[Synthesis of L-2-aminobutyric acid by leucine dehydrogenase coupling with an NADH regeneration system].

In this study, Escherichia coli BL21 (DE3) was used as the host to construct 2 recombinant E. coli strains that co-expressed leucine dehydrogenase (LDH, Bacillus cereus)/formate dehydrogenase (FDH, Ancylobacter aquaticus), or leucine dehydrogenase (LDH, Bacillus cereus)/alcohol dehydrogenase (ADH, Rhodococcus), respectively. L-2-aminobutyric acid was then synthesized by L-threonine deaminase (L-TD) with LDH-FDH or LDH-ADH by coupling with two different NADH regeneration systems. LDH-FDH process and LDH-ADH process were optimized and compared with each other. The optimum reaction pH of LDH-FDH process was 7.5, and the optimum reaction temperature was 35 °C. After 28 h, the concentration of L-2-aminobutyric acid was 161.8 g/L with a yield of 97%, when adding L-threonine in batches for controlling 2-ketobutyric acid concentration less than 15 g/L and using 50 g/L ammonium formate, 0.3 g/L NAD+, 10% LDH-FDH crude enzyme solution (V/V) and 7 500 U/L L-TD. The optimum reaction pH of LDH-ADH process was 8.0, and the optimum reaction temperature was 35 °C. After 24 h, the concentration of L-2-aminobutyric acid was 119.6 g/L with a yield of 98%, when adding L-threonine and isopropanol (1.2 times of L-threonine) in batches for controlling 2-ketobutyric acid concentration less than 15 g/L, removing acetone in time and using 0.3 g/L NAD⁺, 10% LDH-ADH crude enzyme solution (V/V) and 7 500 U/L L-TD. The process and results used in this paper provide a reference for the industrialization of L-2-aminobutyric acid.

Efficient biocatalyst of L-DOPA with Escherichia coli expressing a tyrosine phenol-lyase mutant from Kluyvera intermedia.

L-DOPA (L-dihydroxyphenylalanine) is a promising drug for Parkinson's disease and thereby has a growing annual demand. Tyrosine phenol-lyase (TPL)-based catalysis is considered to be a low-cost yet efficient route for biosynthesis of L-DOPA. TPL is a tetrameric enzyme that catalyzes the synthesis of L-DOPA from pyrocatechol, sodium pyruvate, and ammonium acetate. The implementation of TPL for L-DOPA production has been hampered and the need for the most efficient TPL source with higher L-DOPA production and substrate conversion rate is prevailing. This study involves identifying a novel TPL from Kluyvera intermedia (Ki-TPL) and displayed a robust expression in Escherichia coli. The recombinant strain YW000 carrying Ki-TPL proved strong catalytic activity with a highest L-DOPA yield compared with 16 other TPLs from different organisms. With a further aim to improve this efficiency, random mutagenesis of Ki-TPL was performed and a mutant namely YW021 was obtained. The whole cells of YW021 as biocatalyst yielded 150.4 g L-1 of L-DOPA with a 99.99 % of pyrocatechol conversion at the optimum condition of pH 8.0 at 25 °C, which is the highest level reported to date. Further, the homology modeling and structural analysis revealed the mutant residues responsible for the extensive L-DOPA biosynthesis.

Design guidelines for radio frequency non-contact vital sign detection.

The design guidelines for non-contact vital sign detection are presented in this paper. Firstly, the choice of radio frequency and antenna beamwidth is demonstrated by numerical simulation based on ray-tracing technique and spectrum analysis. Guided by the numerical results of the first part, three typical radar architectures are recently built in one of the ISM bands (the 5.8 GHz band). The characteristics of each architecture are demonstrated and compared based on measurement results. Experiments show that direct-conversion non-quadrature architecture has DC offset and null detection point problem; direct-conversion quadrature architecture can eliminate null detection point but lead to complexity and higher power consumption in effectively combining two quadrature channels. Double-sideband indirect-conversion with simpler radio architecture and less baseband requirement can effectively alleviate the DC offset and null detection point problems.

Robust overnight monitoring of human vital signs by a non-contact respiration and heartbeat detector.

A positive answer is given to the question intrigued by our previous work reported in EMBC 2005: whether it is possible for a non-contact physiological movement detector to detect vital signs from four sides of a human body. In addition to the proof from measured data, theoretical analysis confirms the surprising advantage of detection from the back of the body. Based on this observation, a non-contact system was set up to perform overnight monitoring of vital signs using low power radio waves. Measurement data is presented and analyzed. The challenges and key technologies that improved the performance of our system for overnight monitoring are discussed.

A ka-band low power Doppler radar system for remote detection of cardiopulmonary motion.

A low power Ka-band Doppler radar that can detect human heartbeat and respiration signals is demonstrated. This radar system achieves better than 80% detection accuracy at the distance of 2-m with 16-μW transmitted power. Indirect-conversion receiver architecture is chosen to reduce the DC offset and 1/f noise that can degrade signal-to-noise ratio and detection accuracy. In addition, the radar has also demonstrated the capability of detecting acoustic signals.

Intramolecular dimerization is required for the chlamydia-secreted protease CPAF to degrade host transcriptional factors.

We previously identified a chlamydial protein designated CPAF (chlamydia protease/proteasome-like activity factor) that is secreted into host cell cytosol for degrading host transcription factors required for major histocompatibility complex antigen expression. Here we report that CPAF, synthesized as a 70-kDa proprotein, is processed into two fragments (designated CPAFn and CPAFc) to form intramolecular dimers that are much more stable than the naïve CPAF. Precipitation with antibodies that recognized CPAF dimers removed the proteolytic activity responsible for degrading host transcription factor RFX5 from chlamydia-infected host cell cytosol, while precipitation with antibodies that recognized free CPAF fragments alone did not remove this activity. Separation of CPAFn from CPAFc resulted in a loss of proteolytic activity. Furthermore, neither expressed full-length CPAF that was not processed nor coexpressed CPAFn and CPAFc fragments that failed to form dimers degraded RFX5. These observations demonstrate that intramolecular dimerization is required for CPAF to degrade host transcription factors, a strategy that is utilized by an obligate intracellular bacterial species to evade host defenses.