A low-molecular mass ribonuclease from the brown oyster mushroom.
A ribonuclease, with a molecular mass of 9 kDa and an N-terminal sequence resembling the sequence of a fragment of tRNA/rRNA cytosine-C5-methylase and a fragment of a alanyl-tRNA synthetase, was isolated from fresh fruiting bodies of the brown oyster mushroom Pleurotus ostreatus. The ribonuclease was purified using a very simple protocol that comprised ion-exchange chromatography on carboxymethyl (CM)-cellulose and affinity chromatography on Affi-gel blue gel. Subsequent gel filtration by fast protein liquid chromatography on Superdex 75 and sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis revealed that the ribonuclease was purified after the first two chromatographic steps. The ribonuclease was adsorbed on CM-cellulose and Affi-gel blue gel. The ribonuclease exhibited the highest activity toward poly A, lower activity toward poly C, slight activity toward poly G, and indiscernible activity toward poly U. The enzyme was stimulated upon exposure to 1 microm Mg2+ and 10 microm Zn2+, but was inhibited by the following ions at 10 mm: Ca2+, Mg2+, Zn2+, Cu2+, Fe2+, Mn2+, and Fe3+. The ribonuclease required a pH of 8.0 and a temperature of 50-70 degrees C to express maximal activity. It had a Km of 60 microm toward yeast tRNA. It lacked mitogenic and HIV-1 reverse transcriptase inhibiting activities, but exerted antiproliferative activity toward leukemia L1210 cells.
Xia
L
L
Department of Life Sciences, Shenzhen University, Shenzhen, China.
Chu
K T
KT
Ng
T B
TB
D000595:Amino Acid Sequence; D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D048690:Fruiting Bodies, Fungal / Q000201:enzymology*; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors; D006863:Hydrogen-Ion Concentration; D007939:Leukemia L1210; D051379:Mice; D008810:Mice, Inbred C57BL; D008934:Mitogens / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D020076:Pleurotus / Q000201:enzymology*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D012260:Ribonucleases / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013154:Spleen / Q000166:cytology / Q000187:drug effects
D008934:Mitogens; D018894:Reverse Transcriptase Inhibitors; D054303:HIV Reverse Transcriptase; C436527:Pleurotus tuber-regium ribonuclease; D012260:Ribonucleases