Diacylphosphatidylinositol (PI) is the starting reactant in the process of phosphatidylinositide-related signal transduction mediated through the lipid raft domain. We investigated intermolecular interactions of PI with major raft components, sphingomyelin (SM) and cholesterol (Chol), using surface pressure–molecular area (π–A) isotherm measurements. The classical mean molecular area versus composition plot showed that the measured mean molecular areas are smaller in PI/Chol mixed monolayers and larger in PI/SM mixed monolayers than those calculated on the basis of the ideal additivity. These results indicate that PI interacts attractively with Chol and repulsively with SM. In addition, we energetically evaluated the interaction of PI with SM/Chol mixtures and found that the mixing energy of PI/SM/Chol ternary monolayers decreased as the molar ratio of Chol to SM increased. In order to quantitatively analyze the distribution of PI we calculated the chemical potentials of mixing of PI into the SM/Chol mixed monolayer and into the dioleoylphosphatidylcholine (DOPC) monolayer, which was used as a model for the fluid matrix, on the basis of partial molecular area analysis. Analysis using the chemical potential of mixing of PI suggested that partition of PI molecules between these two monolayers can be changed by a factor of about 1.7 in response to change in Chol molar fraction in the SM/Chol mixed monolayer from 0.3 to 0.6 when the concentration of PI in the DOPC monolayer is kept constant at 7 mol%.

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The kinesin motor converts the chemical energy from ATP turnover into mechanical work, which produces successive 8-nm steps in the forward and backward direction along a microtubule. A key problem for kinesin mechanochemistry is explaining how ATP turnover is coordinated with mechanical work. We investigated this by measuring the ATP dependent properties of kinesin forward and backward steps using optical trapping nanometry. The results showed that the rate for both forward and backward steps are ATP-dependent, indicating that ATP binding to kinesin triggers both forward and backward steps. This suggests that ATP turnover in kinesin is not rigidly coupled to total mechanical work at high load.

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Correlation between fluctuations of the square of electron tunneling matrix element T_DA² and the donor-acceptor distance R_DA in the electron transfer (ET) reaction from bacteriopheophytin anion to the primary quinone of the reaction center in the photosynthetic bacteria Rhodobacter sphaeroides is investigated by a combined study of molecular dynamics simulations of the protein conformation fluctuation and quantum chemical calculations. We adopted two kinds of R_DA; edge-to-edge distance R_EE and center-to-center distance R_CC. The value of T_DA² distributed over more than 5 orders of magnitude and the fluctuation of the value of R_DA distributed over more than 1.8 Å for the 10⁶ instantaneous conformations of 1 ns simulation. We made analysis of the time-averaged correlation step by step as follows. We divide the 10⁶ simulation data into 1000/t parts of small data set to obtain the averaged data points of <T_DA²>_t and <R_EE>_t or <R_CC>_t. Plotting the 1000/t sets of log₁₀ <T_DA²>_t as a function of <R_EE>_t or <R_CC>_t, we made a principal coordinate analysis for these distributions. The slopes <β_E>_t and <β_C>_t of the primary axis are very large at small value of t and they are decreased considerably as t becomes large. The ellipticity for the distribution of <T_DA²>_t vs <R_EE>_t which can be a measure for the degree of correlation became very small when t is large, while it does not hold for the distribution of <T_DA²>_t vs <R_CC>_t. These results indicate that only the correlation between <T_DA²>_t and <R_EE>_t for large t satisfies the well-known linear relation (“Dutton law”), although the slope is larger than the original value 1.4 Å^–1. Based on the present result, we examined the analysis of the dynamic disorder by means of the single-molecule spectroscopy by Xie and co-workers with use of the “Dutton law”.

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Remarkable acceleration of enzymatic proteolysis by pressure at kbar range is reported with ubiquitin as substrate and α-chymotrypsin as enzyme. The acceleration is interpreted in terms of the shift of conformational equilibrium in ubiquitin from the non-degradable folded conformer to the enzyme-degradable unfolded conformer by pressure because of the lower volume of the latter, while the enzymatic activity of α-chymotrypsin is still largely retained. This mechanism is considered generally applicable to most globular proteins and the method of pressure-accelerated proteolysis will have an enormous potential utility in systems wherever efficient removal of proteins is needed.

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