cDNA cloning, functional expression and antifungal activities of a dimeric plant defensin SPE10 from Pachyrrhizus erosus seeds.
SPE10 is an antifungal protein isolated from the seeds of Pachyrrhizus erosus. cDNA encoding a 47 amino acid peptide was cloned by RT-PCR and the gene sequence proved SPE10 to be a new member of plant defensin family. The synthetic cDNA with codons preferred in yeast was cloned into the pPIC9 plasmid directly in-frame with the secretion signal alpha-mating factor, and highly expressed in methylotrophic Pichia pastoris. Activity assays showed the recombinant SPE10 inhibited specifically the growth of several pathogenic fungi as native SPE10. Circular dichroism and fluorescence spectroscopy analysis indicated that the native and recombinant protein should have same folding, though there are eight cystein residues in the sequence. Several evidence suggested SPE10 should be the first dimeric plant defensin reported so far.
Song
Xiaomin
X
National Laboratory of Biomacromolecules, Institute of Biophysics, University of Science and Technology of China, Key Laboratory of Structural Biology, Chinese Academy of Sciences, P.R. China.
Wang
Jing
J
Wu
Fang
F
Li
Xu
X
Teng
Maikun
M
Gong
Weimin
W
D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000494:pharmacology; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D023082:Defensins / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D019281:Dimerization; D005658:Fungi / Q000187:drug effects / Q000254:growth & development; D015870:Gene Expression; D020128:Inhibitory Concentration 50; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D008970:Molecular Weight; D029905:Pachyrhizus / Q000235:genetics* / Q000378:metabolism; D010843:Pichia / Q000235:genetics; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012639:Seeds / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence
D000935:Antifungal Agents; D018076:DNA, Complementary; D023082:Defensins; D010940:Plant Proteins; D011994:Recombinant Proteins