title: Isolation and characterization of cytoplasmic poly(A) polymerase from cryptobiotic gastrulae of Artemia salina.
abstract: Poly(A) polymerase has been purified to near homogeneity from the cytoplasm of Artemia salina cryptobiotic gastrulae by ion-exchange chromatography on DEAE-cellulose, DEAE-Sepharose CL-6B and phosphocellulose P11, gel filtration on CL-Sepharose 6B, affinity chromatography on poly(A)-Sepharose 4B and ATP-agarose. The enzyme is fully dependent on exogeneous oligo(riboadenylic acid) and is free of any nuclease or other enzyme activities. In standard assay conditions the enzyme preparation has a specific activity of 5.6 mumol AMP . h-1 . (mg protein)-1. Sodium dodecyl sulphate/polyacrylamide gel electrophoresis reveals the presence of only two proteins with Mr 94 000 and 70 000. The Mr-70 000 protein has been identified as poly(A) polymerase. The enzyme is exclusively activated by Mn2+. Addition of Ca2+, Mg2+, Zn2+, NH4+, K+ or Na+ inhibits the enzymatic reaction. The activity is specific for ATP and competitive inhibition is observed in the presence of other ribonucleoside 5'-triphosphates. AMP incorporation is time-dependent and is increased non-linearly with protein and primer concentration.
authors.forename: E
authors.initials: E
authors.lastname: Roggen
authors.forename: H
authors.initials: H
authors.lastname: Slegers
mesh_terms: D000818:Animals; D001156:Artemia / Q000201:enzymology*; D055598:Chemical Phenomena; D002621:Chemistry; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003593:Cytoplasm / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D009713:Nucleotidyltransferases / Q000302:isolation & purification*; D011062:Polynucleotide Adenylyltransferase / Q000302:isolation & purification* / Q000378:metabolism; D013347:Subcellular Fractions / Q000201:enzymology
chemical_list: D009713:Nucleotidyltransferases; D011062:Polynucleotide Adenylyltransferase