Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.3 (RNase T1)
 1,228 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A genomic DNA encoding ribonuclease (RNase) T1 from Aspergillus oryzae was cloned using a synthetic oligonucleotide probe. The cloned gene (designated rntA) encoded functional RNase T1, since an A. oryzae transformant with multiple copies of the rntA gene showed higher RNase T1 activity (over 200 times) than a transformant with a vector. A cDNA was cloned by reverse transcription polymerase chain reaction (RT-PCR) with primers corresponding to the 5' terminus and 3' terminus of the reading frame of the rntA gene. Nucleotide sequencing analysis of both DNAs found that RNase T1 had a prepro-sequence consisting of 26 amino acids and the rntA gene had only one intron (114 bp) in the region encoding the signal sequence. The A. oryzae transformant with cDNA controlled by the amyB promoter also showed higher activity (over 300 times), indicating that the cloned cDNA encoded functional RNase T1. On the other hand, the Saccharomyces cerevisiae transformant with cDNA controlled by the GAL1 promoter could not grow on a medium containing galactose. These results suggests that A. oryzae may have a protection mechanism from RNase T1.
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PMID:Cloning and nucleotide sequence of the ribonuclease T1 gene (rntA) from Aspergillus oryzae and its expression in Saccharomyces cerevisiae and Aspergillus oryzae. 853 78

Even though most fungal hydrolytic enzymes have been successfully secreted in S. cerevisiae cells by expression of corresponding cDNA, overexpression of A. oryzae RNase T1 causes severe growth inhibition in yeast. We observed that yeast strains carrying RNase T1 cDNA under control of the GAL1 promoter with a single-copy vector were able to grow on galactose medium while those with a multi-copy vector were not. It was found that overexpression of three mutated versions of RNase T1 with low enzymatic activity did not affect the growth. We also observed that expression of RNase T1 without a signal sequence severely inhibited growth of the transformant even on the single-copy plasmid. Subcellular fractionation showed that overexpressed myc-tagged RNase T1 was localized in the membrane fraction. In the yeast secretory pathway, while the mutants defective in translocation into the ER, ER-Golgi trafficking and vacuole formation had severe growth inhibition during expression of RNase T1 from the single-copy plasmid. These results suggest that a mislocalization of active RNase T1 in cytosol by overflow from the secretory apparatus has toxic effects on the host cells.
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PMID:Genetic analysis of growth inhibition of yeast cells caused by expression of Aspergillus oryzae RNase T1. 1112 88