title: Partial characterization of Acanthamoeba castellanii (T4 genotype) DNase activity.
abstract: The deoxyribonuclease (DNase) activities of Acanthamoeba castellanii belonging to the T4 genotype were investigated. Using zymographic assays, the DNase activities had approximate molecular masses of 25 and 35 kDa. A. castellanii DNases exhibited activity at wide-ranging temperature of up to 60 °C and at pH ranging from 4 to 9. The DNases activities were unaffected by proteinase-K treatment, divalent cations such as Ca(++), Cu(++), Mg(++), and Zn(++), or divalent cation chelating agent ethylenediaminetetraacetic acid (EDTA) or sodium dodecyl sulfate (SDS). The non-reliance on divalent cations and homology data suggests that A. castellanii DNases belong to the class of eukaryotic lysosomal DNase II but exhibit robust properties. The DNases activity in A. castellanii interfered with the genomic DNA extraction. Extraction methods involving EDTA, SDS, and proteinase-K resulted in low yield of genomic DNA. On the other hand, these methods resulted in high yield of genomic DNA from human cells suggesting the robust nature of A. castellanii DNases that are unaffected by reagents normally used in blocking eukaryotic DNases. In contrast, the use of chaotropic agent such as guanidine thiocyanate improved the yield of genomic DNA from A. castellanii cells significantly. Further purification and characterization of Acanthamoeba DNases is needed to study their non-classic distinct properties and to determine their role in the biology, cellular differentiation, cell cycle progression, and arrest of Acanthamoeba.
authors.affiliation: Department of Biological and Biomedical Sciences, Aga Khan University, Stadium Road, Karachi, Pakistan.
authors.forename: Junaid
authors.initials: J
authors.lastname: Iqbal
authors.forename: Shamvil
authors.initials: S
authors.lastname: Panjwani
authors.forename: Ruqaiyyah
authors.initials: R
authors.lastname: Siddiqui
authors.forename: Naveed Ahmed
authors.initials: NA
authors.lastname: Khan
mesh_terms: D048248:Acanthamoeba castellanii / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D000562:Amebiasis / Q000469:parasitology*; D001921:Brain / Q000098:blood supply / Q000166:cytology; D002478:Cells, Cultured; D016054:DNA, Protozoan / Q000235:genetics / Q000302:isolation & purification; D003851:Deoxyribonucleases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells; D005838:Genotype; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007634:Keratitis / Q000469:parasitology; D008018:Life Cycle Stages; D008970:Molecular Weight; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D013696:Temperature
chemical_list: D016054:DNA, Protozoan; D015800:Protozoan Proteins; D003851:Deoxyribonucleases