Table 2.

Restraints Used in Simulated Annealing

Type of Restraint	Details	Notes
Experimental FRET distances	Treated as NOE distances	Additional distance to deal with linker length
Experimentally determined secondary structure	Base pairing	H-bond distances
	Base pair planarity	Not constraining to allow for “propeller” twist of base pair
	Double-helical backbone torsional restraints	Based on values for A-type helix from XPLOR standard library
	Restraints on ribose ring to favor the 3′-endo pucker	Based on standard CNS values
Restraints to prevent “laddering”	Staggered strand-to-strand restraints across base pairs	Staggering is to allow for inclined angle of base pairs in A-form helix
(Applied only to helical regions)	End-to-end restraints on double helical sections	Length adjusted to account for differing numbers of base pairs
	Backbone torsional restraints	Based on X-PLOR values for nucleotides in A-form helix

See also red boxes in Figure 2. The final structure after simulated annealing is shown in Figure 5.