Characterisation of saporin genes: in vitro expression and ribosome inactivation.
A. Saponaria (soapwort) genomic library was screened with a PCR-derived saporin-specific gene probe. The nucleotide sequences of three saporin genomic clones were determined. One of the clones contained a full-length saporin coding sequence whilst the other two were truncated. A hybrid full-length saporin coding sequence was constructed using the two truncated clones. An SP6 promoter sequence and in-frame initiation codon was added to each of the coding sequences using PCR. In vitro translation of saporin coding sequence transcripts in rabbit reticulocyte lysates resulted in the specific depurination of 28S RNA. This indicated that the saporin sequences encoded functional polypeptides with RNA N-glycosidase activity.
Fordham-Skelton
A P
AP
Department of Biological Sciences, University of Durham, UK.
Taylor
P N
PN
Hartley
M R
MR
Croy
R R
RR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004247:DNA; D015870:Gene Expression; D000922:Immunotoxins; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics*; D010944:Plants / Q000235:genetics; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012313:RNA / Q000378:metabolism; D011817:Rabbits; D015183:Restriction Mapping; D054788:Ribosome Inactivating Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D012270:Ribosomes / Q000378:metabolism*; D000076985:Saporins; D016415:Sequence Alignment
D000922:Immunotoxins; D010940:Plant Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D012313:RNA; D004247:DNA; D009699:N-Glycosyl Hydrolases; D054788:Ribosome Inactivating Proteins; D000076985:Saporins