Characterization and biocontrol ability of fusion chitinase in Escherichia coli carrying chitinase cDNA from Trichothecium roseum.
The antifungal mechanism of mycoparasitic fungi involves fungal cell wall degrading enzymes such as chitinases. Trichothecium roseum is an important mycoparasitic fungus with significant antifungal ability, but studies on chitinases of T. roseum were poor. Here, we report a novel chitinase cDNA isolated from T. roseum by PCR amplification based on conserved chitinase sequences. Southern blot analysis suggested that a single copy of the gene exists in the genome of T. roseum. The deduced open reading frame of 1,143 nucleotides encodes a protein of 380 amino acids with a calculated molecular weight of 41.6 kDa. The fusion chitinase expressed in Escherichia coli has been purified by single-step chromatography. It has a pI of pH 5.4 and expresses a thermal stability, but is insensitive to pH in a broad pH range. According to expectation, E. coli efficiently yielded a high amount of active chitinase. Remarkably, the fusion chitinase offered high antifungal activity.
Pan
Hongyu
H
College of Plant Protection, Nanjing Agricultural University, Nanjing/Jiangsu 210095, China.
Wei
Yi
Y
Xin
Furong
F
Zhou
Mingguo
M
Zhang
Shihong
S
D000595:Amino Acid Sequence; D001483:Base Sequence; D002688:Chitinases / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018076:DNA, Complementary; D004271:DNA, Fungal / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D003368:Gossypium / Q000382:microbiology; D006999:Hypocreales / Q000201:enzymology / Q000235:genetics*; D007700:Kinetics; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018076:DNA, Complementary; D004271:DNA, Fungal; D002688:Chitinases