Optimizing the electrical communication between enzymes and electrodes is critical in the development of biosensors, enzymatic biofuel cells, and other bioelectrocatalytic applications. One approach to address this limitation is the attachment of redox mediators or relays to the enzymes. Here we report a simple genetic modification of a glucose oxidase enzyme to display a free thiol group near its active site. This facilitates the site-specific attachment of a maleimide-modified gold nanoparticle to the enzyme, which enables direct electrical communication between the conjugated enzyme and an electrode. Glucose oxidase is of particular interest in biofuel cell and biosensor applications, and the approach of "prewiring" enzyme conjugates in a site-specific manner will be valuable in the continued development of these systems.

This study demonstrates functional independence in the acquisition of mands and tacts. Some subjects first learned to mand the experimenter's placement of objects with the prepositional phrases "On the left" and "On the right." They were regularly tested for collateral appearance of tacts with these same phrases. Other subjects learned to tact the location of objects with these prepositional phrases and were regularly tested for collateral appearance of mands. All subjects were next trained in the repertoire that had not been trained in the first condition (either tact or mand). After all subjects had learned both to mand and to tact correctly, another assessment of mand-tact independence was undertaken. Mands (tacts) were reversed and testing assessed collateral reversal of tacts (mands). The results demonstrated that tacts and mands, even when incorporating identical response forms, were functionally independent during acquisition. Subsequent modification of one repertoire (by reversal training) produced collateral reversal in three of nine subjects.

Recent progress in understanding the Q-cycle mechanism of the bc(1) complex is reviewed. The data strongly support a mechanism in which the Q(o)-site operates through a reaction in which the first electron transfer from ubiquinol to the oxidized iron-sulfur protein is the rate-determining step for the overall process. The reaction involves a proton-coupled electron transfer down a hydrogen bond between the ubiquinol and a histidine ligand of the [2Fe-2S] cluster, in which the unfavorable protonic configuration contributes a substantial part of the activation barrier. The reaction is endergonic, and the products are an unstable ubisemiquinone at the Q(o)-site, and the reduced iron-sulfur protein, the extrinsic mobile domain of which is now free to dissociate and move away from the site to deliver an electron to cyt c(1) and liberate the H(+). When oxidation of the semiquinone is prevented, it participates in bypass reactions, including superoxide generation if O(2) is available. When the b-heme chain is available as an acceptor, the semiquinone is oxidized in a process in which the proton is passed to the glutamate of the conserved -PEWY- sequence, and the semiquinone anion passes its electron to heme b(L) to form the product ubiquinone. The rate is rapid compared to the limiting reaction, and would require movement of the semiquinone closer to heme b(L) to enhance the rate constant. The acceptor reactions at the Q(i)-site are still controversial, but likely involve a "two-electron gate" in which a stable semiquinone stores an electron. Possible mechanisms to explain the cyt b(150) phenomenon are discussed, and the information from pulsed-EPR studies about the structure of the intermediate state is reviewed. The mechanism discussed is applicable to a monomeric bc(1) complex. We discuss evidence in the literature that has been interpreted as shown that the dimeric structure participates in a more complicated mechanism involving electron transfer across the dimer interface. We show from myxothiazol titrations and mutational analysis of Tyr-199, which is at the interface between monomers, that no such inter-monomer electron transfer is detected at the level of the b(L) hemes. We show from analysis of strains with mutations at Asn-221 that there are coulombic interactions between the b-hemes in a monomer. The data can also be interpreted as showing similar coulombic interaction across the dimer interface, and we discuss mechanistic implications.

The form frequently taken by behavior-modification programs is analyzed in terms of the parent science, Behaviorism. Whereas Behaviorism assumes that behavior is the result of contingencies, and that lasting behavior change involves changing the contingencies that give rise to and support the behavior, most behavior-modification programs merely arrange special contingencies in a special environment to eliminate the "problem" behavior. Even when the problem behavior is as widespread as alcoholism and crime, behavior modifiers focus on "fixing" the alcoholic and the criminal, not on changing the societal contingencies that prevail outside the therapeutic environment and continue to produce alcoholics and criminals. The contingencies that shape this method of dealing with behavioral problems are also analyzed, and this analysis leads to a criticism of the current social structure as a behavior control system. Although applied behaviorists have frequently focused on fixing individuals, the science of Behaviorism provides the means to analyze the structures, the system, and the forms of societal control that produce the "problems".

Pulsed EPR spectroscopy was used to explore the structural neighborhood of the semiquinone (SQ) stabilized at the Q i site of the bc 1 complex of Rhodobacter sphaeroides (EC 1.10.2.2) and to demonstrate that the nitrogen atom of a histidine imidazole group donates an H-bond to the SQ. Crystallographic structures show two different configurations for the binding of ubiquinone at the Q i site of mitochondrial bc 1 complexes in which histidine (His-201 in bovine sequence) is either a direct H-bond donor or separated by a bridging water. The paramagnetic properties of the SQ formed at the site provide an independent method for studying the liganding of this intermediate species. The antimycin-sensitive SQ formed at the Q i site by either equilibrium redox titration, reduction of the oxidized complex by ascorbate, or addition of decylubihydroquinone to the oxidized complex in the presence of myxothiazol all showed similar properties. The electron spin echo envelope modulation spectra in the 14 N region were dominated by lines with frequencies at 1.7 and 3.1 MHz. Hyperfine sublevel correlation spectroscopy spectra showed that these were contributed by a single nitrogen. Further analysis showed that the 14 N nucleus was characterized by an isotropic hyperfine coupling of ϳ0.8 MHz and a quadrupole coupling constant of ϳ0.35 MHz. The nitrogen was identified as the N-⑀ or N-␦ imidazole nitrogen of a histidine (it is likely to be His-217, or His-201 in bovine sequence). A distance of 2.5-3.1 Å for the O-N distance between the carbonyl of SQ and the nitrogen was estimated. The mechanistic significance is discussed in the context of a dynamic role for the movement of His-217 in proton transfer to the site.The bc 1 complex family of enzymes plays a central role in all the main pathways of energy conversion, being directly responsible for ϳ30% of all the energy transduction of the biosphere (1-3). The complexes in Rhodobacter exemplify the simplest of these enzymes, with only three or four subunits, including the highly conserved catalytic core common to bacterial and mitochondrial complexes. It is generally accepted that the complex operates through a protonmotive Q cycle (2, 4, 5). Three catalytic subunits, cytochrome b, cytochrome c 1 , and the Rieske iron-sulfur protein, house the mechanism. Two separate internal electron transfer chains connect three catalytic sites for external substrates. At one site, cytochrome c 1 is oxidized by cytochrome c 2 . Two catalytic sites in cytochrome b are involved in the oxidation or reduction of ubiquinone. At the quinol oxidizing site, one electron from quinol is passed to the ironsulfur protein, which transfers it to cytochrome c 1 , whereas the semiquinone (SQ) 1 produced is oxidized by another chain consisting of the two b-hemes of cytochrome b in the bifurcated reaction. At the quinone-reducing site (Q i site), electrons from the b-heme chain are used to generate quinol. The integration of the oxidation and reduction reactions with the release or uptake of protons in the aqueous pha...