Chitinase from Paracoccidioides brasiliensis: molecular cloning, structural, phylogenetic, expression and activity analysis.
A full-length cDNA encoding a chitinase (Pbcts1) was cloned by screening a cDNA library from the yeast cells of Paracoccidioides brasiliensis. The cDNA consists of 1888 bp and encodes an ORF of 1218 bp corresponding to a protein of 45 kDa with 406 amino acid residues. The deduced PbCTS1 is composed of two signature family 18 catalytic domains and seems to belong to fungal/bacterial class. Phylogenetic analysis of PbCTS1 and other chitinases suggests the existence of paralogs of several chitinases to be grouped based on specialized functions, which may reflect the multiple and diverse roles played by fungi chitinases. Glycosyl hydrolase activity assays demonstrated that P. brasiliensis is able to produce and secrete these enzymes mainly during transition from yeast to mycelium. The fungus should be able to use chitin as a carbon source. The presence of an endocytic signal in the deduced protein suggests that it could be secreted by a vesicular nonclassical export pathway. The Pbcts1 expression in mycelium, yeast, during differentiation from mycelium to yeast and in yeast cells obtained from infected mice suggests the relevance of this molecule in P. brasiliensis electing PbCTS1 as an attractive drug target.
Laboratório de Biologia Molecular, Instituto de Ciências Biológicas, Universidade Federal de Goiás, Goiânia-Goiás, Brazil.
Sheyla M R C
SM
Bonfim
Aline H S
AH
Cruz
Rosália S A
RS
Jesuino
Cirano J
CJ
Ulhoa
Eugênia E W I
EE
Molinari-Madlum
Célia M A
CM
Soares
Maristela
M
Pereira
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002688:Chitinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004271:DNA, Fungal / Q000032:analysis; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D025282:Mycelium / Q000201:enzymology; D010228:Paracoccidioides / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity; D010229:Paracoccidioidomycosis / Q000382:microbiology; D010802:Phylogeny; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D004271:DNA, Fungal; D005656:Fungal Proteins; D002688:Chitinases