Folding, DNA recognition, and function of GIY-YIG endonucleases: crystal structures of R.Eco29kI.

Publication Type:

Journal Article

Source:

Structure (London, England : 1993), Volume 18, Issue 10, p.1321-31 (2010)

Keywords:

2010, Amino Acid Sequence, Base Sequence, Basic Sciences Division, Binding Sites, Center-Authored Paper, CRYSTALLIZATION, Crystallography, X-Ray, Deoxyribonucleases, Type II Site-Specific, DNA, Models, Molecular, Molecular Sequence Data, Mutation, Missense, Protein Binding, Protein Folding, Protein Multimerization, Protein Structure, Tertiary, Sequence Homology, Amino Acid

Abstract:

The GIY-YIG endonuclease family comprises hundreds of diverse proteins and a multitude of functions; none have been visualized bound to DNA. The structure of the GIY-YIG restriction endonuclease R.Eco29kI has been solved both alone and bound to its target site. The protein displays a domain-swapped homodimeric structure with several extended surface loops encircling the DNA. Only three side chains from each protein subunit contact DNA bases, two directly and one via a bridging solvent molecule. Both tyrosine residues within the GIY-YIG motif are positioned in the catalytic center near a putative nucleophilic water; the remainder of the active site resembles the HNH endonuclease family. The structure illustrates how the GIY-YIG scaffold has been adapted for the highly specific recognition of a DNA restriction site, in contrast to nonspecific DNA cleavage by GIY-YIG domains in homing endonucleases or structure-specific cleavage by DNA repair enzymes such as UvrC.