Statistical optimization of culture conditions for bacterial cellulose production using Box-Behnken design

Culture conditions in a jar fermentor for bacterial cellulose (BC) production from A. xylinum BPR2001 were optimized by statistical analysis using Box-Behnken design. Response surface methodology was used to predict the levels of the factors, fructose (X1), corn steep liquor (CSL) (X2), dissolved oxygen (DO) (X3), and agar concentration (X4). Total 27 experimental runs by combination of each factor were carried out in a 10-L jar fermentor, and a three-dimensional response surface was generated to determine the effect of the factors and to find out the optimum concentration of each factor for maximum BC production and BC yield. The fructose and agar concentration highly influenced the BC production and BC yield. However, the optimum conditions according to changes in CSL and DO concentrations were predicted at almost central values of tested ranges. The predicted results showed that BC production was 14.3 g/L under the condition of 4.99% fructose, 2.85% CSL, 28.33% DO, and 0.38% agar concentration. On the other hand, BC yield was predicted in 0.34 g/g under the condition of 3.63% fructose, 2.90% CSL, 31.14% DO, and 0.42% agar concentration. Under optimized culture conditions, improvement of BC production and BC yield were experimentally confirmed, which increased 76% and 57%, respectively, compared to BC production and BC yield before optimizing the culture conditions.     

Parameters Effects Evaluation of Microbial Strengthening of Sandy Soils in Mixing Experiments Using Taguchi Methodology

The effects of experimental parameters including soil type, curing duration, inoculum size, and biomass and nutrients concentration on soil strengthening due to calcite precipitation by Sporosarcina pasteurii PTCC 1645 were investigated. The laboratory-scale mixing experiments on remolded samples were designed by the Taguchi method. Soil type proved to be the most incorporating factor, followed by curing time and nutrient concentration. The main effect and the interactions of the parameters were presented and the optimal conditions were obtained. This suggests the importance of local conditions including soil type on any future large-scale, in situ application.     

Construct optimization for protein NMR structure analysis using amide hydrogen/deuterium exchange mass spectrometry

Disordered or unstructured regions of proteins, while often very important biologically, can pose significant challenges for resonance assignment and three‐dimensional structure determination of the ordered regions of proteins by NMR methods. In this article, we demonstrate the application of ¹H/²H exchange mass spectrometry (DXMS) for the rapid identification of disordered segments of proteins and design of protein constructs that are more suitable for structural analysis by NMR. In this benchmark study, DXMS is applied to five NMR protein targets chosen from the Northeast Structural Genomics project. These data were then used to design optimized constructs for three partially disordered proteins. Truncated proteins obtained by deletion of disordered N‐ and C‐terminal tails were evaluated using ¹H‐¹⁵N HSQC and ¹H‐¹⁵N heteronuclear NOE NMR experiments to assess their structural integrity. These constructs provide significantly improved NMR spectra, with minimal structural perturbations to the ordered regions of the protein structure. As a representative example, we compare the solution structures of the full length and DXMS‐based truncated construct for a 77‐residue partially disordered DUF896 family protein YnzC from Bacillus subtilis, where deletion of the disordered residues (ca. 40% of the protein) does not affect the native structure. In addition, we demonstrate that throughput of the DXMS process can be increased by analyzing mixtures of up to four proteins without reducing the sequence coverage for each protein. Our results demonstrate that DXMS can serve as a central component of a process for optimizing protein constructs for NMR structure determination. Proteins 2009. © 2009 Wiley‐Liss, Inc.     

Oxidant and SDS-stable alkaline protease from Bacillus clausii I-52: production and some properties

AIMS: An investigation was carried out on an oxidative and SDS-stable alkaline protease secreted by Bacillus clausii of industrial significance. METHODS AND RESULTS: Maximum enzyme activity was produced when the bacterium was grown in the medium containing (g l-1): soyabean meal, 15; wheat flour, 10; liquid maltose, 25; K2HPO4, 4; Na2HPO4, 1; MgSO4.7H2O, 0.1; Na2CO3, 6. The enzyme has an optimum pH of around 11 and optimum temperature of 60 degrees C. The alkaline protease showed extreme stability towards SDS and oxidizing agents, which retained its activity above 75 and 110% on treatment for 72 h with 5% SDS and 10% H2O2, respectively. Inhibition profile exhibited by phenylmethylsulphonyl fluoride suggested that the protease from B. clausii belongs to the family of serine proteases. CONCLUSIONS: Bacillus clausii produced high levels of an extracellular protease having high stability towards SDS and H2O2. SIGNIFICANCE AND IMPACT OF THE STUDY: The alkaline protease from B. clausii I-52 is significant for an industrial perspective because of its ability to function in broad pH and temperature ranges in addition to its tolerance and stability in presence of an anionic surfactant, like SDS and oxidants like peroxides and perborates. The enzymatic properties of the protease also suggest its suitable application as additive in detergent formulations.     

Protein structure preference, tRNA copy number, and mRNA stem/loop content

From statistical analyses of protein sequences for humans and Escherichia coli we found that the messenger RNA segment of m-codons (for m=2 to 6) with average high tRNA copy number (TCN) (larger than approximately 10.5 for humans or approximately 1.95 for E. coli) preferably code for the alpha helix and that with low TCN (smaller than approximately 7.5 for humans or approximately 1.7 for E. coli) preferably code for coil. Between them there is an intermediate region without correlation to structure preference. For the beta strand the preference/ avoidance tendency is not obvious. All strong preference-modes of TCN for protein secondary structures have been deduced. The mutual interaction between two factors--protein secondary structural type and codon TCN--is tested by F distribution. A phenomenological model on the relation between structure preference and translational efficiency or accuracy is proposed. It is pointed out that the structure preference of codons is related to the distribution of mRNA stem/loop content in three TCN regions.     

Random field models for fitness landscapes

 In many cases fitness landscapes are obtained as particular instances of random fields by randomly assigning a large number of parameters. Models of this type are often characterized reasonably well by their covariance matrices. We characterize isotropic random fields on finite graphs in terms of their Fourier series expansions and investigate the relation between the covariance matrix of the random field model and the correlation structure of the individual landscapes constructed from this random field. Correlation measures are a good characteristic of “rugged landscapes” models as they are closely related to quantities like the number of local optima or the length of adaptive walks. Our formalism suggests to approximate landscape with known autocorrelation function by a random field model that has the same correlation structure. Received: 10 November 1995 / Revised version: 19 February 1996     

Optimization of Operating Temperature for Continuous Immobilized Glucose Isomerase Reactor with Pseudo Linear Kinetics

In this work, the optimal operating temperature for the enzymatic isomerization of glucose to fructose using a continuous immobilized glucose isomerase packed bed reactor is studied. This optimization problem describing the performance of such reactor is based on reversible pseudo linear kinetics and is expressed in terms of a recycle ratio. The thermal deactivation of the enzyme as well as the substrate protection during the reactor operation is considered. The formulation of the problem is expressed in terms of maximization of the productivity of fructose. This constrained nonlinear optimization problem is solved using the disjoint policy of the calculus of variations. Accordingly, this method of solution transforms the nonlinear optimization problem into a system of two coupled nonlinear ordinary differential equations (ODEs) of the initial value type, one equation for the operating temperature profile and the other one for the enzyme activity. The ODE for the operating temperature profile is dependent on the recycle ratio, operating time period, and the reactor residence time as well as the kinetics of the reaction and enzyme deactivation. The optimal initial operating temperature is selected by solving the ODEs system by maximizing the fructose productivity. This results into an unconstrained one‐dimensional optimization problem with simple bounds on the operating temperature. Depending on the limits of the recycle ratio, which represents either a plug flow or a mixed flow reactor, it is found that the optimal temperature of operation is characterized by an increasing temperature profile. For higher residence time and low operating periods the residual enzyme activity in the mixed flow reactor is higher than that for the plug flow reactor, which in turn allows the mixed flow reactor to operate at lower temperature than that of the plug flow reactor. At long operating times and short residence time, the operating temperature profiles are almost the same for both reactors. This could be attributed to the effect of substrate protection on the enzyme stability, which is almost the same for both reactors. Improvement in the fructose productivity for both types of reactors is achieved when compared to the constant optimum temperature of operation. The improvement in the fructose productivity for the plug flow reactor is significant in comparison with the mixed flow reactor.     

华中五味子ISSR-PCR反应体系优化及引物筛选

系统研究了华中五味子ISSR PCR反应体系中的主要影响因子,确立华中五味子ISSR-PCR最适反应体系,并筛选出12条有效引物。单因子实验结果显示,华中五味子ISSR-PCR反应体系中各主要成分的适宜浓度范围为,Mg²⁺ 1.50~3.50 mmol·L^-1,dNTPs 0.10~0.35 mmol·L^-1,引物0.25~0.60 μmol·L^-1,Taq酶0.50~1.50 U。4因子3水平正交实验确立了最适反应体系,即20 μL体系中包含2.50 mmol·L^-1 Mg²⁺、0.20 mmol·L^-1 dNTPs、0.25 μmol·L^-1引物、1.50 U Taq酶、60 ng DNA模板、2.50 μL 10×PCR Buffer。本研究为华中五味子种质资源的评估及遗传多样性分析奠定基础。     

PACS的高级预防维护技术

目的 通过讨论PACS高级预防维护技术,供同他人在日常维护过程中有一定地借鉴作用.方法 从PACS数据库性能检测与优化,存储设备检测及PACS各应用软件模块三方面进行阐述.结果 经过医院一线安装维护人员和PACS厂家多年实践的总结,拥有一定的可参考依据.结论 PACS的高级预防维护是一个复杂的系统工程,包括一般维护和高级维护在内的实践理论才刚刚起步,科学的、规范化的预防维护需要不断地完善和发展.