Evolutionary relationship between defensins in the Poaceae family strengthened by the characterization of new sugarcane defensins.
Plant defensins are small (45-54 amino acids), highly basic, cysteine-rich peptides structurally related to defensins of other organisms, including insects and mammals. Small putative proteins (MW < 10 kDa) containing eight cysteines were screened based on the sugarcane expressed sequence tag (EST) database. We selected ORFs that exhibited 25-100% similarity in primary sequence with other defensins in the NCBI database and that contained eight cysteines. This similarity is sufficient for folding prediction, but not enough for biological activity inference. Six putative defensins (Sd1-6) were selected, and activity assays showed that recombinant Sd1, Sd3 and Sd5 are active against fungi, but not against bacteria. Structural characterization, based on circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy showed that the structures of these Sds were compatible with alpha/beta proteins, a feature expected for plant defensins. Phylogenetic analysis revealed that sugarcane defensins could clearly be grouped within defensins from Poaceae family and Andropogoneae tribe. Our work demonstrates that defensins show strong conservation in the Poaceae family and may indicate that the same conservation occurs in other families. We suggest that evolutionary relationships within plant families can be used as a procedure to predict and annotate new defensins in genomes and group them in evolutionary classes to help in the investigation of their biological function.
De-Paula
V S
VS
Centro Nacional de Ressonância Magnética Nuclear, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Av. Brigadeiro Trompowski, s/n, CCS Bloco E sala 10, Rio de Janeiro, RJ, Brazil.
Razzera
G
G
Medeiros
L
L
Miyamoto
C A
CA
Almeida
M S
MS
Kurtenbach
E
E
Almeida
F C L
FC
Valente
A P
AP
D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D017124:Conserved Sequence; D023082:Defensins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D019143:Evolution, Molecular; D005658:Fungi / Q000187:drug effects; D009682:Magnetic Resonance Spectroscopy; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010802:Phylogeny; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D006109:Poaceae / Q000235:genetics*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D031786:Saccharum; D016415:Sequence Alignment
D000935:Antifungal Agents; D023082:Defensins; D010940:Plant Proteins; D011994:Recombinant Proteins