Reducing the computational cost of NMR crystallography of organic powders at natural isotopic abundance with the help of 13C-13C dipolar couplings

Structure determination of functional organic compounds remains a formidable challenge when the sample exists as a powder. Nuclear magnetic resonance crystallography approaches based on the comparison of experimental and Density Functional Theory (DFT)-computed ¹H chemical shifts have already demonstrated great potential for structure determination of organic powders, but limitations still persist. In this study, we discuss the possibility of using ¹³C-¹³C dipolar couplings quantified on powdered theophylline at natural isotopic abundance with the help of dynamic nuclear polarization, to realize a DFT-free, rapid screening of a pool of structures predicted by ab initio random structure search. We show that although ¹³C-¹³C dipolar couplings can identify structures possessing long range structural motifs and unit cell parameters close to those of the true structure, it must be complemented with other data to recover information about the presence and the chemical nature of the supramolecular interactions.     

Evidence for Multiple Binding Modes in the Initial Contact Between SARS-CoV-2 Spike S1 Protein and Cell Surface Glycans**

Infection of host cells by SARS-CoV-2 begins with recognition by the virus S (spike) protein of cell surface heparan sulfate (HS), tethering the virus to the extracellular matrix environment, and causing the subunit S1-RBD to undergo a conformational change into the ‘open’ conformation. These two events promote the binding of S1-RBD to the angiotensin converting enzyme 2 (ACE2) receptor, a preliminary step toward viral-cell membrane fusion. Combining ligand-based NMR spectroscopy with molecular dynamics, oligosaccharide analogues were used to explore the interactions between S1-RBD of SARS CoV-2 and HS, revealing several low-specificity binding modes and previously unidentified potential sites for the binding of extended HS polysaccharide chains. The evidence for multiple binding modes also suggest that highly specific inhibitors will not be optimal against protein S but, rather, diverse HS-based structures, characterized by high affinity and including multi-valent compounds, may be required.     

Quantification of Intact and Truncated Stromal Cell-Derived Factor-1α in Circulation by Immunoaffinity Enrichment and Tandem Mass Spectrometry

Stromal cell-derived factor 1α (SDF-1α) or CXCL12 is a small pro-inflammatory chemoattractant cytokine and a substrate of dipeptidyl peptidase IV (DPP-IV). Proteolytic cleavage by DPP-IV inactivates SDF-1α and attenuates its interaction with CXCR4, its cell surface receptor. To enable investigation of suppression of such inactivation with pharmacologic inhibition of DPP-IV, we developed quantitative mass spectrometric methods that differentiate intact SDF-1α from its inactive form. Using top-down strategy in quantification, we demonstrated the unique advantage of keeping SDF-1α’s two disulfide bridges intact in the analysis. To achieve the optimal sensitivity required for quantification of intact and truncated SDF-1α at endogenous levels in blood, we coupled nano-flow tandem mass spectrometry with antibody-based affinity enrichment. The assay has a quantitative range of 20 pmol/L to 20 nmol/L in human plasma as well as in rhesus monkey plasma. With only slight modification, the same assay can be used to quantify SDF-1α in mice. Using two in vivo animal studies as examples, we demonstrated that it was critical to differentiate intact SDF-1α from its truncated form in the analysis of biomarkers for pharmacologic inhibition of DPP-IV activity. These novel methods enable translational research on suppression of SDF-1 inactivation with DPP-IV inhibition and can be applied to relevant clinical samples in the future to yield new insights on change of SDF-1α levels in disease settings and in response to therapeutic interventions.

Scission of Carbon Monoxide Using TaR3, R=(N(tBu)Ph) or OSi(tBu)3: A DFT Investigation

The experimentally known reduction of carbon monoxide using a 3‐coordinate [Ta(silox)₃] (silox=OSi(tBu)₃) complex initially forms a ketenylidene [(silox)₃Ta? CCO], followed by a dicarbide [(silox)₃Ta? CC? Ta(silox)₃] structure. The mechanism for this intricate reaction has finally been revealed by using density functional theory, and importantly a likely structure for the previously unknown intermediate [(silox)₃Ta? CO]₂ has been identified. The analysis of the reaction pathway and the numerous intermediates has also uncovered an interesting pattern that results in CO cleavage, that being scission from a structure of the general form [(silox)₃Ta? C_nO] in which n is even. When n is odd, cleavage cannot occur. The mechanism has been extended to consider the effect of altering both the metal species and the ligand environment. Specifically, we predict that introducing electron‐rich metals to the right of Ta in the periodic table to create mixed‐metal dinuclear intermediates shows great promise, as does the ligand environment of the Cummins‐style 3‐coordinate amide structure. This latter environment has the added complexity of improved electron donation from amide rotation that can significantly increase the reaction exothermicity.