Herein, we demonstrate “direct” 13C hyperpolarization of 13C‐acetate via signal amplification by reversible exchange (SABRE). The standard SABRE homogeneous catalyst [Ir‐IMes; [IrCl(COD)(IMes)], (IMes=1,3‐bis(2,4,6‐trimethylphenyl), imidazole‐2‐ylidene; COD=cyclooctadiene)] was first activated in the presence of an auxiliary substrate (pyridine) in alcohol. Following addition of sodium 1‐13C‐acetate, parahydrogen bubbling within a microtesla magnetic field (i.e. under conditions of SABRE in shield enables alignment transfer to heteronuclei, SABRE‐SHEATH) resulted in positive enhancements of up to ≈100‐fold in the 13C NMR signal compared to thermal equilibrium at 9.4 T. The present results are consistent with a mechanism of “direct” transfer of spin order from parahydrogen to 13C spins of acetate weakly bound to the catalyst, under conditions of fast exchange with respect to the 13C acetate resonance, but we find that relaxation dynamics at microtesla fields alter the optimal matching from the traditional SABRE‐SHEATH picture. Further development of this approach could lead to new ways to rapidly, cheaply, and simply hyperpolarize a broad range of substrates (e.g. metabolites with carboxyl groups) for various applications, including biomedical NMR and MRI of cellular and in vivo metabolism.
An atomic view of a main aqueous conformation of cyclosporine A (CycA), an important 11-amino-acid macrocyclic immunosuppressant, is reported. For decades, it has been a grand challenge to determine the conformation of free CycA in an aqueous-like solution given its poor water solubility. Using a combination of X-ray and single-crystal neutron diffraction, we unambiguously resolve a unique conformer (A1) with a novel cis-amide between residues 11 and 1 and two water ligands that stabilize hydrogen bond networks. NMR spectroscopy and titration experiments indicate that the novel conformer is as abundant as the closed conformer in 90/10 (v/v) methanol/water and is the main conformer at 10/90 methanol/water. Five other conformers were also detected in 90/10 methanol/water, one in slow exchange with A1, another one in slow exchange with the closed form and three minor ones, one of which contains two cis amides Abu2−Sar3 and MeBmt1−MeVal11. These conformers help better understand the wide spectrum of membrane permeability observed for CycA analogues and, to some extent, the binding of CycA to protein targets.
We show the simultaneous generation of hyperpolarized C-labeled acetate and N-labeled imidazole following spin-relay of hyperpolarization and hydrolysis of the acetyl moiety on 1- C- N -acetylimidazole. Using SABRE-SHEATH (Signal Amplification by Reversible Exchange in SHield Enables Alignment Transfer to Heteronuclei), transfer of spin order occurs from parahydrogen to acetylimidazole N atoms and the acetyl C site (≈263-fold enhancement), giving rise to relatively long hyperpolarization lifetimes at 0.3 T (T ≈52 s and ≈149 s for C and N, respectively). Immediately following polarization transfer, the C-labeled acetyl group is hydrolytically cleaved to produce hyperpolarized C-acetate/acetic acid (≈140-fold enhancement) and N-imidazole (≈180-fold enhancement), the former with a C T of ≈14 s at 0.3 T. Straightforward synthetic routes, efficient spin-relay of SABRE hyperpolarization, and facile bond cleavage open a door to the cheap and rapid generation of long-lived hyperpolarized states within a wide range of molecular targets, including biologically relevant carboxylic acid derivatives, for metabolic and pH imaging.
Herein, we demonstrate “direct” 13C hyperpolarization of 13C‐acetate via signal amplification by reversible exchange (SABRE). The standard SABRE homogeneous catalyst [Ir‐IMes; [IrCl(COD)(IMes)], (IMes=1,3‐bis(2,4,6‐trimethylphenyl), imidazole‐2‐ylidene; COD=cyclooctadiene)] was first activated in the presence of an auxiliary substrate (pyridine) in alcohol. Following addition of sodium 1‐13C‐acetate, parahydrogen bubbling within a microtesla magnetic field (i.e. under conditions of SABRE in shield enables alignment transfer to heteronuclei, SABRE‐SHEATH) resulted in positive enhancements of up to ≈100‐fold in the 13C NMR signal compared to thermal equilibrium at 9.4 T. The present results are consistent with a mechanism of “direct” transfer of spin order from parahydrogen to 13C spins of acetate weakly bound to the catalyst, under conditions of fast exchange with respect to the 13C acetate resonance, but we find that relaxation dynamics at microtesla fields alter the optimal matching from the traditional SABRE‐SHEATH picture. Further development of this approach could lead to new ways to rapidly, cheaply, and simply hyperpolarize a broad range of substrates (e.g. metabolites with carboxyl groups) for various applications, including biomedical NMR and MRI of cellular and in vivo metabolism.
The self-association of metabolites into well-ordered assemblies at the nanoscale has significant biological and medical implications. The thiol-containing amino acid cysteine (CYS) can assemble into amyloid-like nanofibrils, and its oxidized form, the disulfide-bonded cystine (CTE), forms hexagonal crystals as those found in cystinuria due to metabolic disorder. Yet, there have been no attempts to connect these two phenomena, especially the fibril-to-crystal transition. Here, we reveal that these are not separated events, and the CYS-forming amyloid fibrils are mechanistically linked to hexagonal CTE crystals. For the first time, we demonstrated that cysteine fibrils are a prerequisite for forming cystine crystals, as observed experimentally. To further understand this mechanism, we studied the effects of thiol-containing cystinuria drugs (tiopronin, TIO; and d-penicillamine, PEN) and the canonical epigallocatechin gallate (EGCG) amyloid inhibitor on fibril formation by CYS. The thiol-containing drugs do not solely interact with monomeric CYS via disulfide bond formation but can disrupt amyloid formation by targeting CYS oligomers. On the other hand, EGCG forms inhibitor-dominant complexes (more than one EGCG molecule per cysteine unit) to prevent CYS fibril formation. Interestingly, while CYS can be oxidized into CTE, the thiol drugs can reduce CTE back to CYS. We thus suggest that the formation of crystals in cystinuria could be halted at the initial stage by targeting CYS fibril formation as an alternative to solubilizing the water-insoluble hexagonal CTE crystals at a later stage. Taken together, we depicted a complex hierarchical organization in a simple amino acid assembly with implications for therapeutic intervention.
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