title: Purification and characterization of tilapia (Oreochromis mossambicus) deoxyribonuclease I--primary structure and cDNA sequence.
abstract: DNase I of tilapia (Oreochromis mossambicus) was purified to homogeneity. Tilapia DNase I is most active at pH 8.5 with Mg2+ as activator. The Ca2+/Mg2+ pair has a synergistic effect on activation. The enzyme is readily inactivated by heating above 55 degrees C, but is not inactivated by trypsin or 2-mercaptoethanol under alkaline conditions, with or without CaCl2. Its isoelectric point is 6.0. The 258-amino-acid sequence of tilapia DNase I was derived from overlapping sequences of tryptic, chymotryptic and CNBr peptides. The purified enzyme has two variants differing by a single Lys-->Arg mutation at position 125. The polypeptide chain has one disulfide bridge and one carbohydrate side chain. By mass spectrometry, the purified enzyme shows many molecular mass forms differing by Lys/Arg substitution and sugar-chain length. The major form has a molecular mass of 30,914 Da. A 1061-bp nucleotide sequence for the cDNA of tilapia DNase I, obtained by gene cloning and DNA sequencing, contains an ORF coding for a putative 26-residue transmembrane peptide and the mature DNase I polypeptide.
authors.affiliation: Institute of Biochemistry, College of Medicine, National Taiwan University, Taipei, ROC.
authors.forename: Y M
authors.initials: YM
authors.lastname: Hsiao
authors.forename: H C
authors.initials: HC
authors.lastname: Ho
authors.forename: W Y
authors.initials: WY
authors.lastname: Wang
authors.forename: M F
authors.initials: MF
authors.lastname: Tam
authors.forename: T H
authors.initials: TH
authors.lastname: Liao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003850:Deoxyribonuclease I / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004795:Enzyme Stability; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016415:Sequence Alignment; D017421:Sequence Analysis; D013696:Temperature; D017210:Tilapia / Q000378:metabolism*
chemical_list: D003850:Deoxyribonuclease I