Powering future generations of implanted medical devices will require cumbersome transcutaneous energy transfer or harvesting energy from the human body. No functional solution that harvests power from the body is currently available, despite attempts to use the Seebeck thermoelectric effect, vibrations or body movements. Glucose fuel cells appear more promising, since they produce electrical energy from glucose and dioxygen, two substrates present in physiological fluids. The most powerful ones, Glucose BioFuel Cells (GBFCs), are based on enzymes electrically wired by redox mediators. However, GBFCs cannot be implanted in animals, mainly because the enzymes they rely on either require low pH or are inhibited by chloride or urate anions, present in the Extra Cellular Fluid (ECF). Here we present the first functional implantable GBFC, working in the retroperitoneal space of freely moving rats. The breakthrough relies on the design of a new family of GBFCs, characterized by an innovative and simple mechanical confinement of various enzymes and redox mediators: enzymes are no longer covalently bound to the surface of the electron collectors, which enables use of a wide variety of enzymes and redox mediators, augments the quantity of active enzymes, and simplifies GBFC construction. Our most efficient GBFC was based on composite graphite discs containing glucose oxidase and ubiquinone at the anode, polyphenol oxidase (PPO) and quinone at the cathode. PPO reduces dioxygen into water, at pH 7 and in the presence of chloride ions and urates at physiological concentrations. This GBFC, with electrodes of 0.133 mL, produced a peak specific power of 24.4 µW mL−1, which is better than pacemakers' requirements and paves the way for the development of a new generation of implantable artificial organs, covering a wide range of medical applications.

The ALCR protein is the transcriptional activator of the ethanol utilization pathway in the filamentous fungus AspergiUlus nidulans. This activator belongs to a family of fungal proteins having a conserved DNA-binding domain containing six cysteines (C6 class) with some striking features. At variance with other motifs of this class, the binding domain of ALCR is strongly asymmetrical in relation to the central cysteines and moreover was predicted to adopt a helix-turn-helix structure. This domain of ALCR was synthesized in Escherichia coli and purified as a glutathione-S-transferase fusion protein. Our results show that the transcriptional activator ALCR is a DNA-binding protein. The DNA-binding motif contains zinc that is necessary for the specific DNA binding. The ALCR peptide binds upstream of the coding region of alcR to two specific targets with different affinities that are characterized by a conserved 5-nucleotide core, 5'-CCGCA-3' (or its reverse). One site, the lower-affinity binding site, is a direct repeat, and the other, the higher-affinity binding site, is a palindromic sequence with dyad symmetry. Therefore, the ALCR binding protein is able to recognize one DNA sequence in two different configurations. An alcR mutant obtained by deletion of the two specific targets in the cis-acting region of the alcR gene is unable to grow on ethanol and does not express any alcohol dehydrogenase activity. These results demonstrate that the binding sites are in vivo functional targets (UASaic) for the ALCR protein in A. nidulans. They corroborate prior evidence that alcR is autoregulated.Positive control mechanisms in eukaryotes have been extensively characterized. They are mediated by transcription factors that bind to specific DNA targets. The expression of genes encoding the ethanol utilization enzymes in the ascomycete Aspergillus nidulans is regulated by the pathway-specific transactivator ALCR. In conditions of induction (by ethanol or gratuitous inducers like ethylmethylketone), the ALCR protein is necessary for the expression of the two structural genes alcA, encoding alcohol dehydrogenase I, and aldA, encoding aldehyde dehydrogenase (30, 36). Transcription of these two genes can be strongly induced, and this property was widely used for the expression of heterologous proteins (for a review, see reference 11). The expression of the alcR gene is inducible, positively autoregulated, and subjected to carbon catabolite repression under the control of the negatively acting gene creA (11,25,30). The three genes of the ethanol regulon were cloned and sequenced (12,16,26,32), and the creA gene identified by Bailey and Arst (2) was also cloned (8) and sequenced (9). The transcription factor ALCR is 821 amino acids long (12) and contains a sequence of six Cys residues, Cys-X2-Cys-X6-Cys-X16-Cys-X2-Cys-X6-Cys within its N-terminal part. It is related to the highly conserved DNA-binding domain of the transcription factors of the C6 class of the ascomycetes (23). At variance with other motifs of this class, the putativ...

AlcR is the transactivator mediating transcriptional induction of the alc gene cluster in Aspergillus nidulans. The AlcR DNA-binding domain consists of a zinc binuclear cluster different from the other members of the Zn 2 Cys 6 family by several features. In particular, it is able to bind to symmetric and asymmetric sites with the same affinity, with both sites being functional in A. nidulans. Here, we show that unlike the other proteins of the Zn 2 Cys 6 binuclear cluster family, AlcR binds most probably as a monomer to its cognate targets. Two molecules of the AlcR protein can simultaneously bind in a noncooperative manner to inverted repeats. The consensus core has been determined precisely (5 -CCGCN-3 ), and the AlcR-binding site in the aldA promoter has been localized. The sequence downstream of the zinc cluster is necessary for high affinity binding. Furthermore, our data show that the use of the carrier protein glutathione S-transferase in AlcR binding experiments introduces an important bias in the recognition of DNA sites due to its tertiary dimeric structure.