Biocatalytic production of D-p-hydroxyphenylglycine by optimizing protein expression and cell wall engineering in Escherichia coli.

D-p-hydroxyphenylglycine (D-HPG) functions as an intermediate and has important value in antibiotic industries. The high pollution and costs from chemical processes make biotechnological route for D-HPG highly desirable. Here, a whole-cell transformation process by D-hydantoinase(Hase) and D-carbamoylase(Case) was developed to produce D-HPG from DL-hydroxyphenylhydantoin(DL-HPH) in Escherichia coli. The artificially designed ribosome binding site with strong intensity significantly facilitated the protein expression of limiting step enzyme Case. Next, the cell wall permeability was improved by disturbing the peptidoglycan structure by overproduction of D,D-carboxypeptidases without obviously affecting cell growth, to increase the bioavailability of low soluble hydantoin substrate. By fine-tuning regulation of expression level of D,D-carboxypeptidase DacB, the final production yield of D-HPG increased to 100% with 140 mM DL-HPH substrate under the optimized transformation conditions. This is the first example to enhance bio-productivity of chemicals by cell wall engineering and creates a new vision on biotransformation of sparingly soluble substrates. Additionally, the newly demonstrated 'hydroxyl occupancy' phenomenon when Case reacts with hydroxyl substrates provides a referential information for the enzyme engineering in future.

Hydrogen bond network in the hydration layer of the water confined in nanotubes increasing the dielectric constant parallel along the nanotube axis.

The water confined in nanotubes has been extensively studied, because of the potential usages in drug delivery and desalination. The radial distribution of the dielectric constant parallel along the nanotube axis was obtained by molecular dynamics simulations in a carbon nanotube and a nanotube with a very small van der Waals potential. The confined water was divided into two parts, the middle part water and the hydration water. In both cases, the hydrogen bond orientation of the middle water is isotropic, while the hydrogen bonds in hydration layers are apt to parallel along the nanotube axis. Therefore, the hydration water has higher dipole correlations increasing the dielectric constant along the nanotube axis.

Anisotropic dielectric relaxation of the water confined in nanotubes for terahertz spectroscopy studied by molecular dynamics simulations.

The dynamics and structure of the hydrogen-bond network in confined water are of importance in understanding biological and chemical processes. Recently, terahertz (THz) time domain spectroscopy was widely applied for studying the kinetics of molecules and the hydrogen-bond network in water. However, the characteristics of the THz spectroscopy varying with respect to the confinement and the mechanism underlying the variation are still unclear. Here, on the basis of molecular dynamics simulations, the relationship between the anisotropic dielectric relaxation and the structure of the water confined in a carbon nanotube (CNT) was investigated. The results show that there are two preferred hydrogen-bond orientations of the confined water in the nanotube: (1) parallel to the CNT axis and (2) perpendicular to the CNT axis, which are clearly different. Moreover, the response of the orientations to the increment of the CNT diameters is opposite, leading to the opposite variations of the dielectric relaxation times along the two directions. The anisotropy in the relaxation time can be presented by the anisotropic dielectric permittivity which is able to be observed through THz spectroscopy. The anormal behaviors above are attributed to the special structure of the water close to the nanotube wall due to the confinement and hydrophobicity of CNT. These studies contribute an important step in understanding the THz experiments of water in nanoscales, and designing a chamber for specific chemical and biological reactions by controlling the diameters and materials of the nanotube.

DNA base pair hybridization and water-mediated metastable structures studied by molecular dynamics simulations.

The base pair hybridization of a DNA segment was studied using molecular dynamics simulation. The results show the obvious correlation between the probability of successful hybridization and the accessible surface area to water of two successive base pairs, including the unpaired base pair adjacent to paired base pair and this adjacent paired base pair. Importantly, two metastable structures in an A-T base pair were discovered by the analysis of the free energy landscape. Both structures involved addition of a water molecule to the linkage between the two nucleobases in one base pair. The existence of the metastable structures provide potential barriers to the Watson-Crick base pair, and numerical simulations show that those potential barriers can be surmounted by thermal fluctuations at higher temperatures. These studies contribute an important step toward the understanding of the mechanism in DNA hybridization, particularly the effect of temperature on DNA hybridization and polymerase chain reaction. These observations are expected to be helpful for facilitating experimental bio/nanotechnology designs involving fast hybridization.

Kinetics of water filling the hydrophobic channels of narrow carbon nanotubes studied by molecular dynamics simulations.

The kinetics of water filling narrow single-walled carbon nanotubes was studied using molecular dynamics simulations. The time required to fully fill a nanotube was linear with respect to the tube length. We observed that water molecules could enter into nanotubes of different lengths, either from one end or from both ends. The probability of having a nanotube filled completely from both ends increased exponentially with the tube length. For short tubes, filling usually proceeded from only one end. For long tubes, filling generally proceeded from both tube ends over three stages, i.e., filling from one end, filling from both ends, and filling from both ends with the dipole reorientation of water molecules to give a concerted ordering within the fully filled tube. The water molecules in the partially filled nanotube were hydrogen bonded similarly to those in the fully filled nanotube. Simulations for the reference Lennard-Jones fluid without hydrogen bonds were also performed and showed that the filling behavior of water molecules can be attributed to strong intermolecular hydrogen bonding.

DNA structural changes under different stretching methods studied by molecular dynamics simulations.

We present a molecular dynamics simulation study of 22-mer DNA conformational variations obtained by stretching both 3'-termini and both 5'-termini. Stretching 3'-termini by 3.5 nm required 142 kJ mol(-1) and the force plateau was approximately 80 pN, whereas stretching 5'-termini by the same length required 190 kJ mol(-1) and the force plateau was approximately 100 pN. Stretching 3'-termini led to a larger untwisting of the double helix and the successive base pairs rolled to the side of the DNA minor groove, while stretching 5'-termini resulted in the base pairs rolling to the major groove side and reducing of the diameter of DNA molecule. The most distinctive difference between stretching 3'-termini and 5'-termini was that at the force plateau region stretching the 5'-termini resulted in breakage of the base pairs, which considerably disturbed the structure of the DNA double helix. All of the variations of base rotation and translation for both stretching methods took place when the relative length of DNA l was longer than 1.2, which was the point the force plateau appeared.

Size dependence of nanoscale confinement on chiral transformation.

Molecular dynamic simulations of the chiral transition of a difluorobenzo[c]phenanthrene molecule (C(18)H(12)F(2), D molecule) in single-walled boron-nitride nanotubes (SWBNNTs) revealed remarkable effects of the nanoscale confinement. The critical temperature, above which the chiral transition occurs, increases considerably with the nanotube diameter, and the chiral transition frequency decreases almost exponentially with respect to the reciprocal of temperature. The chiral transitions correlate closely with the orientational transformations of the D molecule. Furthermore, the interaction energy barriers between the D molecule and the nanotube for different orientational states can characterize the chiral transition. This implies that the temperature threshold of a chiral transition can be controlled by a suitable nanotube. These findings provide new insights to the effect of nanoscale confinement on molecular chirality.

Graphene on Au(111): a highly conductive material with excellent adsorption properties for high-resolution bio/nanodetection and identification.

Based on numerical simulations and experimental studies, we show that a composite material which consists of a sheet of graphene on a Au(111) surface exhibits both an excellent conductivity and the ability to stably adsorb biomolecules. If we use this material as a substrate, the signal-to-noise ratios can be greatly enhanced. The key to this unique property is that graphene can stably adsorb carbon-based rings, which are widely present in biomolecules, due to pi-stacking interactions while the substrate retains the excellent conductivity of gold. Remarkably, the signal-to-noise ratio is found to be so high that the signal is clearly distinguishable for different nucleobases when an ssDNA is placed on this graphene-on-Au(111) material. Our finding opens opportunities for a range of bio/nano-applications including single-DNA-molecule-based biodevices and biosensors, particularly, high-accuracy sequencing of DNA strands with repeating segments.

Manipulating biomolecules with aqueous liquids confined within single-walled nanotubes.

Confinement of molecules inside nanoscale pores has become an important method for exploiting new dynamics not happening in bulk systems and for fabricating novel structures. Molecules that are encapsulated in nanopores are difficult to control with respect to their position and activity. On the basis of molecular dynamics simulations, we have achieved controllable manipulation, both in space and time, of biomolecules with aqueous liquids inside a single-walled nanotube by using an external charge or a group of external charges. The remarkable manipulation abilities are attributed to the single-walled structure of the nanotube that the electrostatic interactions of charges inside and outside the single-walled nanotube are strong enough, and the charge-induced dipole-orientation ordering of water confined in the nanochannel so that water has a strong interaction with the external charge. These designs are expected to serve as lab-in-nanotube for the interactions and chemical reactions of molecules especially biomolecules, and have wide applications in nanotechnology and biotechnology.