| Title |
A new family of CRISPR-type V nucleases with C-rich PAM recognition / |
| Authors |
Urbaitis, Tomas ; Gasiūnas, Giedrius ; Young, Joshua K ; Hou, Zhenglin ; Paulraj, Sushmitha ; Godliauskaitė, Eglė ; Juškevičienė, Mantvyda M ; Štitilytė, Miglė ; Jasnauskaitė, Monika ; Mabuchi, Megumu ; Robb, G Brett ; Šikšnys, Virginijus |
| DOI |
10.15252/embr.202255481 |
| Full Text |
|
| Is Part of |
EMBO reports.. Hoboken : Wiley. 2022, vol. 23, iss. 12, art. no. e55481, p. [1-15].. ISSN 1469-221X. eISSN 1469-3178 |
| Keywords [eng] |
CRISPR-Cas ; PAM ; RNA-guided nuclease ; genome editing ; nucleic acid detection |
| Abstract [eng] |
Most CRISPR-type V nucleases are stimulated to cleave double-stranded (ds) DNA targets by a T-rich PAM, which restricts their targeting range. Here, we identify and characterize a new family of type V RNA-guided nuclease, Cas12l, that exclusively recognizes a C-rich (5'-CCY-3') PAM. The organization of genes within its CRISPR locus is similar to type II-B CRISPR-Cas9 systems, but both sequence analysis and functional studies establish it as a new family of type V effector. Biochemical experiments show that Cas12l nucleases function optimally between 37 and 52°C, depending on the ortholog, and preferentially cut supercoiled DNA. Like other type V nucleases, it exhibits collateral nonspecific ssDNA and ssRNA cleavage activity that is triggered by ssDNA or dsDNA target recognition. Finally, we show that one family member, Asp2Cas12l, functions in a heterologous cellular environment, altogether, suggesting that this new group of CRISPR-associated nucleases may be harnessed as genome editing reagents. |
| Published |
Hoboken : Wiley |
| Type |
Journal article |
| Language |
English |
| Publication date |
2022 |
| CC license |
|
