Ribonuclease V1 Explained

Ribonuclease V1 (RNase V1) is a ribonuclease enzyme found in the venom of the Caspian cobra (Naja oxiana).^[1] It cleaves double-stranded RNA in a non-sequence-specific manner, usually requiring a substrate of at least six stacked nucleotides.^[2] Like many ribonucleases, the enzyme requires the presence of magnesium ions for activity.^[3]

Laboratory use

Purified RNase V1 is a commonly used reagent in molecular biology experiments. In conjunction with other ribonucleases that cleave single-stranded RNA after specific nucleotides or sequences – such as RNase T1 and RNase I – it can be used to map internal interactions in large RNA molecules with complex secondary structure or to perform footprinting experiments on macromolecular complexes containing RNA.

RNase V1 is the only commonly used laboratory RNase that provides positive evidence for the presence of double-stranded helical conformations in target RNA.^[4] Because RNase V1 has some activity against RNA that is base-paired but single-stranded,^[5] dual susceptibility to both RNase V1 and RNase I at a single site in a target RNA molecule provides evidence of this relatively unusual conformation found in RNA loops.^[6]

Structural discoveries

RNase V1 played a particularly important role in the elucidation of the distinctive stem-loop structure of transfer RNA.^[7] It has also been extensively used to study the highly structured RNA genomes of retroviruses, such as hepatitis C,^[8] dengue virus,^[9] and HIV.^[10] Together with S1 nuclease, which specifically cleaves single-stranded RNA, it can be used to profile the secondary structure propensities of messenger RNA molecules, a procedure that can be applied to whole transcriptomes when paired with deep sequencing.^{[11] [12]}

Notes and References

1. Favorova OO, Fasiolo F, Keith G, Vassilenko SK, Ebel JP . Partial digestion of tRNA--aminoacyl-tRNA synthetase complexes with cobra venom ribonuclease . Biochemistry . 20 . 4 . 1006–11 . February 1981 . 7011369 . 10.1021/bi00507a055 .
2. Book: Ying. Shao Yao. MicroRNA Protocols. Humana Press. 23. registration. 9781597451239. 2006-01-01.
3. Nilsen TW . RNA structure determination using nuclease digestion . Cold Spring Harbor Protocols . 2013 . 4 . 379–82 . April 2013 . 23547152 . 10.1101/pdb.prot072330 . free .
4. Book: Melodie. Duval. Cedric. Romilly. Anne-Catherine. Helfer. Olivier. Fuchsbauer. Pascale. Romby. Stefano. Marzi . Klostermeier. Dagmar. Hammann. Christian. RNA Structure and Folding: Biophysical Techniques and Prediction Methods. 2013. Walter de Gruyter. 32. 9783110284959.
5. Lowman HB, Draper DE . On the recognition of helical RNA by cobra venom V1 nuclease . The Journal of Biological Chemistry . 261 . 12 . 5396–403 . April 1986 . 10.1016/S0021-9258(19)57229-5 . 2420800 . free .
6. Chaulk SG, Xu Z, Glover MJ, Fahlman RP . MicroRNA miR-92a-1 biogenesis and mRNA targeting is modulated by a tertiary contact within the miR-17~92 microRNA cluster . Nucleic Acids Research . 42 . 8 . 5234–44 . April 2014 . 24520115 . 4005684 . 10.1093/nar/gku133 .
7. Lockard RE, Kumar A . Mapping tRNA structure in solution using double-strand-specific ribonuclease V1 from cobra venom . Nucleic Acids Research . 9 . 19 . 5125–40 . October 1981 . 7031604 . 327503 . 10.1093/nar/9.19.5125 .
8. Blight KJ, Rice CM . Secondary structure determination of the conserved 98-base sequence at the 3' terminus of hepatitis C virus genome RNA . Journal of Virology . 71 . 10 . 7345–52 . October 1997 . 10.1128/JVI.71.10.7345-7352.1997 . 9311812 . 192079 .
9. Polacek C, Foley JE, Harris E . Conformational changes in the solution structure of the dengue virus 5' end in the presence and absence of the 3' untranslated region . Journal of Virology . 83 . 2 . 1161–6 . January 2009 . 19004957 . 2612390 . 10.1128/JVI.01362-08 .
10. Harrison GP, Lever AM . The human immunodeficiency virus type 1 packaging signal and major splice donor region have a conserved stable secondary structure . Journal of Virology . 66 . 7 . 4144–53 . July 1992 . 10.1128/JVI.66.7.4144-4153.1992 . 1602537 . 241217 . .
11. Kertesz M, Wan Y, Mazor E, Rinn JL, Nutter RC, Chang HY, Segal E. Howard Y. Chang . Genome-wide measurement of RNA secondary structure in yeast . Nature . 467 . 7311 . 103–7 . September 2010 . 20811459 . 3847670 . 10.1038/nature09322 . 2010Natur.467..103K .
12. Book: Silverman . Ian M. . Berkowitz . Nathan D. . Gosai . Sager J. . Gregory . Brian D. . Yeo . Gene W. . RNA Processing . 2016 . Springer . 978-3-319-29071-3 . 29–59 . Genome-Wide Approaches for RNA Structure Probing.