Bacteriorhodopsin (bR) has a retinal with all-trans and 13-cis, 15-syn configurations whose isomeric ratio is close to 1 in the dark-adapted bR, and the population of the 13-cis, 15-syn configuration can be increased under pressure. In our previous study, we applied pressure to bR using centrifugal force introduced by sample spinning and demonstrated that the 15 N nuclear magnetic resonance (NMR) signal intensity of the Schiff base in 13-cis, 15-syn retinal increased. In this study, we demonstrate a pressure-induced change in the local dynamics of the protein side in the vicinity of retinal. In the 13 C dipolar decoupling and magic angle-spinning NMR spectra of [3-13 C]Ala-labeled bR under fast spinning at 10 and 12 kHz, corresponding to 44 and 63 bar, respectively, the 13 C NMR signal at 16.6 p.p.m. appeared prominently as a sharp peak. This peak is assigned to Ala81 and Ala84 residues located in the vicinity of the retinal. It was observed that the change in the local dynamics in the vicinity of the retinal was caused by the applied pressure. These experiments demonstrate that fast magic angle spinning in NMR provides a pressure source for investigating the structure and change in the dynamics of biomacromolecules.

Genetic evidence indicates that only the β-anomer of galactose is transported to Kluyveromyces lactis cells by galactose/glucose transporter Hgt1p, and that aldose-1-epimerase encoded by GAL10 is a prerequisite for growth on galactose. Minor aldose-1-epimerases other than Gal10p also exist in K. lactis. Using a mutant defective in both aldose-1-epimerases, we show by solid-state nuclear magnetic resonance spectroscopy that only β-anomer is transported in the cell and stays without or with a slow rate of conversion to α-anomer. Signals due to intracellular β-galactose appeared at two positions, both of which were shifted towards higher magnetic fields than that of β-galactose in aqueous solution, suggesting that incorporated galactose binds to cellular components, probably proteins.

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