ArticleProtein expression and purification2010
Engineering BspQI nicking enzymes and application of N.BspQI in DNA labeling and production of single-strand DNA.
Article in Protein expression and purification, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed, 34 citations in OpenAlex.
- Cryo-EM structures of DNA-free and DNA-bound BsaXI: architecture of a Type IIB restriction-modification enzyme.Nucleic acids research · 2025Article
- Strand-selective base editing of human mitochondrial DNA using mitoBEs.Nature biotechnology · 2024Article
- Types and Applications of Nicking Enzyme-Combined Isothermal Amplification.International journal of molecular sciences · 2022Review
- Engineering Infrequent DNA Nicking Endonuclease by Fusion of aFrontiers in microbiology · 2021Article
- A micropatterned substrate for on-surface enzymatic labelling of linearized long DNA molecules.Scientific reports · 2019Article
- Article
- Microscale Objects via Restructuring of Large, Double-Stranded DNA Molecules.ACS applied materials & interfaces · 2018Article
- Helraiser intermediates provide insight into the mechanism of eukaryotic replicative transposition.Nature communications · 2018Article
- Guiding the folding pathway of DNA origami.Nature · 2015Article
- Article
- Engineering nicking enzymes that preferentially nick 5-methylcytosine-modified DNA.Nucleic acids research · 2014Article
- Natural zinc ribbon HNH endonucleases and engineered zinc finger nicking endonuclease.Nucleic acids research · 2013Article
- Vectors for ligation-independent construction of lacZ gene fusions and cloning of PCR products using a nicking endonuclease.Plasmid · 2011Article
- Labeling DNA for single-molecule experiments: methods of labeling internal specific sequences on double-stranded DNA.Nanoscale · 2011Review
- Linear nicking endonuclease-mediated strand-displacement DNA amplification.Analytical biochemistry · 2011Article
- Degenerate sequence recognition by the monomeric restriction enzyme: single mutation converts BcnI into a strand-specific nicking endonuclease.Nucleic acids research · 2011Article
- Efficient preparation of internally modified single-molecule constructs using nicking enzymes.Nucleic acids research · 2011Article
- Natural and engineered nicking endonucleases--from cleavage mechanism to engineering of strand-specificity.Nucleic acids research · 2011Review
- Article
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Authors and funding
12 authors at 3 institutions in 2 countries.
Funding
Abstract
BspQI is a thermostable Type IIS restriction endonuclease (REase) with the recognition sequence 5'GCTCTTC N1/N4 3'. Here we report the cloning and expression of the bspQIR gene for the BspQI restriction enzyme in Escherichia coli. Alanine scanning of the BspQI charged residues identified a number of DNA nicking variants. After sampling combinations of different amino acid substitutions, an Nt.BspQI triple mutant (E172A/E248A/E255K) was constructed with predominantly top-strand DNA nicking activity. Furthermore, a triple mutant of BspQI (Nb.BspQI, N235A/K331A/R428A) was engineered to create a bottom-strand nicking enzyme. In addition, we demonstrated the application of Nt.BspQI in optical mapping of single DNA molecules. Nt or Nb.BspQI-nicked dsDNA can be further digested by E. coli exonuclease III to create ssDNA for downstream applications. BspQI contains two potential catalytic sites: a top-strand catalytic site (Ct) with a D-H-N-K motif found in the HNH endonuclease family and a bottom-strand catalytic site (Cb) with three scattered Glu residues. BlastP analysis of proteins in GenBank indicated a putative restriction enzyme with significant amino acid sequence identity to BspQI from the sequenced bacterial genome Croceibacter atlanticus HTCC2559. This restriction gene was amplified by PCR and cloned into a T7 expression vector. Restriction mapping and run-off DNA sequencing of digested products from the partially purified enzyme indicated that it is an EarI isoschizomer with 6-bp recognition, which we named CatHI (CTCTTC N1/N4).
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.