A magnetic field and high-pressure are effective parameters to control protein crystallization. A magnetic field significantly affects the number, orientation and growth rate (morphology) of protein crystals and damps the buoyancy convection in protein solutions. Quality of protein crystals can be also improved by a magnetic field. The improvement in the crystal quality is due to a magnetic orientation effect. High-pressure can also give large effects on the solubility and growth kinetics of protein crystals. Changes in the solubility with pressure results from a difference in the volume between bulk water and the water hydrating the contact regions of protein molecules inside the crystal.
The force field of a protein is an important and fundamental tool for protein scientists in modeling the structures. Although currently available all-atom protein force fields have been well tuned, it is still a difficult and time-consuming task to quantitatively reproduce the thermodynamic parameters by computer simulation, especially in water. In this article, a new all-atom protein force field (named as the SAAP force field), which is based on a novel idea to compose a protein force field by using single amino acid potential (SAAP) functions, is presented.
To elucidate the role of flexible loops in the stability and function of Escherichia coli dihydrofolate reductase (DHFR), glycine-67, glycine-121, and alanine-145 in the three different loops were substituted with several amino acids by site-directed mutagenesis. The free energy change of unfolding by urea decreased with an increase in the hydrophobicity and volume of amino acids introduced at any site. In the enzyme reaction, Km was only slightly influenced, but kcat was decreased by mutation, especially with significant effects at site 121. Adiabatic compressibility also largely changed by mutation, indicating the modified volume fluctuation of the native state. Further, nonadditive effects were observed in the stability and function of the double mutants at sites 67 and 121 although they are separated by 27.4 Å. These results indicate that the effects of mutations extend to almost the whole protein molecule via reconstituted atomic packing or long-range interaction, and then the loop regions play important roles in the stability, fluctuation, and function of this enzyme.
Kinesin is an ATP-driven molecular motor that moves processively along a microtubule in a stepwise manner. The steps occur not only in the forward direction, but also in the backward. Here, we have studied the bidirectional stepping mechanism of kinesin motors. The stepping mechanism of the forward and backward movements was well characterized by Feynman's thermal ratchet model. The driving force of the stepwise movement is essentially Brownian motion, but it is biased in the forward direction by utilizing the free energy released from the hydrolysis of ATP.
Plant heme oxygenase (HO), which catalyzes conversion of heme to biliverdin, being further converted to photo-receptive phycobilins, had been known only in the cell extracts of primitive red algae and cyanobacteria for these twenty years. Recent development of gene analysis, however, has shown that HO is ubiquitously contained in variety of living species. We have succeeded for the first time in obtaining recombinant HO protein of cyanobacteria and in characterizing the HO protein and its heme complex by spectroscopic analyses. Here, we review the existence, role, and characteristics of cyanobacterial and plant HOs in comparison with those of mammalian and bacterial HOs.