David Auble
Here you can see two different complexes of the BamHI restriction endonuclease bound to DNA. On the left is the complex of a BamHI homodimer bound to its palindromic restriction site GGATCC. The two subunits are shown in blue and green. This is referred to as the "pre-reactive" complex because the structure was solved in a solution containing calcium, an inhibitor of DNA cleavage, rather than another divalent cation such as magnesium or manganese which is required for DNA cleavage. Two metal ions (orange) are found in the complex, and calcium is bound in approximately the same positions as the normal metal co-factor. When BamHI binds its specific site, C-terminal α-helical arms unfold and form interactions with DNA. The interactions of the arms are actually asymmetric: one arm interacts with the minor groove while the other arm interacts with the sugar phosphate backbone. However, both subunits present catalytic groups (red) to the DNA similarly, leading to strand scission at symmetrical positions.
On the right is shown the structure of a complex of BamHI bound to a non-specific site, GAATTC. This non-specific site differs from the specific one by only a single base pair, and yet BamHI cutting at this non-specific site is impaired by about a million-fold. In the non-specific complex you can see that the DNA is not bound tightly in the BamHI DNA binding cleft, and in fact, there are no base-specific side chain interactions formed in the non-specific complex. The DNA binding cleft is widened due to movement of the BamHI monomers with respect to one another and the buried solvent-accessible surface area is much reduced compared to the specific complex. The C-terminal sequences form α-helices in the non-specific complex, resembling their conformation in the free enzyme. DNA cutting doesn't occur in the non-specific complex because the catalytic groups of the enzyme (shown in red) are quite distal from the DNA. However, BamHI does interact with the DNA backbone, and as discussed in class, this is what would be expected for a protein that slides or can hop along DNA. The non-specific complex thus provides a picture of what a protein might look like as it scans DNA for a specific site.
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2bam: specific complex |
1esg: non-specific complex |