Molecular cloning, characterization, and expression of a chitinase from the entomopathogenic fungus Paecilomyces javanicus.
Paecilomyces javanicus is an entomopathogenic fungus of coleopteran and lepidopteran insects. Here we report on cloning, characterization, and expression patterns of a chitinase from P. javanicus. A strong chitinase activity was detected in P. javanicus cultures added to chitin. The full-length cDNA, designated PjChi-1, was cloned from mycelia by using both degenerate primer/reverse transcription polymerase chain reaction (RT-PCR) amplification and 5'-/3'-RACE extension. The 1.18-kb cDNA gene contains a 1035-bp open reading frame and encodes a 345-amino acid polypeptide with a deduced molecular mass of 37 kDa. A conserved motif for chitinase activity -F82DGIDIDWE90- was present in deduced amino acid sequence. Both RT-PCR and Northern analysis revealed that the expression of the PjChi gene was constitutive at low level, but enhanced to high level when chitin was the substrate. Fungal inhibitory assay showed that PjChi-1 inhibited the growth of phytopathogenic fungi such as Sclerotium rolfsii, Colletotrichum gloeosporioides, Aspergillus nidulans, and Rhizoctonia solani.
Chen
Chien-Cheng
CC
Institute of Plant Biology, College of Life Science, National Taiwan University, 106, Taipei, Taiwan.
Kumar
H G Ashok
HG
Kumar
Senthil
S
Tzean
Shean-Shong
SS
Yeh
Kai-Wun
KW
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000898:Antibiosis; D015152:Blotting, Northern; D002686:Chitin / Q000378:metabolism; D002688:Chitinases / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D015870:Gene Expression; D015966:Gene Expression Regulation, Fungal; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D010143:Paecilomyces / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic
D018076:DNA, Complementary; D004271:DNA, Fungal; D002686:Chitin; D002688:Chitinases