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Phosphorylation and dephosphorylation of proteins by kinases and phosphatases are central to cellular responses and function. The structural effects of serine and threonine phosphorylation were examined in peptides and in proteins, by circular dichroism, NMR spectroscopy, bioinformatics analysis of the PDB, small-molecule X-ray crystallography, and computational investigations. Phosphorylation of both serine and threonine residues induces substantial conformational restriction in their physiologically more important dianionic forms. Threonine exhibits a particularly strong disorder-to-order transition upon phosphorylation, with dianionic phosphothreonine preferentially adopting a cyclic conformation with restricted φ (φ ~ –60 ̊) stabilized by three noncovalent interactions: a strong intraresidue phosphate-amide hydrogen bond, an n→π* interaction between consecutive carbonyls, and an n→σ* interaction between the phosphate Oγ lone pair and the antibonding orbital of C–Hβ that restricts the χ2 side chain conformation. Proline is unique among the canonical amino acids for its covalent cyclization on the backbone. Phosphothreonine can mimic proline's backbone cyclization via noncovalent interactions. The preferred torsions of dianionic phosphothreonine are φ,ψ = polyproline II helix > α-helix (φ ~ –60 ̊); χ1 = g–; χ2 ~ +115 ̊ (eclipsed C–H/O–P bonds). This structural signature is observed in diverse proteins, including in the activation loops of protein kinases and in protein-protein interactions. In total, these results suggest a structural basis for the differential use and evolution of threonine versus serine phosphorylation sites in proteins, with serine phosphorylation typically inducing smaller, rheostat-like changes, versus threonine phosphorylation promoting larger, step function-like switches, in proteins. 

The preparation of a new class of reactive porous solids, prepared via straightforward salt metathesis reactions, is described here. Reaction of the dimethylammonium salt of a magnesium‐based porous coordination cage with the chloride salt of [Cr^IICl(Me₄cyclam)]⁺affords a porous solid with concomitant removal of dimethylammonium chloride. The salt consists of the ions combined in the expected ratio based on their charge as confirmed by UV–vis and X‐ray photoelectron spectroscopies, ion chromatography (IC), and inductively coupled plasma mass spectrometry (ICP‐MS). The porous salt boasts a Brunauer‐Emmett‐Teller (BET) surface area of 213 m² g⁻¹. Single crystal X‐ray diffraction reveals the chromium(II) cations in the structure reside in the interstitial space between porous cages. Importantly, the chromium(II) centers, previously shown to react with O₂to afford reactive chromium(III)‐superoxide adducts, are still accessible in the solid state as confirmed by UV–vis spectroscopy. The site‐isolated reactive centers have competence toward hydrogen atom abstraction chemistry and display significantly increased stability and reactivity as compared to dissolved ions.

 

Proline residues within proteins lack a traditional hydrogen bond donor. However, the hydrogens of the proline ring are all sterically accessible, with polarized C−H bonds at Hα and Hδ that exhibit greater partial positive character and can be utilized as alternative sites for molecular recognition. C−H/O interactions, between proline C−H bonds and oxygen lone pairs, have been previously identified as modes of recognition within protein structures and for higher‐order assembly of protein structures. In order to better understand intermolecular recognition of proline residues, a series of proline derivatives was synthesized, including 4R‐hydroxyproline nitrobenzoate methyl ester, acylated on the proline nitrogen with bromoacetyl and glycolyl groups, and Boc‐4S‐(4‐iodophenyl)hydroxyproline methyl amide. All three derivatives exhibited multiple close intermolecular C−H/O interactions in the crystallographic state, with H⋅⋅⋅O distances as close as 2.3 Å. These observed distances are well below the 2.72 Å sum of the van der Waals radii of H and O, and suggest that these interactions are particularly favorable. In order to generalize these results, we further analyzed the role of C−H/O interactions in all previously crystallized derivatives of these amino acids, and found that all 26 structures exhibited close intermolecular C−H/O interactions. Finally, we analyzed all proline residues in the Cambridge Structural Database of small‐molecule crystal structures. We found that the majority of these structures exhibited intermolecular C−H/O interactions at proline C−H bonds, suggesting that C−H/O interactions are an inherent and important mode for recognition of and higher‐order assembly at proline residues. Due to steric accessibility and multiple polarized C−H bonds, proline residues are uniquely positioned as sites for binding and recognition via C−H/O interactions.