6EKO

Crystal structure of Type IIP restriction endonuclease PfoI with cognate DNA

Summary for 6EKO

• Entry DOI: 10.2210/pdb6eko/pdb
• Related: 6EK1
• Descriptor: Restriction endonuclease PfoI, DNA (5'-D(*CP*GP*CP*TP*CP*CP*CP*GP*GP*AP*GP*CP*GP*T)-3'), CALCIUM ION, ... (4 entities in total)
• Functional Keywords: restriction endonuclease, pd-(d/e)xk nuclease, hydrolase
• Biological source: Pseudomonas fluorescens; More
• Total number of polymer chains: 4
• Total formula weight: 79851.21

Authors

Tamulaitiene, G.,Manakova, E.,Jovaisaite, V.,Grazulis, S.,Siksnys, V. (deposition date: 2017-09-26, release date: 2018-10-10, Last modification date: 2024-10-16)

Primary citation

Tamulaitiene, G.,Manakova, E.,Jovaisaite, V.,Tamulaitis, G.,Grazulis, S.,Bochtler, M.,Siksnys, V.
Unique mechanism of target recognition by PfoI restriction endonuclease of the CCGG-family.
Nucleic Acids Res., 47:997-1010, 2019

PubMed Abstract: Restriction endonucleases (REs) of the CCGG-family recognize a set of 4-8 bp target sequences that share a common CCGG or CCNGG core and possess PD…D/ExK nuclease fold. REs that interact with 5 bp sequence 5'-CCNGG flip the central N nucleotides and 'compress' the bound DNA to stack the inner base pairs to mimic the CCGG sequence. PfoI belongs to the CCGG-family and cleaves the 7 bp sequence 5'-T|CCNGGA ("|" designates cleavage position). We present here crystal structures of PfoI in free and DNA-bound forms that show unique active site arrangement and mechanism of sequence recognition. Structures and mutagenesis indicate that PfoI features a permuted E…ExD…K active site that differs from the consensus motif characteristic to other family members. Although PfoI also flips the central N nucleotides of the target sequence it does not 'compress' the bound DNA. Instead, PfoI induces a drastic change in DNA backbone conformation that shortens the distance between scissile phosphates to match that in the unperturbed CCGG sequence. Our data demonstrate the diversity and versatility of structural mechanisms employed by restriction enzymes for recognition of related DNA sequences.

PubMed: 30445642
DOI: 10.1093/nar/gky1137

Experimental method

X-RAY DIFFRACTION (2.284 Å)