Tense State

Diagram 5: DesT tense state bound to SFA ligand.

The C terminus of DesT is well ordered and reveals the adenine ring interactions - it stacks between Ile199 and His205' adjescent to Trp204' and the N1 and N6 nitrogen atoms form Hydrogen bonds with the amide nitrogen of His205' and the carbonyl oxygen of His203' respectively.

The entrance to the ligand-binding pocket is largely unaffected by the ligand swap because the first seven carbons of the 16:0 chain form similar interactions to those of the oleoyl chain. It is  the tail of the 16:0 linear chain that inserts directly into the hydrophobic core and creates a new 9Å hydrophobic pocket - the acyl chain penetrates the phenyalanine-rich cluster below α6, and these residues adjust their positions to accommodate the ligand.

This results in 3 conformational changes :

α4 and α7 pack closer together, and a slight bend is induced in α4 due to the local unwinding of residues 68–71. This unwinding is associated with the movement of Phe71 and is stabilized by a hydrogen bond between the side chain of Thr70 and the carbonyl oxygen of Glu66.

The C terminus of α6, loop L6-L7 and the N terminus of α7 engage loop L8-L9, which extends across the interface and becomes ordered. L8-L9 contains five proline residues and appears to act as a clamp that specifically stabilizes the tense-state conformation.

The C termini of the paired α6 helices slide closer together by one turn (~4.4 Å), as seen in diagram 5, and this new location is stabilized by new interactions centered on Tyr115.

The recognition helices α3 and α3′ rotate by some 5°, and their center-to-center distance increases from the required DNA binding distance of 36.7 Å to 41.9 Å. It also reveals the substantial relative movements of the DNA-binding domains in the tense form, away from their optimal DNA-binding orientations in the relaxed form.

<previous page                                                                                                                                                                 next page>