:Seed Storage Proteins; C059777:cruciferin protein, Brassica napus