Programing a cyanide-free transformation of aldehydes to nitriles and one-pot synthesis of amides through tandem chemo-enzymatic cascades.

Nitriles are broadly applied to synthesize pharmaceuticals, agrochemicals, and materials because of their versatile transformation. Although various methods have been developed for introducing a nitrile group into organic molecules, most of them entail the use of highly toxic chemicals, transition metals, or harsh conditions. In this work, we reported a greener chemo-enzymatic cascade to synthesize alky and aryl nitriles from readily accessible aldehydes, that were further transformed into corresponding amides via an artificial enzyme cascade. A biphasic reaction system was designed to bridge chemical synthesis and enzymatic catalysis through simple phase separation. The biphasic system mainly perfectly avoided the inactivation of hydroxylamine on aldoxime dehydratase from Pseudomonas putida (OxdF1) and nitrile hydratase from Aurantimonas manganoxydans ATCC BAA-1229 (NHase1229). For the synthesis of various nitriles, moderate isolation yields of approximately 60% were obtained by the chemo-enzymatic cascade. Interestingly, two seemingly conflicting reactions of dehydration and hydration were sequentially proceeded to synthesize amides by the synergistic catalysis of OxdF1 and NHase1229 in E. coli cells. An isolation yield of approximately 62% was achieved for benzamide at the one-liter scale. In addition, the shuttle transport of substrates and products between two phases is convenient for the product separation and n-hexane recycling. Thus, the chemo-enzymatic cascade shows a potential application in the cyanide-free and large-scale synthesis of nitriles and amides.

Catalytically active inclusion bodies (CatIBs) induced by terminally attached self-assembling coiled-coil domains: To enhance the stability of (R)-hydroxynitrile lyase.

The catalytically-active inclusion bodies (CatIBs) represent a promising strategy for immobilizing enzyme without additional carriers and chemicals, which has aroused great attention in academic and industrial communities. In this work, we discovered two natural parallel right-handed coiled-coil tetramer peptides from PDB database by a structural mining strategy. The two self-assembling peptides, NSPdoT from rotavirus and HVdoT from human Vasodilator-stimulated phosphoprotein, efficiently induced the CatIBs formation of a (R)-Hydroxynitrile lyase from Arabidopsis thaliana (AtHNL) in Escherichia coli cells. This is convenient to simultaneously purify and immobilize the target proteins as biocatalysts. As expected, HVdoT-AtHNL and NSPdoT-AtHNL possessed drastically increased tolerance toward lower pH values, which will be very critical to synthesize cyanohydrins under acidic condition for suppressing the non-enzymatic side reaction. In addition. AtHNL-CatIBs are produced at high yield in host cells as bioactive microparticles, which exhibited high thermal and pH stabilities. Therefore, the CatIBs method represent a promising application for the immobilization of enzymes in the biocatalysis field.

Cyanide-free synthesis of aromatic nitriles from aldoximes: Discovery and application of a novel heme-containing aldoxime dehydratase.

Aromatic nitriles are important structural motifs that frequently existed in pharmaceutical drugs. Due to the convenient synthesis of aldoximes from aldehydes, the dehydration of aldoximes to corresponding nitriles by aldoxime dehydratases (Oxds) is considered as a safe and robust enzymatic production route. Although the Oxd genes are widely distributed in microbial kingdom, so far less than ten Oxds were expressed and further characterized. In this study, we found 26 predicted putative Oxd genes from the GenBank database using a genome mining strategy. The Oxd gene from Pseudomonas putida F1 was cloned and functionally expressed in Escherichia coli BL21 (DE3). The amino acid sequence of OxdF1 shows high identities of 33∼85 % to other characterized Oxds, and contained a ferrous heme as the catalytic site. The optimum reaction pH and temperature of recombinant OxdF1 were 7.0 and 35 °C, respectively. OxdF1 was stable in pH 7.0 potassium phosphate buffer at 30 °C, and its half-life was approximately 3.8 h. OxdF1 can efficiently dehydrate aromatic and heterocyclic aldoximes to nitriles, such as 2-bromobenzaldoxime, 2-chloro-6-fluorobenzaldoxime, thiophene-2-carboxaldoxime, and pyridine-3-aldoxime. Therefore, the recombinant OxdF1 shows a potential application in the cyanide-free synthesis of aromatic nitriles.

Effects of acute heat stress on protein expression and histone modification in the adrenal gland of male layer-type country chickens.

The adrenal gland responds to heat stress by epinephrine and glucocorticoid release to alleviate the adverse effects. This study investigated the effect of acute heat stress on the protein profile and histone modification in the adrenal gland of layer-type country chickens. A total of 192 roosters were subject to acute heat stress and thereafter classified into a resistant or susceptible group according to body temperature change. The iTRAQ analysis identified 80 differentially expressed proteins, in which the resistant group had a higher level of somatostatin and hydroxy-δ-5-steroid dehydrogenase but a lower parathymosin expression in accordance with the change of serum glucocorticoid levels. Histone modification analysis identified 115 histone markers. The susceptible group had a higher level of tri-methylation of histone H3 lysine 27 (H3K27me3) and showed a positive crosstalk with K36me and K37me in the H3 tails. The differential changes of body temperature projected in physiological regulation at the hypothalamus-pituitary-adrenal axis suggest the genetic heterogeneity in basic metabolic rate and efficiency for heat dissipation to acclimate to thermal stress and maintain body temperature homeostasis. The alteration of adrenal H3K27me3 level was associated with the endocrine function of adrenal gland and may contribute to the thermotolerance of chickens.

A new deep learning method for displacement tracking from ultrasound RF signals of vascular walls.

It is necessary to monitor the mechanical properties of arteries which directly related to cardiovascular diseases (CVDs) in the early stages. In this study, we proposed a new method based on deep learning (DL) to track the displacement of the vessel wall from the ultrasound radio-frequency (RF) signals, which is a key technique to achieve quantitative measurement of vascular biomechanics. In comparison with traditional method, both results on simulation and experimental carotid artery data demonstrated that the DL method has higher accuracy for motion tracking of artery walls. Hence, the DL method can be widely applied so that can predict the early pathology of cardiovascular system.

A novel transcranial ultrasound imaging method with diverging wave transmission and deep learning approach.

Real time brain transcranial ultrasound imaging is extremely intriguing because of its numerous applications. However, the skull causes phase distortion and amplitude attenuation of ultrasound signals due to its density: the speed of sound is significantly different in bone tissue than in soft tissue. In this study, we propose an ultrafast transcranial ultrasound imaging technique with diverging wave (DW) transmission and a deep learning approach to achieve large field-of-view with high resolution and real time brain ultrasound imaging. DW transmission provides a frame rate of several kiloHz and a large field of view that is suitable for human brain imaging via a small acoustic window. However, it suffers from poor image quality because the diverging waves are all unfocused. Here, we adopted adaptive beamforming algorithms to improve both the image contrast and the lateral resolution. Both simulated and in situ experiments with a human skull resulted in significant image improvements. However, the skull still introduces a wavefront offset and distortion, which degrades the image quality even when adaptive beamforming methods are used. Thus, we also employed a U-Net neural network to detect the contour and position of the skull directly from the acquired RF signal matrix. This approach avoids the need for beamforming, image reconstruction, and image segmentation, making it more suitable for clinical use.

A novel transcranial ultrasound imaging method with diverging wave.

Real time transcranial ultrasound imaging of brain can be extremely intriguing because of its numerous applications. In this study, we proposed an ultrafast transcranial ultrasound imaging technique with diverging wave (DW) transmission, which has been a promising technique to image moving objects, such as complex blood flow field and transient elastography. However, diverging waves are all unfocused waves, which makes their image quality, especially the lateral resolution and contrast, has not yet been satisfactory. Here we tried to apply the adaptive beamforming algorithms to improve both the image contrast and the lateral resolution. Simulation and phantom experiments proved that our methods can significantly improve the DW image quality. Finally, transcranial ultrasound imaging collected through temporal bone were presented and analyzed. The ultrasound frequency used in this study ranges from 2 MHz to 4 MHz, centered at 2.8 MHz. Since the wavefront was offset and distorted after passing through temporal bone, the image quality will be slightly degraded. Even then, it was demonstrated that these adaptive algorithms can significantly improve the transcranial image quality, especially the image contrast.

Coherence Plane-Wave Compounding with Angle Coherence Factor for Ultrafast Ultrasound Imaging.

Ultrafast ultrasound imaging with plane wave transmission has been a promising technique to image moving objects, however, implies compromises among resolution, contrast and sensitivity. Coherence plane-wave compounding (CPWC) can balance the image quality and frame rate. The image quality, especiallyin terms of the suppression of artifacts stemmed from side lobes, is greatly comprised by reducing the number of the tilted plane-waves. However, in some special scenarios, such as tracking shear wave propagation inside soft tissue, and imaging the complex blood flow, it's better to keep a very high frame rate. How to realize a B-mode image of equivalent quality to the standard focused approach at a very high frame rate? Here we proposed a new imaging framework by combining CPWC with angle coherence factor. The B-mode images from simulation, experimental phantoms demonstrated that our proposed methodology greatly suppressed the side-lobes artifacts by around 20 dB compared with CPWC imaging, while the image quality, especially lateral resolution and contrast kept equivalent.