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Ribonucleases from rat and bovine liver: purification, specificity and structural characterization
W Zhao1, Z Kote-Jarai, Y van Santen
1Biochemisch Laboratorium, Rijksuniversiteit Groningen, Netherlands.
Biochimica Et Biophysica Acta
|May 29, 1998
Summary
Researchers identified four pyrimidine-specific ribonucleases in rat liver, with three purified enzymes showing ribonuclease activity. This study details their characterization and sequence similarities to other ribonuclease superfamily members.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- The ribonuclease superfamily encompasses enzymes with diverse functions, including RNA degradation.
- Pyrimidine-specific ribonucleases play roles in cellular processes and have implications in various biological contexts.
- Understanding the diversity and function of these enzymes in different mammalian tissues is crucial.
Purpose of the Study:
- To identify and characterize pyrimidine-specific ribonucleases in rat liver.
- To elucidate the sequence and functional relationships of these rat liver ribonucleases with known members of the ribonuclease superfamily.
- To investigate the substrate specificity of liver-type ribonucleases.
Main Methods:
- Purification of ribonuclease enzymes from rat liver.
- Biochemical assays to determine ribonuclease activity using yeast RNA as a substrate.
- N-terminal sequencing and peptide analysis following chemical cleavage (acidic, CNBr).
- Amino acid sequence determination from a liver cDNA library.
- Comparative sequence analysis with known ribonucleases and angiogenins.
Main Results:
- Four pyrimidine-specific ribonucleases were detected in rat liver.
- Three purified ribonucleases (RL1, RL2, RL3) exhibited enzymatic activity.
- RL1 was identified as rat pancreatic ribonuclease (ribonuclease 1).
- RL2 showed high sequence similarity to neurotoxin-type ribonucleases.
- Rat liver-type ribonuclease (ribonuclease 4) sequence was determined, differing from other mammalian counterparts.
- A peptide from RL3 matched the liver-type ribonuclease sequence.
- A contaminant in RL3 fraction showed similarity to angiogenins.
- Liver-type ribonucleases (bovine, porcine, rat) demonstrated a poly(U) over poly(C) preference.
Conclusions:
- Rat liver expresses multiple pyrimidine-specific ribonucleases, including known and novel forms.
- RL2 and RL3 represent distinct members of the ribonuclease superfamily with unique sequence characteristics.
- The determined sequence of rat liver-type ribonuclease (ribonuclease 4) expands the understanding of mammalian liver ribonucleases.
- The observed poly(U) preference is a distinguishing feature of mammalian liver-type ribonucleases.