bluuebunny/crossref_metadata_2025_split huggingface dataset:
Overcoming the thermodynamic limitation in asymmetric hydrogen transfer reactions catalyzed by whole cells
Authors: Goldberg Katja , Edegger Klaus , Kroutil Wolfgang , Liese Andreas 
Published: June 2006

AbstractWhole lyophilized cells of an Escherichia coli overexpressing the alcohol dehydrogenase (ADH‐‘A’) from Rhodococcus ruber DSM 44541 were used for the asymmetric reduction of ketones to secondary alcohols. The recycling of the required nicotinamide cofactor (NADH) was achieved in a coupled‐substrate process. In the course of the reaction the ketone is reduced to the alcohol and the hydrogen donor 2‐propanol is oxidized to acetone by one enzyme. This leads to a thermodynamic equilibrium between all four components determining the maximum achievable conversion. To overcome this limitation an in situ product removal technique (ISPR) for the application with whole cells was developed. In this method the most volatile compound is separated from the reaction vessel by an air flow resulting in a shift of the equilibrium towards the desired secondary alcohol. The so‐called stripping process represents a simple and efficient method to overcome the thermodynamic limitation in biocatalytic reactions. Employing this method, the conversion of selected biotransformations was increased up to completeness. © 2006 Wiley Periodicals, Inc.
Characterization of sol‐gel entrapped chlorophyllase
Authors: Yi Yunyu , Kermasha Selim , Neufeld Ronald 
Published: June 2006

AbstractImmobilization of membrane proteins remains a challenge compared to soluble proteins. The membrane protein—chlorophyllase was successful entrapped in tetramethoxysilane (TMOS)‐based sol‐gel in the presence of lipid. Activity was examined against mixing rate, incubation temperature, time, substrate, acetone, and canola oil concentration. The external mass transfer of chlorophyll is not the rate‐limiting step at higher mixing rates. Stability against temperature and acetone as denaturant was enhanced. In spite of the fact that an initial reaction lag phase was observed, 20% more chlorophyll was hydrolyzed, compared to reaction with free enzyme by the end of a 12 h assay. The initial lower activity demonstrated by entrapped chlorophyllase is likely due to the diffusion resistance of chlorophyll into and within the entrapment matrix. This hypothesis was substantiated by a low diffusion coefficient on the order of 10−14 m2/s obtained for chlorophyll in nanoporous sol‐gel particles. Pore size distribution of nanoporous wet TMOS‐based sol‐gel with or without protein was determined by thermoporometry. The change in pore morphology upon doping with chlorophyllase suggests that protein acts as a template during the sol‐gel process. © 2006 Wiley Periodicals, Inc.
A Unique Mechanism for Methyl Ester Formation via an Amide Intermediate Found in Myxobacteria
Authors: Müller Inga , Weinig Stefan , Steinmetz Heinrich , Kunze Birgitte , Veluthoor Sheeba , Mahmud Taifo , Müller Rolf 
Published: June 2006

AbstractSecondary metabolism involves a broad diversity of biochemical reactions that result in a wide variety of biologically active compounds. Terminal amide formation during the biosynthesis of the myxobacterial electron‐transport inhibitor, myxothiazol, was analyzed by heterologous expression of the unique nonribosomal‐peptide synthetase, MtaG, and incubation with a synthesized substrate mimic. These experiments provide evidence that the terminal amide is formed from a carrier protein‐bound myxothiazol acid that is thioesterified to MtaF. This intermediate is transformed to an amide by extension with glycine and subsequent oxidative cleavage by MtaG. The final steps of melithiazol assembly involve a highly similar protein‐bound intermediate (attached to MelF, a homologue of MtaF), which is transformed to an amide by MelG (homologue of MtaG). In this study, we also show that the amide moiety of myxothiazol A can be hydrolyzed in vivo to the formerly unknown free myxothiazol acid by heterologous expression of melJ in the myxothiazol producer Stigmatella aurantiaca DW4/3‐1. The methyltransferase MelK can finally methylate the acid to give rise to the methyl ester, which is produced as the final product in the melithiazol A biosynthetic pathway. These experiments clarify the role of MelJ and MelK during melithiazol assembly.
Resonance Structures of the Amide Bond: The Advantages of Planarity
Authors: Mujika Jon I. , Matxain Jon M. , Eriksson Leif A. , Lopez Xabier 
Published: June 2006

AbstractDelocalization indexes based on magnitudes derived from electron‐pair densities are demonstrated to be useful indicators of electron resonance in amides. These indexes, based on the integration of the two‐electron density matrix over the atomic basins defined through the zero‐flux condition, have been calculated for a series of amides at the B3LYP/6‐31+G* level of theory. These quantities, which can be viewed as a measure of the sharing of electrons between atoms, behave in concordance with the traditional resonance model, even though they are integrated in Bader atomic basins. Thus, the use of these quantities overcomes contradictory results from analyses of atomic charges, yet keeps the theoretical appeal of using nonarbitrary atomic partitions and unambiguously defined functions such as densities and pair densities. Moreover, for a large data set consisting of 24 amides plus their corresponding rotational transition states, a linear relation was found between the rotational barrier for the amide and the delocalization index between the nitrogen and oxygen atoms, indicating that this parameter can be used as an ideal physical‐chemical indicator of the electron resonance in amides.
An Experimental and Theoretical Investigation of Gas‐Phase Reactions of Ca2+with Glycine
Authors: Corral Inés , Mó Otilia , Yáñez Manuel , Salpin Jean‐Yves , Tortajada Jeanine , Moran Damian , Radom Leo 
Published: June 2006

AbstractThe gas‐phase reactions between Ca2+and glycine ([Ca(gly)]2+) have been investigated through the use of mass spectrometry techniques and B3‐LYP/cc‐pWCVTZ density functional theory computations. The major peaks observed in the electrospray MS/MS spectrum of [Ca(gly)]2+correspond to the formation of the [Ca,C,O2,H]+, NH2CH2+, CaOH+, and NH2CH2CO+fragment ions, which are produced in Coulomb explosion processes. The computed potential energy surface (PES) shows that not only are these species the most stable product ions from a thermodynamic point of view, but they may be produced with barriers lower than for competing processes. Carbon monoxide is a secondary product, derived from the unimolecular decomposition of some of the primary ions formed in the Coulomb explosions. In contrast to what is found for the reactions of Ca2+with urea ([Ca(urea)]2+), minimal unimolecular losses of neutral fragments are observed for the gas‐phase fragmentation processes of [Ca(gly)]2+, which is readily explained in terms of the topological differences between their respective PESs.
Diels–Alder Ligation of Peptides and Proteins
Authors: de Araújo Aline Dantas , Palomo Jose M. , Cramer Janina , Seitz Oliver , Alexandrov Kirill , Waldmann Herbert 
Published: June 2006

AbstractThe development of the Diels–Alder cycloaddition as a new method for the site‐specific chemoselective ligation of peptides and proteins under mild conditions is reported. Peptides equipped with a 2,4‐hexadienyl ester and an N‐terminal maleimide react in aqueous media to give cycloadducts in high yields and depending on the amino acid sequence with high stereoselectivity. Except for the cysteine SH group the transformation is compatible with all amino acid side chain functional groups. For ligation to proteins the hexadienyl group was attached to avidin and streptavidin noncovalently by means of complex formation with a biotinylated peptide or by covalent attachment of a hexadienyl ester‐containing label to lysine side chains incorporated into the proteins. Site‐specific attachment of the hexadienyl unit into a Rab protein was achieved by means of expressed protein ligation followed by protection of the generated cysteine SH by means of Ellman's reagent. The protein reacted with different maleimido‐modified peptides under mild conditions to give the fully functional cycloadducts in high yield. The results demonstrate that the Diels–Alder ligation offers an advantageous and technically straightforward new opportunity for the site‐specific equipment of peptides and proteins with further functional groups and labels. It proceeds under very mild conditions and is compatible with most functional groups found in proteins. Its combination with other ligation methods, in particular expressed protein ligation is feasible.
Individual cell‐based models of the spatial‐temporal organization of multicellular systems—Achievements and limitations
Authors: Galle J. , Aust G. , Schaller G. , Beyer T. , Drasdo D. 
Published: June 2006

AbstractComputational approaches of multicellular assemblies have reached a stage where they may contribute to unveil the processes that underlie the organization of tissues and multicellular aggregates. In this article, we briefly review and present some new results on a number of 3D lattice free individual cell‐based mathematical models of epithelial cell populations. The models we consider here are parameterized by bio‐physical and cell‐biological quantities on the level of an individual cell. Eventually, they aim at predicting the dynamics of the biological processes on the tissue level. We focus on a number of systems, the growth of cell populations in vitro, and the spatial‐temporal organization of regenerative tissues. For selected examples we compare different model approaches and show that the qualitative results are robust with respect to many model details. Hence, for the qualitative features and largely for the quantitative features many model details do not matter as long as characteristic biological features and mechanisms are correctly represented. For a quantitative prediction, the control of the bio‐physical and cell‐biological parameters on the molecular scale has to be known. At this point, slide‐based cytometry may contribute. It permits to track the fate of cells and other tissue subunits in time and validated the organization processes predicted by the mathematical models. © 2006 International Society for Analytical Cytology
Toward complete laser ablation of melanoma contaminant cells in a co‐culture outgrowth model via image cytometry
Authors: Shen Feimo , Price Jeffrey H. 
Published: June 2006

AbstractBackground:Contaminant cancer cells in autologous transplant tissue can cause relapse and the rates are unknown. A method capable of removing all contaminant cells with a high probability detected by cytomic analyses would be useful. Neither 100% cell purging nor techniques for measuring the probability of success have been developed. Here, we report a method for removing 100% of the cells under ideal staining conditions and quantify the probability of success.Methods:Laser ablation was combined with previously reported automated microscopy to purge contaminant cells and evaluate 100% ablation in a co‐culture model of prestained mouse melanoma cells mixed with mouse NIH‐3T3 cells. Melanoma passage efficiency was measured by: (1) micropipetting single cells into microtiter wells and (2) ablating all but one melanoma cell in co‐cultures.Results:(74 ± 5)% of single melanoma cells pipetted into microtiter plate wells divided at least once. With ablation of all but one contaminant cell in co‐cultures, melanoma dominated in (62 ± 8)% cultures in 21 days. With 100% ablation in six additional experiments, no melanoma outgrowth was observed, giving a >99.1% probability that all contaminant melanoma cells were purged.Conclusions:We successfully demonstrated a model for complete ablation within a defined probability using automated high‐content image cytometry with ideal staining conditions. The results show that the instrumentation is capable of delivering 100% ablation at a defined probability and establishes the basis for further studies with clinical models wherein pretherapeutic cytomic analyses of unique cellular expression and/or morphological characteristics will be key for contaminant cancer cell identification. © 2006 International Society for Analytical Cytology
Scanning cytometry with a LEAP: Laser‐enabled analysis and processing of live cells in situ
Authors: Szaniszlo Peter , Rose William A. , Wang Nan , Reece Lisa M. , Tsulaia Tamara V. , Hanania Elie G. , Elferink Cornelis J. , Leary James F. 
Published: June 2006

AbstractBackground:Scanning cytometry now has many of the features (and power) of multiparameter flow cytometry while keeping its own advantages as an imaging technology. Modern instruments combine capabilities of scanning cytometry with the ability to manipulate cells. A new technology, called LEAP™ (laser‐enabled analysis and processing), offers a unique combination of capabilities in cell purification and selective macromolecule delivery (optoinjection).Methods:LEAP‐mediated cell purification and optoinjection effects were assessed in model experiments using adherent and suspension cell types and cell mixtures plated and processed at different densities. Optoinjection effects were visualized by delivering fluorescent dextrans into cells. Results were analyzed using the LEAP instrument's own imaging system as well as by fluorescence and confocal microscopy.Results:Live cell samples (adherent and suspension) could be purified to 90–100% purity with 50–90% yield, causing minimal cell damage depending on the cell type and plating density. Nearly one hundred percent of the targeted cells of all cell types examined could be successfully optoinjected with dextrans of 3–70 kDa, causing no visual damage to the cells. Indirect optoinjection effects were observed on untargeted cells within 5–60 μm to targeted areas under conditions used here.Conclusions:LEAP provides solutions in cell purification and targeted macromolecule delivery for traditional and challenging applications where other methods fall short. © 2006 International Society for Analytical Cytology
Quality assessment of confocal microscopy slide‐based systems: Instability
Authors: Zucker Robert M. 
Published: June 2006

AbstractBackground:All slide‐based fluorescence cytometry detections systems basically include an excitation light source, intermediate optics, and a detection device (CCD or PMT). Occasionally, this equipment becomes unstable, generating unreliable and inferior data.Methods:A number of tests have been devised to evaluate equipment performance and instability. The following four instability tests are described: galvanometer scanning, stage drift, correct wavelength spectral detection, and long‐term laser power.Results:Quality assurance tests revealed that a confocal microscope can become unstable in the following parameters, yielding inaccurate data: laser power, PMTs functionality, spectrophotometer accuracy, galvanometer scanning and laser stability, and stage drift. Long‐term laser power stability has been observed to vary greatly.Conclusions:Confocal systems can become unstable in the following parameters: long‐term laser power, galvanometer scanning, spectrophotometer accuracy, and stage stability. Instability in any of these parameters will affect image quality. Laser power fluctuations result from either a defective Acousto‐optic tunable filter or improper heat dissipation. Spectrophotometer instability will generate unreliable spectra data, extra light reflections, and poor image quality. Galvanometer scanning instability yields poor image quality while microscope stage drift results in a sample going out of the plane of focus. With minor modifications, these tests may be applicable to other slide‐based systems. © 2006 International Society for Analytical Cytology
Quality assessment of confocal microscopy slide based systems: Performance
Authors: Zucker Robert M. 
Published: June 2006

AbstractBackground:All fluorescence slide‐based cytometry detections systems basically include the following components: (1) an excitation light source, (2) intermediate optics, and (3) a detection device consisting of a CCD camera or a PMT. The optical principles employed is slide‐based systems are similar to those of confocal microscopes (CLSM).Methods:The following tests evaluated confocal equipment performance: dichroic reflectivity, field illumination, lens performance, laser power output, spectral registration, axial resolution, PMT reliability, and system noise.Results:Quality assurance tests provide a basis to determine if the equipment is operating correctly. Laser power, PMTs function, dichroic reflection, spectral registration, axial registration, system noise and sensitivity, lens performance and laser stability were tested colocalization of UV and visible peaks of a bead should be less than 210 nm. Interference contrast optics decrease fluorescence resolution.Conclusions:QA tests that assess CLSM system performance are also applicable to other slide‐based systems. By utilization this type of testing approach, the subjective nature of assessing the CLSM may be eliminated. These tests serve as guidelines for other investigators to ensure that their machines are providing data that is accurate with the necessary resolution, sensitivity and precision. © 2006 International Society for Analytical Cytology
Slide‐based cytometry for cytomics—A minireview
Authors: Tárnok Attila 
Published: June 2006

AbstractIn the postgenomic era, to gain the most detailed quantitative data from biological specimens has become increasingly important in the emerging new fields of high‐content and high‐throughput single‐cell analysis for systems biology and cytomics. Areas of research and diagnosis with the demand to virtually measure “anything” in the cell include immunophenotyping, rare cell detection and characterization in the case of stem cells and residual tumor cells, tissue analysis, and drug discovery. Systemic analysis is also a prerequisite for predictive medicine by genomics, proteomics, and cytomics.This issue of Cytometry Part A is dedicated to innovative concepts of system wide single cells analysis and manipulation, new technologies, data analysis and display, and, finally, quality assessment. The manuscripts to these chapters are provided by cutting edge experts in the fields. This overview will briefly highlight the most important aspects of this continuously developing field. © 2006 International Society for Analytical Cytology
A Ruthenium(II) Complex with a Tripodal Ligand Containing Three Imidazole Groups
Authors: Yamaguchi Tomoka , Harada Kazutoshi , Sunatsuki Yukinari , Kojima Masaaki , Nakajima Kiyohiko , Matsumoto Naohide 
Published: June 2006

AbstractThe orange [RuII(H3L)](ClO4)2·3H2O (1) complex was synthesized from cis‐[RuCl2(dmso)4] and H3L, where H3L (tris{[2‐{(imidazol‐4‐yl)methylidene}amino]ethyl}amine) is a tripodal ligand obtained by condensation of tris(2‐aminoethyl)amine and 4‐formylimidazole in a 1:3 mol ratio. The X‐ray crystal structure analysis revealed that the complex has an octahedral structure coordinated by three imidazole nitrogen atoms and three Schiff base (imine) nitrogen atoms. The uncoordinated NH groups of the imidazole moieties of 1 are easily deprotonated by the action of a base. The effect of deprotonation on the RuIII/RuII redox potential was studied by cyclic voltammetry in methanol containing 0.1 M LiClO4. Complete deprotonation shifts the RuIII/RuII potential to a much more negative value from –0.18 to –0.72 V vs. Ag/Ag+, making it easier to oxidize. The bluish‐purple RuIII complex, [RuIII(H3L)]3+, was formed by controlled‐potential electrolysis of 1, and the electronic spectrum is reported. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)
Metal Atom Vibrational Amplitudes in Decamethylferrocene and Related Organometallics
Authors: Herber Rolfe H. , Nowik Israel , Kahlenberg Volker , Kopacka Holger , Schottenberger Herwig 
Published: June 2006

AbstractRecent experiments have shown that root‐mean‐square amplitudes‐of‐vibration (rmsav) for iron atoms in organometallic complexes at various temperatures can be extracted from temperature‐dependent 57Fe Mössbauer spectroscopic data related to the recoil‐free fraction of the metal atom. The applicability of this procedure has been validated by comparison with rmsav data extracted from Uij values obtained from single‐crystal X‐ray diffraction parameters at various temperatures. This analysis has been applied in detail to a study of decamethylferrocene (DMFc), for which the agreement between the rmsav values derived from the two experimental procedures is compared in the present study. A comparison of these rmsav values for a number of related structures is also reported over a similar temperature range. An improved method for the quantitative preparation of isotopically enriched DMFc is described. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)
Simultaneous Regio‐ and Enantiodifferentiation in Carbohydrate Coupling
Authors: Cid M. Belén , Alonso Inés , Alfonso Francisco , Bonilla Julia B. , López‐Prados Javier , Martín‐Lomas Manuel 
Published: June 2006

AbstractThe glycosylation of 1,2‐trans‐diequatorial diols derived from tetrabenzoylated and tetrabenzylated D‐ and L‐chiro‐inositol with several glycosyl donors has been investigated. An unprecedented dependence of the regioselectivity on the absolute configuration of the acceptor has been found. However this trend is also modulated by the nature of the protecting groups on both the donor and acceptor, with benzoylated acceptors affording higher levels of regioselectivity. Most of the results have been rationalized by DFT calculations which indicate that stereoelectronic factors and hydrogen bonding between the donor and acceptor govern their relative orientation and determine the regiochemical outcome of the process. These studies also highlight the role of the acyl group adjacent to the OH to be glycosylated in facilitating the glycosylation reaction. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)
The Synthesis of cis‐ and trans‐Fused Bicyclic Sugar Amino Acids
Authors: Risseeuw Martijn D. P. , Grotenbreg Gijsbert M. , Witte Martin D. , Tuin Adriaan W. , Leeuwenburgh Michiel A. , Van der Marel Gijsbert A. , Overkleeft Herman S. , Overhand Mark 
Published: June 2006

AbstractFour isomeric bicyclic sugar amino acids (SAAs) were prepared from α‐acetylenic‐C‐glucoside 6 by employing a Petasis olefination and a ring‐closing metathesis (RCM) as key steps. The applicability of the resulting SAAs in solid‐phase peptide synthesis was demonstrated by the synthesis of tetrapeptide 36. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)