Specific and reversible immobilization of proteins tagged to the affinity polypeptide C-LytA on functionalized graphite electrodes

We have developed a general method for the specific and reversible immobilization of proteins fused to the choline-binding module C-LytA on functionalized graphite electrodes. Graphite electrode surfaces were modified by diazonium chemistry to introduce carboxylic groups that were subsequently used to anchor mixed self-assembled monolayers consisting of N,N-diethylethylenediamine groups, acting as choline analogs, and ethanolamine groups as spacers. The ability of the prepared electrodes to specifically bind C-LytA-tagged recombinant proteins was tested with a C-LytA-β-galactosidase fusion protein. The binding, activity and stability of the immobilized protein was evaluated by electrochemically monitoring the formation of an electroactive product in the enzymatic hydrolysis of the synthetic substrate 4-aminophenyl β-D-galactopyranoside. The hybrid protein was immobilized in an specific and reversible way, while retaining the catalytic activity. Moreover, these functionalized electrodes were shown to be highly stable and reusable. The method developed here can be envisaged as a general, immobilization procedure on the protein biosensor field.

Analysis of the Brominated Dioxin and Furan Emission Congener Pattern from Different Sources

Polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/F) have been studied for several decades and are well-known as unintentionally generated persistent organic pollutants (POPs), which pose serious health and environmental risks on a global scale1. Polybrominated dibenzo-p-dioxins and dibenzofurans (PBDD/F) have similar properties and effects to PCDD/F, as they are structural analogs with all the chlorine atoms substituted by bromine atoms. PBDD/F have been found in various matrices such as air, sediments, marine products, and human adipose samples.

A Comprehensive Discussion of HMBC Pulse Sequences: 4. Establishing Two-Bond Correlations from HMBC and Related Experiments

The utility of the HMBC experiment for structure elucidation is unquestionable, but the nature of the coupling pathways leading to correlations in an HMBC experiment creates the potential for misinterpretation. This misinterpretation potential is intimately linked to the size of the long-range heteronuclear couplings involved, and may become troublesome in those cases of a particularly strong 2JCH correlation that might be mistaken for a 3JCH correlation or a 4JCH correlation of appreciable strength that could be mistaken for a weaker 3JCH correlation. To address these potential avenues of confusion, work from several laboratories has been focused on the development of what might be considered “coupling pathway edited” long-range heteronuclear correlation experiments that are derived from or related to the HMBC experiment. The first example of an effort to address the problems associated with correlation path length was seen in the heteronucleus-detected XCORFE experiment described by Reynolds and co-workers that predated the development of the HMBC experiment. Proton-detected analogs of the HMBC experiment intended to differentiate 2JCH correlations from nJCH correlations where n = 3, 4, include the 2J,3J-HMBC, HMBC-RELAY, H2BC, edited-HMBC, and HAT H2BC experiments. The principles underlying the critical components of each of these experiments are discussed and experimental verification of the results that can be obtained using model compounds are shown. This contribution concludes with a brief discussion of the 1,1-ADEQUATE experiments that provide an alternative means of identifying adjacent protonated and non-protonated carbon correlations by exploiting 1JCC correlations at natural abundance.

(Table 1) Ooze/chalk cycles: frequency of occurrence at DSDP Leg 74 Sites

Shallow- to deep-water environments are represented by the sediments and rocks recovered from the Walvis Ridge- Angola Basin transect. These calcareous oozes, chalks, limestones, and volcaniclastic sedimentary rocks are used to define and correlate four lithostratigraphic units. The sediments were deposited in cycles which represent recurring tectonic or Oceanographic events and may be related to climatic fluctuations and orbital perturbations. Turbidites are the most common and easily identified sedimentary cycle. They are Late Cretaceous to Paleocene in age and are repeated in intervals ranging from thousands to tens of thousands of years. They are also found interbedded between basalt layers. Turbidites are easily distinguished from the other cycles present by their sedimentary structures, mineral composition, alteration products, and physical properties (GRAPE) data. Large-scale turbidites, debris, or slump breccias are found at or just above the Cretaceous/Tertiary boundary and indicate an event of considerable energy possibly related to intense tectonic activity. Diagenetic cycles, interpreted as small-scale dissolution cycles or sequences produced by biogenic activity, occur in early Paleocene chalks. The recurrence intervals average -20,000 y. but have a wide range of values. Variations in CaCO3 content, color, gradational boundaries, and trace fossil content characterize these sediments. These cycles reflect bottom-water conditions. Ooze-chalk cycles occur in upper Oligocene to upper Paleocene sediments and represent conditions that once existed at the sediment/water interface where they obtained their diagenetic potential. These oscillations are repeated over tens of thousands of years and may have no modern analogs. Color variations in sediments at the Cretaceous/Tertiary boundary indicate local fluctuations in oxygen content within the sediments or the water column. This situation lasted for several hundred thousand years and is not repeated elsewhere in the sequence. Large dissolution cycles are recorded in the sediments at Site 527 that are of middle Miocene and early Oligocene to middle Eocene age. During this time the seafloor at this site appears to have been located at or subsided to a depth occupied by a fluctuating CCD and lysocline.