Team:Vilnius-Lithuania/Basic Part
Basic part
In this page we describe our favorite basic part, which is I-F type CRISPR-Cas Cascade complex. It can be found in Parts Registry - BBa_K1773005.
I-F type CRISPR-Cas Cascade complex
I-F type CRISPR-Cas Cascade complex, which has a mutated EcoRI restriction site to make it biobrick compatible. This protein complex needs to be coexpressed with a crRNA region to make a functional Cascade ribonucleoprotein complex. Together with Cas3 (BBa_K1773003) degrades DNA.
Usage and Biology
Cascade (CRISPR Associated Complex for Antiviral Defence) complex is an essential component of I-type CRISPR-Cas systems. This ribonucleoprotein complex is coded by operon of 4 genes: Csy1, Csy2, Csy3 and Cas6f, with stoichiometry of 1:1:6:1 respectively. Also it has a crRNA molecule, which acts like a guide, that brings the whole complex to the target dsDNA sequence. Each Cascade complex has a unique Protospacer adjacent motive (PAM) recognition site. The recognisable PAM sequence of this specific I-F Cascade complex from Aggregatibacter actinomycetemcomitans recognises CC nucleotides directly upstream of the protospacer (the target sequence). In other words, a target sequence needs to have a CC pair on the non-complementary strand of DNA -2 -1 positions of the protospacer. When Cascade complex finds and recognises the DNA target, then Cas3 (BBa_K1773003) is recruited for DNA degradation.