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The structure and conformational behavior of sulfonium salt glycosidase inhibitors in solution: a combined quantum mechanical NMR approach.

ABSTRACT: We present the conformational analysis of an inhibitor of alpha-mannosidase, based on a novel sulfonium salt structure (1) that mimics the mannosyl cation intermediate. Because of the number of possible isomeric structures for 1, as well as its complex molecular structure, traditional conformational analysis by NMR was not applicable. Instead, a single experimentally consistent structure was obtained from finite perturbation quantum mechanical calculations of the NMR J-couplings at the B3LYP/6-311G** level. Using a full relaxation matrix analysis, we showed that the quantum-predicted NMR structure was the only isomer that was consistent with the experimental NOE intensities. The results illustrate the potential for finite perturbation calculations to be useful in the analysis of complex charged molecules.

SUBMITTER: Gonzalez-Outeirino J

PROVIDER: S-EPMC1386731 | biostudies-literature | 2004 Jun

REPOSITORIES: biostudies-literature

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The structure and conformational behavior of sulfonium salt glycosidase inhibitors in solution: a combined quantum mechanical NMR approach.

Gonzalez-Outeiriño Jorge J Glushka John J Siriwardena Aloysius A Woods Robert J RJ

Journal of the American Chemical Society 20040601 22

We present the conformational analysis of an inhibitor of alpha-mannosidase, based on a novel sulfonium salt structure (1) that mimics the mannosyl cation intermediate. Because of the number of possible isomeric structures for 1, as well as its complex molecular structure, traditional conformational analysis by NMR was not applicable. Instead, a single experimentally consistent structure was obtained from finite perturbation quantum mechanical calculations of the NMR J-couplings at the B3LYP/6-3 ...[more]

PMID: 15174846

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Project description:BackgroundHalophiles are extremophiles that thrive in environments with very high concentrations of salt. Although the salt reliance and physiology of these extremophiles have been widely investigated, the molecular working mechanisms of their enzymes under salty conditions have been little explored.Methodology/principal findingsA halophilic esterolytic enzyme LipC derived from archeaon Haloarcula marismortui was overexpressed from Escherichia coli BL21. The purified enzyme showed a range of hydrolytic activity towards the substrates of p-nitrophenyl esters with different alkyl chains (n = 2-16), with the highest activity being observed for p-nitrophenyl acetate, consistent with the basic character of an esterase. The optimal esterase activities were found to be at pH 9.5 and [NaCl] = 3.4 M or [KCl] = 3.0 M and at around 45 degrees C. Interestingly, the hydrolysis activity showed a clear reversibility against changes in salt concentration. At the ambient temperature of 22 degrees C, enzyme systems working under the optimal salt concentrations were very stable against time. Increase in temperature increased the activity but reduced its stability. Circular dichroism (CD), dynamic light scattering (DLS) and small angle neutron scattering (SANS) were deployed to determine the physical states of LipC in solution. As the salt concentration increased, DLS revealed substantial increase in aggregate sizes, but CD measurements revealed the maximal retention of the alpha-helical structure at the salt concentration matching the optimal activity. These observations were supported by SANS analysis that revealed the highest proportion of unimers and dimers around the optimal salt concentration, although the coexistent larger aggregates showed a trend of increasing size with salt concentration, consistent with the DLS data.Conclusions/significanceThe solution alpha-helical structure and activity relation also matched the highest proportion of enzyme unimers and dimers. Given that all the solutions studied were structurally inhomogeneous, it is important for future work to understand how the LipC's solution aggregation affected its activity.

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The structure and conformational behavior of sulfonium salt glycosidase inhibitors in solution: a combined quantum mechanical NMR approach.

Publications

The structure and conformational behavior of sulfonium salt glycosidase inhibitors in solution: a combined quantum mechanical NMR approach.

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