Glutarate and N-acetyl-L-glutamate buffers for cell-free synthesis of selectively 15N-labelled proteins

Cell-free protein synthesis provides rapid and economical access to selectively 15N-labelled proteins, greatly facilitating the assignment of 15N-HSQC spectra. While the best yields are usually obtained with buffers containing high concentrations of potassium L-glutamate, preparation of selectively 15N-Glu labelled samples requires non-standard conditions. Among many compounds tested to replace the L-Glu buffer, potassium N-acetyl-L-glutamate and potassium glutarate were found to perform best, delivering high yields for all proteins tested, with preserved selectivity of 15N-Glu labelling. Assessment of amino-transferase activity by combinatorial 15N-labelling revealed that glutarate and N-acetyl-L-glutamate suppress the transfer of the 15N-alpha-amino groups between amino acids less well than the conventional L-Glu buffer. On balance, the glutarate buffer appears most suitable for the preparation of samples containing 15N-L-Glu while the conventional L-Glu buffer is advantageous for all other samples

Introducing energetic and capacitive energy efficiency measures in the sector of chilled and frozen food storage via an electronic platform

The freshness of agricultural products is kept well when chilled or frozen. Additionally, chilled and frozen food is en vogue due to convenience reasons. The growing demand for chilled and frozen food on a global scale has some negative ecological side effects, though. The GEMIT Institute for Business process Management and IT developed the idea to combine an electronic market place for idle cold storage capacities with a tool that helps coldstores increase their energy efficiency. It is ecologically worthwhile in a branch characterized by high CO2-emissions and energy consumption. A future marketplace does not only serve as a spot market for short term demand in storage, where it guarantees cheaper storage space in proximity and higher independency from common cold storage services. Additionally, the supplier of empty cold storage space generates new revenues and at the same time spends the maintenance energy more efficiently. Finally, the participating suppliers can benefit from a benchmarking tool and information about their own energy savings potentials. The current market situation is especially advantageous for an introduction of ECO-Store, with growing cold storage spaces in Germany and an increasing consumption of chilled food worldwide. Aspects that can impose a barrier to the project are considered as well. It was found out that companies might not want to disclose data or cooperate with possible competitors and operating providers of cold storage space fear a decline in prices. Reaching a critical mass of participants in order to assure the cost effectiveness of the investment and to increase the attractiveness itself is a crucial issue. Finally, resembling concepts already exist on the market. Yet, what it makes unique is the combination of the electronic market place and a tool aiming at reducing individual energy consumption

Introducing energetic and capacitive energy efficiency measures in the sector of chilled and frozen food storage via an electronic platform

The freshness of agricultural products is kept well when chilled or frozen. Additionally, chilled and frozen food is en vogue due to convenience reasons. The growing demand for chilled and frozen food on a global scale has some negative ecological side effects, though. The GEMIT Institute for Business process Management and IT developed the idea to combine an electronic market place for idle cold storage capacities with a tool that helps coldstores increase their energy efficiency. It is ecologically worthwhile in a branch characterized by high CO2-emissions and energy consumption. A future marketplace does not only serve as a spot market for short term demand in storage, where it guarantees cheaper storage space in proximity and higher independency from common cold storage services. Additionally, the supplier of empty cold storage space generates new revenues and at the same time spends the maintenance energy more efficiently. Finally, the participating suppliers can benefit from a benchmarking tool and information about their own energy savings potentials. The current market situation is especially advantageous for an introduction of ECO-Store, with growing cold storage spaces in Germany and an increasing consumption of chilled food worldwide. Aspects that can impose a barrier to the project are considered as well. It was found out that companies might not want to disclose data or cooperate with possible competitors and operating providers of cold storage space fear a decline in prices. Reaching a critical mass of participants in order to assure the cost effectiveness of the investment and to increase the attractiveness itself is a crucial issue. Finally, resembling concepts already exist on the market. Yet, what it makes unique is the combination of the electronic market place and a tool aiming at reducing individual energy consumption

OR3 operator of bacteriophage lambda in a 23 base-pair DNA fragment: sequence-specific 1H NMR assignments for the non-labile protons and comparison with the isolated 17 base-pair operator

Sequence-specific 1H NMR assignments are presented for a non-selfcomplementary 23-base-pair DNA duplex of molecular weight 15,000 daltons, containing the OR3 repressor binding site of bacteriophage lambda as the central core. The NMR techniques used were mainly phase-sensitive two-dimensional NOE and 2Q spectroscopy, the latter to overcome overlap problems within the spectral region of the deoxyribose spin-systems. Direct sequential NOE connectivities are observed between adenine 2 H and deoxyribose 1' protons. We propose the use of these connectivities as a check of the assignments of C1' and A2 protons, which have independently been derived via other assignment pathways

Application of 13C(ω1)-half-filtered [1H,1H]-NOESY for studies of a complex formed between DNA and a 13C-labeled minor-groove-binding drug

AbstractThe complex formed between the anticancer drug 4-[p-[p-(4-quinolylamino)benzamido]anilino]pyridine (SN 6999) and the decadeoxyribonucleoside nonaphosphate d-(GCATTAATGC)2 was investigated using two-dimensional nuclear Overhauser enhancement spectroscopy (NOESY) with a 13C(ω1)-half-filter. The two quaternary methyl groups in SN 6999 had been labeled with 13C for these experiments. The simplified subspectra of [1H,1H]-NOESY obtained with this procedure greatly facilitate the identification and assignment of intermolecular NOEs. Quite generally, the combined use of isotope labeling and heteronuclear filters in [1H,1H]-NOESY provides an improved experimental basis for structural studies of drug/ DNA complexes

Sequential NMR assignments of labile protons in DNA using two-dimensional nuclear-overhauser-enhancemnt spectroscopy with three jump-and-return pulse sequences

Two-dimensional nuclear Overhauser enhancement (NOESY) spectra of labile protons were recorded in H2O solutions of a protein and of a DNA duplex, using a modification of the standard NOESY experiment with all three 90° pulses replaced by jump-and-return sequences. For the protein as well as the DNA fragment the strategically important spectral regions could be recorded with good sensitivity and free of artifacts. Using this procedure, sequence-specific assignments were obtained for the imino protons, C2H of adenine, and C4NH2 of cytosine in a 23-base-pair DNA duplex which includes the 17-base-pair OR3 repressor binding site of bacteriophage λ. Based on comparison with previously published results on the isolated OR3 binding site, these data were used for a study of chain termination effects on the chemical shifts of imino proton resonances of DNA duplexes

Hydrogen Bond Dynamics Near A Micellar Surface: Origin of the Universal Slow Relaxation at Complex Aqueous Interfaces

The dynamics of hydrogen bonds among water molecules themselves and with the polar head groups (PHG) at a micellar surface have been investigated by long molecular dynamics simulations. The lifetime of the hydrogen bond between a PHG and a water molecule is found to be much longer than that between any two water molecules, and is likely to be a general feature of hydrophilic surfaces of organized assemblies. Analyses of individual water trajectories suggest that water molecules can remain bound to the micellar surface for more than a hundred picosecond. The activation energy for such a transition from the bound to a free state for the water molecules is estimated to be about 3.5kcal/mole.Comment: 12 pages. Phys. Rev. Lett. (Accepted) (2002

The mosaic of KIR haplotypes in rhesus macaques

To further refine and improve biomedical research in rhesus macaques, it is necessary to increase our knowledge concerning both the degree of allelic variation (polymorphism) and diversity (gene copy number variation) in the killer cell immunoglobulin-like receptor (KIR) gene cluster. Pedigreed animals in particular should be studied, as segregation data will provide clues to the linkage of particular KIR genes/alleles segregating on a haplotype and to its gene content as well. A dual strategy allowed us to screen the presence and absence of genes and the corresponding transcripts, as well as to track differences in transcription levels. On the basis of this approach, 14 diverse KIR haplotypes have been described. These haplotypes consist of multiple inhibitory and activating Mamu-KIR genes, and any gene present on one haplotype may be absent on another. This suggests that the cost of accelerated evolution by recombination may be the loss of certain framework genes on a haplotype

Sequential NMR assignments of labile protons in DNA using two-dimensional nuclear-Overhauser-enhancement spectroscopy with three jump-and-return pulse sequences

Two-dimensional nuclear Overhauser enhancement (NOESY) spectra of labile protons were recorded in H2O solutions of a protein and of a DNA duplex, using a modification of the standard NOESY experiment with all three 90 degree pulses replaced by jump-and-return sequences. For the protein as well as the DNA fragment the strategically important spectral regions could be recorded with good sensitivity and free of artifacts. Using this procedure, sequence-specific assignments were obtained for the imino protons, C2H of adenine, and C4NH2 of cytosine in a 23-base-pair DNA duplex which includes the 17-base-pair OR3 repressor binding site of bacteriophage lambda. Based on comparison with previously published results on the isolated OR3 binding site, these data were used for a study of chain termination effects on the chemical shifts of imino proton resonances of DNA duplexes