Transgenic expression in Arabidopsis of a polyprotein construct leading to production of two different antimicrobial proteins.
We developed a method for expression in Arabidopsis of a transgene encoding a cleavable chimeric polyprotein. The polyprotein precursor consists of a leader peptide and two different antimicrobial proteins (AMPs), DmAMP1 originating from Dahlia merckii seeds and RsAFP2 originating from Raphanus sativus seeds, which are linked by an intervening sequence ("linker peptide") originating from a natural polyprotein occurring in seed of Impatiens balsamina. The chimeric polyprotein was found to be cleaved in transgenic Arabidopsis plants and the individual AMPs were secreted into the extracellular space. Both AMPs were found to exert antifungal activity in vitro. It is surprising that the amount of AMPs produced in plants transformed with some of the polyprotein transgene constructs was significantly higher compared with the amount in plants transformed with a transgene encoding a single AMP, indicating that the polyprotein expression strategy may be a way to boost expression levels of small proteins.
François
Isabelle E J A
IE
Centre of Microbial and Plant Genetics, Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Heverlee, Belgium.
De Bolle
Miguel F C
MF
Dwyer
Geoff
G
Goderis
Inge J W M
IJ
Woutors
Piet F J
PF
Verhaert
Peter D
PD
Proost
Paul
P
Schaaper
Wim M M
WM
Cammue
Bruno P A
BP
Broekaert
Willem F
WF
D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000302:isolation & purification* / Q000494:pharmacology; D000935:Antifungal Agents / Q000302:isolation & purification / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000235:genetics*; D017360:Arabidopsis / Q000235:genetics*; D023082:Defensins; D005670:Fusarium / Q000187:drug effects; D018506:Gene Expression Regulation, Plant; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism; D029069:Impatiens / Q000235:genetics; D008969:Molecular Sequence Data; D010936:Plant Extracts / Q000737:chemistry; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified; D010957:Plasmids / Q000235:genetics; D020815:Polyproteins / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification; D016415:Sequence Alignment
D000890:Anti-Infective Agents; D000935:Antifungal Agents; D023181:Antimicrobial Cationic Peptides; D023082:Defensins; C099969:Dm-AMP1 protein, Dahlia merckii; D010936:Plant Extracts; D010940:Plant Proteins; D020815:Polyproteins; D011993:Recombinant Fusion Proteins; D005954:Glucosephosphate Dehydrogenase