Molecular characterization of a peroxiredoxin from the hard tick Haemaphysalis longicornis.
Antioxidant enzymes in eukaryotes play an important role in protection against the oxygen radicals generated during aerobic metabolism. Here we report the cloning and characterization of a cDNA encoding the antioxidant enzyme peroxiredoxin from the hard tick Haemaphysalis longicornis (HlPrx). HlPrx is 939 bp long and contains a 101 bp non-translated sequence at the 5' end and a polyadenylation singnal followed by a poly(A) tail at the 3' end. HlPrx encodes a full-length protein with a predicted molecular mass of 26 kDa that possesses one cysteine residue at amino acid 49 that is conserved among Prx proteins of various species. GenBanktrade mark analysis showed that the deduced amino acid sequence had significant similarity to mammalian and plant Prxs at the amino acid level. A DNA-nicking assay revealed that Escherichia coli-expressed recombinant HlPrx (rHlPrx) inhibited oxidative-nicking of supercoiled plasmid DNA. Two-dimensional immunoblot analysis with mouse antirHlPrx serum showed reaction with a major constituent protein spot in extracts of adult ticks. In addition, immunoblot analysis showed that rHlPrx was immunoreacted with serum from rabbits repeatedly infested with H. longicornis. Localization analysis using mouse antirHlPrx serum revealed that native HlPrx was highly expressed in the salivary gland of the tick. Moreover, Northern blot analysis showed that the level of HlPrx transcripts was increased during blood sucking. The present results indicate that HlPrx may be an important detoxifying enzyme during the normal life span as well as during blood sucking in ticks.
Tsuji
N
N
Laboratory of Parasitic Diseases, National Institute of Animal Health, Ministry of Agriculture, Forestry and Fisheries, 3-1-1 Kannondai, Tsukuba, Ibaraki 305-0856 Japan. tsujin@niah.affrc.go.jp
Kamio
T
T
Isobe
T
T
Fujisaki
K
K
D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D010544:Peroxidases / Q000235:genetics* / Q000378:metabolism; D054464:Peroxiredoxins; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics; D013987:Ticks / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism
D000975:Antioxidants; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D010544:Peroxidases; D054464:Peroxiredoxins