Automated solid-phase synthesis of metabolically stabilized triazolo-peptidomimetics

The use of 1,4-disubstituted 1,2,3-triazoles as trans-amide bond surrogates has become an important tool for the synthesis of metabolically stabilized peptidomimetics. These heterocyclic bioisosters are generally incorporated into the peptide backbone by applying a diazo-transfer reaction followed by CuAAC (click chemistry) with an α-amino alkyne. Even though the manual synthesis of backbone-modified triazolo-peptidomimetics has been reported by us and others, no procedure has yet been described for an automated synthesis using peptide synthesizers. In order to efficiently adapt these reactions to an automated setup, different conditions were explored, putting special emphasis on the required long-term stability of both the diazo-transfer reagent and the Cu(I) catalyst in solution. ISA·HCl is the reagent of choice to accomplish the diazo-transfer reaction; however, it was found instable in DMF, the most commonly used solvent for SPPS. Thus, an aqueous solution of ISA·HCl was used to prevent its degradation over time, and the composition in the final diazo-transfer reaction was adjusted to preserve suitable swelling conditions of the resins applied. The CuAAC reaction was performed without difficulties using [Cu (CH₃CN)₄]PF₆ as a catalyst and TBTA as a stabilizer to prevent oxidation to Cu(II). The optimized automated two-step procedure was applied to the synthesis of structurally diverse triazolo-peptidomimetics to demonstrate the versatility of the developed methodology. Under the optimized conditions, five triazolo-peptidomimetics (8–5 amino acid residues) were synthesized efficiently using two different resins. Analysis of the crude products by HPLC-MS revealed moderate to good purities of the desired triazolo-peptidomimetics (70–85%). The synthesis time ranged between 9 and 12.5 h.     

Dynamic concentration challenges for biosensor characterization

A method and apparatus are described for characterization of the steady state and dynamic response of biosensors. The apparatus produces a steady stream of homogeneously mixed analyte whose concentration can be fixed at discrete values or varied continously. The device is ideally suited for continously operated biosensors, but is also effective for biosensors that operate in discrete sampling modes. The system permits simultaneous testing of several sensors and determination of the accuracy, precision and repeatability of sensor response. The characteristics of this testing apparatus were validated with ferrocyanide and glucose as indicators. As an example of use of the apparatus, concentration ramps were created and used to complement conventional step changes for characterizing an implantable glucose sensor. The ramp rate can be adjusted easily by scaling the apparatus to simulate the rate of concentration change anticipated during actual monitoring situations.     

运动红细胞形变结构的图象自动定量分析的研究

建立了运动目标图象自动分析ＭＣＥＬＬ应用软件和方法,并用于微血管中运动红细胞变形参数的定量分析。该图象处理主要特征功能包括压缩数字图象的灰度窗口设置,运动目标的抽取,形态相似性判决和运动目标的跟踪。同时给出运动图象处理的主要算式、动态曲线和应用结果。此外,还抽提和录取在微血管中红细胞快速运动情况下,所呈现拖鞋形的二值图象。研究结果证明红细胞具有可变性生理功能,这对于心脑血管的发病机理、早期病变的预测和有关药物疗效的研究有着十分重要的意义。     

Escher: A new docking procedure applied to the reconstruction of protein tertiary structure

Evaluation of Surface Complementarity, Hydrogen bonding, and Electrostatic interaction in molecular Recognition (ESCHER) is a new docking procedure consisting of three modules that work in series. The first module evaluates the geometric complementarity and produces a set of rough solutions for the docking problem. The second module identifies molecular collisions within those solutions, and the third evaluates their electrostatic complementarity. We describe the algorithm and its application to the docking of cocrystallized protein domains and unbound components of protein-protein complexes. Furthermore, ESCHER has been applied to the reassociation of secondary and supersecondary structure elements. The possibility of applying a docking method to the problem of protein structure prediction is discussed. Proteins 28:556–567, 1997. © 1997 Wiley-Liss, Inc.     

Phenotype analysis of cultivation processes via unsupervised machine learning: Demonstration for Clostridium pasteurianum

A novel approach of phenotype analysis of fermentation‐based bioprocesses based on unsupervised learning (clustering) is presented. As a prior identification of phenotypes and conditional interrelations is desired to control fermentation performance, an automated learning method to output reference phenotypes (defined as vector of biomass‐specific rates) was developed and the necessary computing process and parameters were assessed. For its demonstration, time series data of 90 Clostridium pasteurianum cultivations were used which feature a broad spectrum of solventogenic and acidogenic phenotypes, while 14 clusters of phenotypic manifestations were identified. The analysis of reference phenotypes showed distinct differences, where potential conditionalities were exemplary isolated. Further, cluster‐based balancing of carbon and ATP or the use of reference phenotypes as indicator for bioprocess monitoring were demonstrated to highlight the perks of this approach. Overall, such analysis depends strongly on the quality of the data and experimental validations will be required before conclusions. However, the automated, streamlined and abstracted approach diminishes the need of individual evaluation of all noisy dataset and showed promising results, which could be transferred to strains with comparably wide‐ranging phenotypic manifestations or as indicators for repeated bioprocesses with clearly defined target.     

Studies on Image Automated Analysis and Recognition of Plant Chromosome

A plant chromosome image analysis CHROM. HUK software system has been developed in the light of the theory of image analysis and recognition and applied in karyotype automated analysis of Xizang wild barley (Hordeum agriocrithon Aberg var. nigrum) and Cupressus gigantea Cheng et L. K. Fu. The main features of automated analysis include the pre-processing of chromosome image, determinating peak-valley threshold of chromosome, separation of overlapping chromosome, evaluation of centre line of chromosome,limit erotion recognition of position of centromere and second constric, extraction of characterictic parameters of chromosome and karyotype analysis. The vast amount of data obtained could be stored, operated and used for further statistical analysis. According to the estimation in the 95% confidence interval and the tree type sort, the chromosome were paired and sorted. In the meantime, a karyogram and an idiogram of karyotype were generated automatically through computerization.     

Automated Flight Test and System Identification for Rotary Wing Small Aerial Platform Using Frequency Responses Analysis

This paper proposes an autopilot system that can be used to control the small scale rotorcraft during the flight test for linear-frequency-domain system identification. The input frequency-sweep is generated automatically as part of the autopilot control command. Therefore the bandwidth coverage and consistency of the frequency-sweep are guaranteed to produce high quality data for system identification. Beside that, we can set the safety parameters during the flight test （maximum roll/pitch value, minimum altitude, etc.） so the safety of the whole flight test is guaranteed. This autopilot system is validated using hardware in the loop simulator for hover flight condition.     

Automated docking of flexible ligands: Applications of autodock

AutoDock is a suite of C programs used to predict the bound conformations of a small, flexible ligand to a macromolecular target of known structure. The technique combines simulated annealing for conformation searching with a rapid grid-based method of energy evaluation. This paper reviews recent applications of the technique and describes the enhancements included in the current release.     

A method for accurate detection of translocation junctions in ewing family of tumors

The Ewing family of tumors (ET) generally contain translocations involving the EWS gene and the FLI or ERG genes. Identification of the translocation confirms the diagnosis of ET. Currently, diagnosis of the translocation is made by several methods. In general, these methods require different primer sets for amplifying different translocations and subsequent efforts to identify the amplified product. The need to employ different sets of primers to amplify different translocation junctions presents some limitations. We have developed a method based on PCR with consensus primers followed by direct automated sequencing of the amplified product. With this method we have correctly determined known as well as unknown ET-associated EWS-FLI and EWS-ERG translocations in appropriate specimens. Use of our consensus primers eliminates the need for separate PCRs to amplify EWS-FLI and EWS-ERG translocation junctions, and because direct sequencing is used for confirming the identity of the amplification product, the accuracy of detection becomes 100%. The method might also accurately diagnose ET-associated translocations other than EWS-FLI and EWS-ERG translocations.     

Description of heterotrophic bacteria occurring in paper mills and paper products

AIMS: To isolate aerobic mesophilic bacilli and thermophilic bacteria from different paper mill samples and to evaluate their potential harmfulness. METHODS AND RESULTS: A total of 109 mesophilic and 68 thermophilic isolates were purified and characterized by automated ribotyping and partial 16S rDNA sequencing. The mesophilic isolates belonged to the genera Bacillus (13 taxa), Brevibacillus (three taxa) and Paenibacillus (five taxa). The thermophilic bacteria represented seven taxa of Bacillus, Geobacillus or Paenibacillus, four of proteobacteria and one of actinobacteria. The most frequently occurring bacteria were Bacillus cereus, B. licheniformis, Pseudoxanthomonas taiwanensis and bacteria closely related to Paenibacillus stellifer, P. turicensis or Leptothrix sp. One mill was contaminated throughout with bacteria of a novel mesophilic genus most closely related to Brevibacillus centrosporus and another with bacteria of a novel thermophilic genus most closely related to Hydrogenophilus thermoluteolus. One B. cereus isolate producing haemolytic diarrhoeal enterotoxin was detected and all the tested B. licheniformis isolates produced a metabolite toxic to boar sperm cells. CONCLUSIONS: The bacilli and thermophilic bacteria isolated represent species which should not present occupational hazards in paper mill environments. The most harmful bacterium detected was B. licheniformis and potentially also B. cereus. SIGNIFICANCE AND IMPACT OF THE STUDY: Knowledge of the microbial diversity in a paper mill provides a rational basis for development of an effective controlling programme. A database constructed from the fingerprints generated using automated ribotyping helps to identify and trace the contamination routes of bacteria occurring in paper mills.