Intestinal DNases of 36 and 38.5kDa from the parasitic nematode Haemonchus contortus have non-classic DNase characteristics and produce DNA fragments with 3'-hydroxyls.
Treatment with the anthelmintic fenbendazole induces fragmentation of genomic DNA in intestinal cells of Haemonchus contortus. This effect is characterized by DNA fragments with 3'-hydroxyls (OH). Investigation into DNases responsible identified intestinal DNase activities that produce DNA fragments with 3'-OH. However, this interpretation was complicated by a mixture of activities in the intestinal fractions evaluated. In addition, intestinal activities displayed non-classic characteristics. Here it is shown that heparin sulfate (HS) fractionation enriched for intestinal DNases that produce 3'-OH. The 2.0M NaCl fraction of HS contained DNase activity that produced 3'-OH with minimal contamination by activity that produced 3'-phosphates (P). 3'-OH were produced under acidic (pH 5.0) or neutral (pH 7.0) conditions by DNases in this fraction. These DNases were sensitive to EDTA under each condition. Furthermore, EDTA-sensitive DNase activity in this fraction digested H. contortus intestinal cell nuclear DNA in histological sections, producing 3'-OH under acidic and neutral conditions. DNases at 36 and 38.5kDa in this fraction each produced 3'-OH at pH 5.0 when gel eluted, and each activity was sensitive to EDTA. Hence, the 36 and 38.5kDa DNases in the 2.0M NaCl HS intestinal fraction have characteristics expected for candidate DNases that mediate DF in H. contortus intestinal cell nuclei induced by fenbendazole. DNase activity that produces 3'-OH under acidic condition with sensitivity to EDTA is unconventional for classic acidic or neutral DNases and is a unique finding for nematodes. Excretory/secretory products from the worm and whole worm lysates were also explored as sources to fractionate intestinal DNases identified. HS fractionation of those worm samples did not clearly resolve the intestinal DNases of interest, although DNases with distinct characteristics were identified in each source.
Kwak
Dongmi
D
Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, USA.
Jasmer
Douglas P
DP
D000818:Animals; D053938:DNA Fragmentation; D018509:DNA, Helminth / Q000737:chemistry* / Q000378:metabolism; D003851:Deoxyribonucleases / Q000737:chemistry / Q000378:metabolism*; D006189:Haemonchus / Q000201:enzymology* / Q000235:genetics; D017665:Hydroxyl Radical / Q000737:chemistry; D007422:Intestines / Q000201:enzymology; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism
D018509:DNA, Helminth; D017665:Hydroxyl Radical; D003851:Deoxyribonucleases; D010744:Phosphoric Monoester Hydrolases