Pore Size Dependence of Mass Transfer of Zinc Myoglobin in a Single Mesoporous Silica Particle.

This study investigated the pore size dependence of the mass transfer of zinc myoglobin (ZnMb) in a single mesoporous silica particle through confocal fluorescence microspectroscopy. The ZnMb's fluorescence depth profile in the particle was analyzed by a spherical diffusion model, and the intraparticle diffusion coefficient was obtained. The intraparticle diffusion coefficient in the silica particle with various pore sizes (10, 15, 30, and 50 nm) was furthermore analyzed based on a pore and surface diffusion model. Although the mass transfer mechanism in all silica particles followed the pore and surface diffusion model, the adsorption and desorption of ZnMb affected the mass transfer depending on the pore size. The influence of the slow desorption of ZnMb became pronounced for large pore sizes (30 and 50 nm), which was revealed by simulation using a diffusion equation combined with the adsorption-desorption kinetics. The distribution of ZnMb was suppressed in small pore sizes (10 and 15 nm) owing to the adsorption of ZnMb onto the entrance of the pore. Thus, we revealed the mass transfer mechanism of ZnMb in the silica particle with different pore sizes.

Heme-bound tyrosine vibrations in hemoglobin M: Resonance Raman, crystallography, and DFT calculation.

Hemoglobins M (Hbs M) are human hemoglobin variants in which either the α or ß subunit contains a ferric heme in the α2ß2 tetramer. Though the ferric subunit cannot bind O2, it regulates O2 affinity of its counterpart ferrous subunit. We have investigated resonance Raman spectra of two Hbs, M Iwate (α87His → tyrosine [Tyr]) and M Boston (α58His → Tyr), having tyrosine as a heme axial ligand at proximal and distal positions, respectively, that exhibit unassigned resonance Raman bands arising from ferric (not ferrous) hemes at 899 and 876 cm-1. Our quantum chemical calculations using density functional theory on Fe-porphyrin models with p-cresol and/or 4-methylimidazole showed that the unassigned bands correspond to the breathing-like modes of Fe3+-bound Tyr and are sensitive to the Fe-O-C(Tyr) angle. Based on the frequencies of the Raman bands, the Fe-O-C(Tyr) angles of Hbs M Iwate and M Boston were predicted to be 153.5° and 129.2°, respectively. Consistent with this prediction, x-ray crystallographic analysis showed that the Fe-O-C(Tyr) angles of Hbs M Iwate and M Boston in the T quaternary structure were 153.6° and 134.6°, respectively. It also showed a similar Fe-O bond length (1.96 and 1.97 Å) and different tilting angles.

Kinetic Analysis of the Mass Transfer of Zinc Myoglobin in a Single Mesoporous Silica Particle by Confocal Fluorescence Microspectroscopy.

The adsorption/desorption mechanisms of biomolecules in porous materials have attracted significant attention because of their applications in many fields, including environmental, medical, and industrial sciences. Here, we employ confocal fluorescence microspectroscopy to reveal the diffusion behavior of zinc myoglobin (ZnMb, 4.4 nm × 4.4 nm × 2.5 nm) as a spherical protein in a single mesoporous silica particle (pore size of 15 nm). The measurement of the time course of the fluorescence depth profile of the particle reveals that intraparticle diffusion is the rate-limiting process of ZnMb in the particle. The diffusion coefficients of ZnMb in the particle for the distribution (Ddis) and release (Dre) processes are determined from the rate constants, e.g., Ddis = 1.65 × 10-10 cm2 s-1 and Dre = 3.68 × 10-10 cm2 s-1, for a 10 mM buffer solution. The obtained D values for various buffer concentrations are analyzed using the pore and surface diffusion model. Although surface diffusion is the main distribution process, the release process involves pore and surface diffusion, which have not been observed with small organic molecules; the mechanism of transfer of small molecules is pore diffusion alone. We demonstrate that the mass transfer kinetics of ZnMb in the silica particle can be explained well on the basis of pore and surface diffusion.