Chemical shift assignments of domain 4 from the phosphohexomutase from Pseudomonas aeruginosa suggest that freeing perturbs its coevolved domain interface

A domain needed for the catalytic efficiency of an enzyme model of simple processivity and domain–domain interactions has been characterized by NMR. This domain 4 from phosphomannomutase/phosphoglucomutase (PMM/PGM) closes upon glucose phosphate and mannose phosphate ligands in the active site, and can modestly reconstitute activity of enzyme truncated to domains 1–3. This enzyme supports biosynthesis of the saccharide-derived virulence factors (rhamnolipids, lipopolysaccharides, and alginate) of the opportunistic bacterial pathogen Pseudomonas aeruginosa. ¹H, ¹³C, and ¹⁵N NMR chemical shift assignments of domain 4 of PMM/PGM suggest preservation and independence of its structure when separated from domains 1–3. The face of domain 4 that packs with domain 3 is perturbed in NMR spectra without disrupting this fold. The perturbed residues overlap both the most highly coevolved positions in the interface and residues lining a cavity at the domain interface.     

Experimental support for a right-handed vertical double helix

The experimentally observed geometry of a miniature double helix of high anti (χCN fixed ? 120°) nucleic acid structure indicates substantial interstrand base stacking with little intrastrand base stacking. This geometry is consistent with the right-handed vertical double helix in which the base planes are parallel to helical axis proposed by Olson [(1977) Proc. Natl. Acad. Sci. USA 74 , 1775] from theoretical calculations. The experimental data do not agree with the left-handed model in which the base planes are perpendicular to helical axis that has been proposed by Yathindra and Sundaralingam [(1976) Nucl. Acids Res. 3 , 729]. The theoretically computed chemical shift changes for the various double-helical configurations show that for the system examined, only the vertical model can explain the experimentally observed shift trends. The melting curve for the helix–coil transition for high anti dinucleoside monophosphate has been observed for the first time. Even though the experimental data support vertical double helices when χCN is fixed at 120°, data on naturally occurring nucleic acid structures indicate that they have no proclivity to enter into vertically stabilized double-helical arrays.