Expression, Purification and Properties of Shikimate Dehydrogenase from Mycobacterium Tuberculosis

Zhang, Xuelian,Zhang, Shunbao,Hao, Fang,Lai, Xuhui,Yu, Haidong,Huang, Yishu,Wang, Honghai Korean Society for Biochemistry and Molecular Biol 2005 Journal of biochemistry and molecular biology Vol.38 No.5

Tuberculosis, caused by Mycobacterium tuberculosis, continues to be one of the main diseases to mankind. It is urgent to discover novel drug targets for appropriate antimicrobial agents against this human pathogen. The shikimate pathway is onsidered as an attractive target for the discovery of novel antibiotics for its essentiality in bacteria and absence in mammalian cells. The Mycobacterium tuberculosis aroE-encoded shikimate dehydrogenase was cloned, expressed and purified. Sequence alignment analysis shows that shikimate dehydrogenase of Mycobacterium tuberculosis exhibit the pattern of G-X-(N/S)-V-(T/S)-X-PX-K, which is highly conserved within the shikimate dehydrogenase family. The recombinant shikimate dehydrogenase spectrum determined by CD spectroscopy showed that the percentages for $\alpha$-helix, $\beta$-sheet, $\beta$-turn, and random coil were 29.2%, 9.3%, 32.7%, and 28.8%, respectively. The enzymatic characterization demonstrates that it appears to be fully active at pH from 9.0 to 12, and temperature $63^{\circ}C$. The apparent Michaelis constant for shikimic acid and $NADP^+$ were calculated to be about $29.5\;{\mu}M$ and $63\;{\mu}M$. The recombinant shikimate dehydrogenase catalyzes the substrate in the presence of $NADP^+$ with an enzyme turnover number of $399\;s^{-1}$. Zymological studies suggest that the cloned shikimate dehydrogenase from M. tuberculosis has a pretty activity, and the work should help in the discovery of enzyme inhibitors and further of possible antimicrobial agents against Mycobacterium tuberculosis.

Proportional-Integral Controller for Stabilization of Second-Order Delay Processes

Honghai Wang,Yilian Zhang,Jian-Chang Liu,Feisheng Yang 제어·로봇·시스템학회 2014 International Journal of Control, Automation, and Vol.12 No.6

This paper considers the problem of determining the complete stabilizing set of proportional-integral (PI) controllers for a second-order process with time delay by employing a version of the Hermite-Biehler theorem applicable to quasipolynomials. With the poles of open-loop system being complex, we first provide the result to find the admissible range of the proportional gain. Then by choosing a fixed proportional gain in this range, we can ascertain the complete region of integral gain which can stabilize the second-order delay process. Similarly, the result for the case of open-loop real poles is also obtained. It is mentioned that the condition to obtain the parameter set for stabilizing the given plant is sufficient and necessary.

Expression, Purification and Properties of Shikimate Dehydrogenasefrom Mycobacterium Tuberculosis

Xuhui Lai,Honghai Wang,Xuelian Zhang,Shunbao Zhang,Fang Hao,Haidong Yu,Yishu Huang 한국생화학분자생물학회 2005 BMB Reports Vol.38 No.5

Cell Surface Display of Four Types of Solanum nigrum Metallothionein on Saccharomyces cerevisiae for Biosorption of Cadmium

( Qinguo Wei ),( Honghai Zhang ),( Dongge Guo ),( Shisheng Ma ) 한국미생물 · 생명공학회 2016 Journal of microbiology and biotechnology Vol.26 No.5

We displayed four types of Solanum nigrum metallothionein (SMT) for the first time on the surface of Saccharomyces cerevisiae using an α-agglutinin-based display system. The SMT genes were amplified by RT-PCR. The plasmid pYES2 was used to construct the expression vector. Transformed yeast strains were confirmed by PCR amplification and custom sequencing. Surface-expressed metallothioneins were indirectly indicated by the enhanced cadmium sorption capacity. Flame atomic absorption spectrophotometry was used to examine the concentration of Cd2+ in this study. The transformed yeast strains showed much higher resistance ability to Cd2+ compared with the control. Strikingly, their Cd2+ accumulation was almost twice as much as that of the wild-type yeast cells. Furthermore, surface-engineered yeast strains could effectively adsorb ultra-trace cadmium and accumulate Cd2+ under a wide range of pH levels, from 3 to 7, without disturbing the Cu2+ and Hg2+. Four types of surfaceengineered Saccharomyces cerevisiae strains were constructed and they could be used to purify Cd2+-contaminated water and adsorb ultra-trace cadmium effectively. The surface-engineered Saccharomyces cerevisiae strains would be useful tools for the bioremediation and biosorption of environmental cadmium contaminants.

Biosynthesis of Isoprenoids: Characterization of a Functionally Active Recombinant 2-C-methyl-D-erythritol 4-phosphate Cytidyltransferase (IspD) from Mycobacterium tuberculosis H37Rv

Shi, Wenjun,Feng, Jianfang,Zhang, Min,Lai, Xuhui,Xu, Shengfeng,Zhang, Xuelian,Wang, Honghai Korean Society for Biochemistry and Molecular Biol 2007 Journal of biochemistry and molecular biology Vol.40 No.6

Tuberculosis, caused by Mycobacterium tuberculosis, continues to be one of the leading infectious diseases to humans. It is urgent to discover novel drug targets for the development of antitubercular agents. The 2-C-methyl-Derythritol-4-phosphate (MEP) pathway for isoprenoid biosynthesis has been considered as an attractive target for the discovery of novel antibiotics for its essentiality in bacteria and absence in mammals. MEP cytidyltransferase (IspD), the third-step enzyme of the pathway, catalyzes MEP and CTP to form 4-diphosphocytidyl-2-C-methylerythritol (CDP-ME) and PPi. In the work, ispD gene from M. tuberculosis H37Rv (MtIspD) was cloned and expressed. With N-terminal fusion of a histidine-tagged sequence, MtIspD could be purified to homogeneity by one-step nickel affinity chromatography. MtIspD exists as a homodimer with an apparent molecular mass of 52 kDa. Enzyme property analysis revealed that MtIspD has high specificity for pyrimidine bases and narrow divalent cation requirements, with maximal activity found in the presence of CTP and $Mg^{2+}$. The turnover number of MtIspD is $3.4 s^{-1}$. The Km for MEP and CTP are 43 and $92{\mu}M$, respectively. Furthermore, MtIspD shows thermal instable above $50^{\circ}C$. Circular dichroism spectra revealed that the alteration of tertiary conformation is closely related with sharp loss of enzyme activity at higher temperature. This study is expected to help better understand the features of IspD and provide useful information for the development of novel antibiotics to treat M. tuberculosis.

J-Sim: a simulation and emulation environment for wireless sensor networks

Sobeih, A.,Hou, J.C.,Lu-Chuan Kung,Ning Li,Honghai Zhang,Wei-Peng Chen,Hung-Ying Tyan,Hyuk Lim IEEE 2006 IEEE wireless communications Vol.13 No.4

<P>Wireless sensor networks have gained considerable attention in the past few years. They have found application domains in battlefield communication, homeland security, pollution sensing, and traffic monitoring. As such, there has been an increasing need to define and develop simulation frameworks for carrying out high-fidelity WSN simulation. In this article we present a modeling, simulation, and emulation framework for WSNs in J-Sim - an open source, component-based compositional network simulation environment developed entirely in Java. This framework is built on the autonomous component architecture and extensible internetworking framework of J-Sim, and provides an object-oriented definition of target, sensor, and sink nodes, sensor and wireless communication channels, and physical media such as seismic channels, mobility models, and power models (both energy-producing and energy-consuming components). Application-specific models can be defined by subclassing classes in the simulation framework and customizing their behaviors. We also include in J-Sim a set of classes and mechanisms to realize network emulation. We demonstrate the use of the proposed WSN simulation framework by implementing several well-known localization, geographic routing, and directed diffusion protocols, and perform performance comparisons (in terms of the execution time incurred and memory used) in simulating WSN scenarios in J-Sim and ns-2. The simulation study indicates the WSN framework in J-Sim is much more scalable than ns-2 (especially in memory usage). We also demonstrate the use of the WSN framework in carrying out real-life full-fledged Future Combat System (FCS) simulation and emulation</P>