Biomedical subjects

Yann Astier

Publications and source records attributed to Yann Astier.

4 recordsLinked to original sources

PubMed2006

Toward single molecule DNA sequencing: direct identification of ribonucleoside and deoxyribonucleoside 5'-monophosphates by using an engineered protein nanopore equipped with a molecular adapter.

Individual nucleic acid molecules might be sequenced by the identification of nucleoside 5'-monophosphates as they are released by processive exonucleases. Here, we show that single molecule detection with a modified protein nanopore can be used to identify ribonucleoside and 2'-deoxyribonucleoside 5'-monophosphates, thereby taking a step along this path. Distinct levels of current block are observed for each of the four members of a set of nucleoside 5'-monophosphates when the molecules bind within a mutant alpha-hemolysin pore, (M113R)(7), equipped with the molecular adapter heptakis-(6-deoxy-6-amino)-beta-cyclodextrin. While our results compare favorably with alternative approaches, further work will be required to improve the accuracy of identification of the nucleic acid bases, to feed each released nucleotide into the pore, and to ensure that every nucleotide is captured by the adapter.

DNA↗

PubMed2005

Protein components for nanodevices.

A long-term goal of nanobiotechnology is to build tiny devices that respond to the environment, perform computations and carry out tasks. Considerable progress has been made in building protein components for such devices, and here we describe examples, including self-assembling protein arrays, pores with triggers and switches, and motor proteins harnessed for specific tasks. A major issue that has been successfully addressed in this recent work is the interface between the proteins and other components of the system, such as a metal surface. While further progress is expected in the coming years, the assembly of devices from the components has seen more limited accomplishments. For example, although a wide variety of sensors based on nanobiotechnology has been developed, unresolved problems still confront the construction of complex nanobioelectronic circuits, and the development of nanorobotics with biological components remains a distant dream.

Microchip Analytical Procedures↗

PubMed2005

Sensing nitrite through a pseudoazurin-nitrite reductase electron transfer relay.

Nitrite is converted to nitric oxide by haem or copper-containing enzymes in denitrifying bacteria during the process of denitrification. In designing an efficient biosensor, this enzymic turnover must be quantitatively assessed. The enzyme nitrite reductase from Alcaligenes faecalis contains a redox-active blue copper centre and a nonblue enzyme-active copper centre. It can be covalently tethered to modified gold-electrode surfaces in configurations in which direct electron transfer is possible. A surface cysteine mutant of the enzyme can be similarly immobilised on bare electroactive gold substrates. Under such circumstances, however, electron transfer cannot be effectively coupled with substrate catalytic turnover. In using either the natural redox partner, pseudoazurin, or ruthenium hexammine as an "electron-shuttle" or "conduit" between enzyme and a peptide-modified electrode surface, the coupling of electron transfer to catalysis can be utilised in the development of an amperometric nitrite sensor.

Azurin↗

PubMed2003

Cofactor-independent oxygenation reactions catalyzed by soluble methane monooxygenase at the surface of a modified gold electrode.

Soluble methane monooxygenase (sMMO) is a three-component enzyme that catalyses dioxygen- and NAD(P)H-dependent oxygenation of methane and numerous other substrates. Oxygenation occurs at the binuclear iron active centre in the hydroxylase component (MMOH), to which electrons are passed from NAD(P)H via the reductase component (MMOR), along a pathway that is facilitated and controlled by the third component, protein B (MMOB). We previously demonstrated that electrons could be passed to MMOH from a hexapeptide-modified gold electrode and thus cyclic voltammetry could be used to measure the redox potentials of the MMOH active site. Here we have shown that the reduction current is enhanced by the presence of catalase or if the reaction is performed in a flow-cell, probably because oxygen is reduced to hydrogen peroxide, by MMOH at the electrode surface and the hydrogen peroxide then inactivates the enzyme unless removed by catalase or a continuous flow of solution. Hydrogen peroxide production appears to be inhibited by MMOB, suggesting that MMOB is controlling the flow of electrons to MMOH as it does in the presence of MMOR and NAD(P)H. Most importantly, in the presence of MMOB and catalase, the electrode-associated MMOH oxygenates acetonitrile to cyanoaldehyde and methane to methanol. Thus the electochemically driven sMMO showed the same catalytic activity and regulation by MMOB as the natural NAD(P)H-driven reaction and may have the potential for development into an economic, NAD(P)H-independent oxygenation catalyst. The significance of the production of hydrogen peroxide, which is not usually observed with the NAD(P)H-driven system, is also discussed.

Acetonitriles↗