Mung bean nuclease
Mung bean nuclease is a nuclease derived from sprouts of the mung bean Vigna radiata that removes nucleotides in a step-wise manner from single-stranded DNA molecules (ssDNA) and is used in biotechnological applications to remove such ssDNA from a mixture also containing double-stranded DNA (dsDNA). This enzyme is useful for transcript mapping, removal of single-stranded regions in DNA hybrids or single-stranded overhangs produced by restriction enzymes, etc.
The enzyme degrades single-stranded DNA or RNA to nucleoside 5’-monophosphates, but does not digest double-stranded DNA, double-stranded RNA, or DNA / RNA hybrids. Mung Bean Nuclease catalyzes the specific degradation of single-stranded DNA or RNA, and produces mono and oligonucleotides carrying a 5′-P terminus.
Mung bean nuclease has a theoretical molecular weight of 39 kDa. Mung bean nuclease has a stringent single-stranded specificity for DNA or RNA and produces 5’-phosphoryl oligo- and mononucleotides.
Requirements
Mung bean nuclease requires Zn2+. The addition of EDTA or SDS causes irreversible inactivation. Mung bean nuclease is not active at pH below 4.6, nor at low salt concentration.
Description
A single-strand specific DNA and RNA endonuclease which will degrade single-stranded extensions from the ends of DNA and RNA molecules, leaving blunt, ligatable ends.
Because Mung Bean Nuclease has higher specificity for ssDNA and RNA than S1 Nuclease, it is the enzyme of choice for most applications requiring a single-strand-specific nuclease.
Unlike S1 Nuclease, Mung Bean Nuclease will not cleave the intact strand of nicked duplex DNA.
Its ability to recognise double-stranded nucleic acids depends on the base sequence.
It tends to cleave at ApN and at T(U) pN. It completely degrades ApA, but does not degrade G and C. Unlike S1 Nuclease, it does not cleave the strand opposite to that which has been nicked.
Mung Bean Nuclease catalyzes the specific degradation of single-stranded DNA or RNA, and produces mono- and oligonucleotides carrying a 5′-P terminus.
More than 1000- fold amount of enzyme can degrade oligomer into all mononucleotides.
An excess of the enzyme is required to degrade double-stranded DNA or RNA and DNA-RNA hybrids, and in this case, AT-rich regions are selectively degraded.
This enzyme work well at A↓pN, T ↓pN sites, and especially A↓pN sites are 100% degraded.
However, it is difficult to degrade C↓pC, C↓pG site.
Mung bean exonuclease is a nuclease derived from mung beans that removes nucleotides in a step-wise manner from single stranded DNA molecules and is used to remove such ssDNA from a mixture also containing double stranded DNA (dsDNA).
Unit Definition:
One unit of Mung Bean Nuclease converts 1 µg of heat-denatured calf thymus DNA into an acid-soluble form in 1 minute at 37 °C under standard assay conditions.
Applications in biotechnology and biochemical research
- Removal of hairpin loops during cDNA synthesis.
- High-resolution mapping of the termini and exon structures of RNA transcripts (commonly termed Berk-Sharp or S1 Mapping) using either internal-labelled or end-labelled probes.
- Restriction-site modification or removal by digestion of single-stranded protruding ends.
- Cleavage of single-basepair mismatches, as a replacement for CEL 1 Nuclease in TILLING.
- Unidirectional deletion of large DNA (in combination with Exonuclease III) to generate ordered deletions for sequencing.
- Removal of 3´ and 5´ extensions from DNA or RNA termini.
- Transcriptional mapping.
- Cleavage of hairpin loops.
- Excision of gene coding sequences from genomic DNA.