title: A novel ribonuclease with antiproliferative activity from fresh fruiting bodies of the edible mushroom Hypsizigus marmoreus.
abstract: An 18-kDa ribonuclease (RNase) with a novel N-terminal sequence was purified from fresh fruiting bodies of the mushroom Hypsizigus marmoreus. The purification protocol comprised ion exchange chromatography on DEAE cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on CM-cellulose and Q-Sepharose and gel filtration by fast protein liquid chromatography on Superdex 75. The starting buffer was 10 mM Tris-HCl buffer (pH 7.2), 10 mM Tris-HCl buffer (pH 7.2), 10 mM NH(4)OAc buffer (pH 5), 10 mM NH(4)HCO(3) buffer (pH 9.4) and 200 mM NH(4)HCO(3) (pH 8.5), respectively. Absorbed proteins were desorbed using NaCl added to the starting buffer. A 42-fold purification of the enzyme was achieved. The RNase was unadsorbed on DEAE cellulose, Affi-gel blue gel and CM-cellulose but adsorbed on Q-Sepharose. It exhibited maximal RNase activity at pH 5 and 70 degrees C. Some RNase activity was detectable at 100 degrees C. It demonstrated the highest ribonucleolytic activity (196 U/mg) toward poly C, the next highest activity (126 U/mg) toward poly A, and much weaker activity toward poly U (48 U/mg) and poly G (41 U/mg). The RNase inhibited [(3)H-methyl]-thymidine uptake by leukemia L1210 cells with an IC(50) of 60 microM.
authors.affiliation: State Key Laboratory for Agrobiotechnology and Department of Microbiology, China Agricultural University, Beijing, China.
authors.forename: G P
authors.initials: GP
authors.lastname: Guan
authors.forename: H X
authors.initials: HX
authors.lastname: Wang
authors.forename: T B
authors.initials: TB
authors.lastname: Ng
mesh_terms: D000363:Agaricales / Q000201:enzymology*; D000595:Amino Acid Sequence; D049109:Cell Proliferation / Q000187:drug effects*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D012260:Ribonucleases / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*
chemical_list: D012260:Ribonucleases