title: Multiple forms of DNA polymerase from the thermo-acidophilic eubacterium Bacillus acidocaldarius: purification, biochemical characterization and possible biological role.
abstract: Two DNA polymerase isoenzymes, called DpA and DpB on the basis of their elution order from DEAE cellulose, were purified to homogeneity from the thermo-acidophilic eubacterium Bacillus acidocaldarius. The enzymes are weakly acidophilic proteins constituted by a single subunit of 117 and 103 kDa respectively. DpA and DpB differ in thermostability, in thermophilicity, in sensitivity to assay conditions and in resistance to sulphydryl-group blocking agents such as N-ethylmaleimide and p-hydroxymercuriobenzoate. They differ also in synthetic template-primer utilization, in the apparent Km for dNTPs and in processivity. In particular, DpA utilizes more effic iently synthetic templates-primers such as poly(dA).poly(dT), poly(dT). (rA)12-18 and poly(rA).(dT)12-18 and presents a greater tendency to accept dNTP analogues modified in the sugar or in the base ring, such as cytosine beta-d-arabinofuranoside 5'-triphosphate, 2',3'-dideoxyribonucleosides 5'-triphosphate, butylphenyl-dGTP and digoxigenin-conjugated dUTP. In addition, DpA presents an exonuclease activity that preferentially hydrolyses DNA in the 5'-3' direction, whereas DpB lacks this activity. The possible biological role of the enzymes is discussed.
authors.affiliation: Istituto di Biochimica delle Proteine ed Enzimologia del Consiglio Nazionale delle Ricerche, Via Toiano 6, 80072 Arco Felice, Naples, Italy.
authors.forename: M
authors.initials: M
authors.lastname: De Falco
authors.forename: P
authors.initials: P
authors.lastname: Grippo
authors.forename: M
authors.initials: M
authors.lastname: Rossi
authors.forename: P
authors.initials: P
authors.lastname: Orlando
mesh_terms: D001407:Bacillus / Q000201:enzymology*; D002852:Chromatography, Ion Exchange; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D007525:Isoelectric Focusing; D007700:Kinetics; D019384:Nucleic Acid Synthesis Inhibitors; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D019384:Nucleic Acid Synthesis Inhibitors; D004259:DNA-Directed DNA Polymerase