Identification and molecular characterization of a chitinase from the hard tick Haemaphysalis longicornis.
A cDNA encoding tick chitinase was cloned from a cDNA library of mRNA from Haemaphysalis longicornis eggs and designated as CHT1 cDNA. The CHT1 cDNA contains an open reading frame of 2790 bp that codes for 930 amino acid residues with a coding capacity of 104 kDa. The deduced amino acid sequence shows a 31% amino acid homology to Aedes aegypti chitinase and a multidomain structure containing one chitin binding peritrophin A domain and two glycosyl hydrolase family 18 chitin binding domains. The endogenous chitinase of H. longicornis was identified by a two-dimensional immunoblot analysis with mouse anti-rCHT1 serum and shown to have a molecular mass of 108 kDa with a pI of 5.0. A recombinant baculovirus AcMNPV.CHT1-expressed rCHT1 is glycosylated and able to degrade chitin. Chitin degradation was ablated by allosamidin in a dose-dependent manner. The optimal temperature and pH for activity of the purified chitinase were 45 degrees C and pH 5-7. The CHT1 cDNA has an ELR motif for chemokine-mediated angiogenesis and appears to be a chitinase of the chemokine family. Localization analysis using mouse anti-rCHT1 serum revealed that native chitinase is highly expressed in the epidermis and midgut of the tick. AcMNPV.CHT1 topically applied to H. longicornis ticks exhibited replication. This is the first report of insect baculovirus infection of ticks. The importance of AcMNPV.CHT1 as a novel bio-acaricide for tick control is discussed.
You
Myungjo
M
Department of Basic Veterinary Science, the United Graduate School of Veterinary Sciences, Gifu University, Yanagito, Japan.
Xuan
Xuenan
X
Tsuji
Naotoshi
N
Kamio
Tsugihiko
T
Taylor
DeMar
D
Suzuki
Naoyoshi
N
Fujisaki
Kozo
K
D000595:Amino Acid Sequence; D000818:Animals; D001096:Arachnid Vectors; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D002688:Chitinases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D018411:Spodoptera; D013987:Ticks / Q000201:enzymology*
D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; D002688:Chitinases