Cloning and expression of cDNA coding for bouganin.
Bouganin is a ribosome-inactivating protein that recently was isolated from Bougainvillea spectabilis Willd. In this work, the cloning and expression of the cDNA encoding for bouganin is described. From the cDNA, the amino-acid sequence was deduced, which correlated with the primary sequence data obtained by amino-acid sequencing on the native protein. Bouganin is synthesized as a pro-peptide consisting of 305 amino acids, the first 26 of which act as a leader signal while the 29 C-terminal amino acids are cleaved during processing of the molecule. The mature protein consists of 250 amino acids. Using the cDNA sequence encoding the mature protein of 250 amino acids, a recombinant protein was expressed, purified and characterized. The recombinant molecule had similar activity in a cell-free protein synthesis assay and had comparable toxicity on living cells as compared to the isolated native bouganin.
den Hartog
Marcel T
MT
Tanox Pharma B.V., Amsterdam, the Netherlands; Dipartimento di Patologia Sperimentale, Università di Bologna, Bologna, Italy. mtdenhartog@hotmail.com
Lubelli
Chiara
C
Boon
Louis
L
Heerkens
Sijmie
S
Ortiz Buijsse
Antonio P
AP
de Boer
Mark
M
Stirpe
Fiorenzo
F
D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D019684:Magnoliopsida / Q000737:chemistry*; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D054788:Ribosome Inactivating Proteins; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D011994:Recombinant Proteins; D009699:N-Glycosyl Hydrolases; D054788:Ribosome Inactivating Proteins