Molecular, biochemical and immunological analyses of canine pancreatic DNase I.
The DNase I from canine pancreas was purified 260-fold to electrophoretic homogeneity with a 35% yield using three-step column chromatography. The activity of the purified enzyme was completely inhibited by 20 mM EDTA, an antibody specific to the purified enzyme and G-actin. A 1,373-bp cDNA encoding canine DNase I was constructed from the total canine pancreatic RNA using a rapid amplification of cDNA ends method, followed by sequencing. The mature canine DNase I protein was found to consist of 262 amino acids. A survey of DNase I in 13 different canine tissues revealed the highest levels of both DNase I enzyme activity and gene expression in the pancreas; therefore, the canine DNase I is of the pancreatic type. Phylogenetic and sequence identity analyses, studies of immunological properties and the tissue-distribution patterns of DNase I indicated that the canine enzyme is more closely related to the human DNase I than to other mammalian DNases I. Therefore, canine DNase I is found to be one of the best substitutes in studies of human DNase I.
Kaneko
Yasushi
Y
Department of Legal Medicine and Molecular Genetics, Gunma University, Graduate School of Medicine, Maebashi, Gunma 371-8511.
Takeshita
Haruo
H
Mogi
Kouichi
K
Nakajima
Tamiko
T
Yasuda
Toshihiro
T
Itoi
Masako
M
Kuwano
Hiroyuki
H
Kishi
Koichiro
K
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D003850:Deoxyribonuclease I / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D004285:Dogs; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D018076:DNA, Complementary; D003850:Deoxyribonuclease I