A novel tRNA precursor cleaving endoribonuclease from Zea mays.
In a search for tRNA-processing nucleases in Zea mays an activity was found which cleaves the precursor to Escherichia coli tyrosine tRNA in the loop of the extra arm within the mature tRNA sequence. The activity (named RNase Zma) was partially purified by ion exchange and gel filtration chromatography; the latter step enabled estimation of the molecular weight of the enzyme at 34,000. The optimal pH and Mg2+ concentration varied in an interdependent manner; 23 mM Mg2+ and pH 7.5 gave the best combination of cleavage by RNase Zma and inhibition of cleavage by (apparently) contaminating nuclease(s). Increasing concentrations of both sodium and ammonium chloride inhibited activity, in both cases by about 80% at 0.5 M. RNase Zma was also inhibited by mature tRNA (52% inhibition at 200 micrograms/ml tRNA). The activity was destroyed completely by heating at 100 degrees C for 5 min but was increasingly active over the temperature range of 23-42 degrees C; the latter experiments yielded an activation energy for the cleavage reaction of 6.2 kcal/mole. RNase Zma was also sensitive to protease digestion even though fractions which contain it consist primarily of RNA. The in vivo role of RNase Zma is unknown, but it does produce a product similar to a tRNA fragment which may be used to prime reverse transcription of copia elements in Drosophila.
Hayashi
D K
DK
Department of Chemical and Biological Sciences, Illinois Institute of Technology, IIT Center, Chicago 60616.
Stark
B C
BC
D000643:Ammonium Chloride / Q000494:pharmacology*; D000818:Animals; D001483:Base Sequence; D004330:Drosophila / Q000235:genetics; D004722:Endoribonucleases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012322:RNA Precursors / Q000378:metabolism*; D016337:RNA, Catalytic / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism*; D012358:RNA, Transfer, Met / Q000378:metabolism; D043262:Ribonuclease P; D012964:Sodium / Q000494:pharmacology; D003313:Zea mays / Q000201:enzymology*
D029968:Escherichia coli Proteins; D012322:RNA Precursors; D016337:RNA, Catalytic; D012358:RNA, Transfer, Met; D000643:Ammonium Chloride; D012343:RNA, Transfer; D012964:Sodium; D004722:Endoribonucleases; D043262:Ribonuclease P; C470865:ribonuclease P, E coli; D008274:Magnesium