Electron transfer in acetohydroxy acid synthase as a side reaction of catalysis. implications for the reactivity and partitioning of the carbanion/enamine form of (alpha-hydroxyethyl)thiamin diphosphate in a "nonredox" flavoenzyme

Acetohydroxy acid synthases (AHAS) are thiamin diphosphate- (ThDP-) and FAD-dependent enzymes that catalyze the first common step of branched-chain amino acid biosynthesis in plants, bacteria, and fungi. Although the flavin cofactor is not chemically involved in the physiological reaction of AHAS, it has been shown to be essential for the structural integrity and activity of the enzyme. Here, we report that the enzyme-bound FAD in AHAS is reduced in the course of catalysis in a side reaction. The reduction of the enzyme-bound flavin during turnover of different substrates under aerobic and anaerobic conditions was characterized by stopped-flow kinetics using the intrinsic FAD absorbance. Reduction of enzyme-bound FAD proceeds with a net rate constant of k' = 0.2 s(-1) in the presence of oxygen and approximately 1 s(-1) under anaerobic conditions. No transient flavin radicals are detectable during the reduction process while time-resolved absorbance spectra are recorded. Reconstitution of the binary enzyme-FAD complex with the chemically synthesized intermediate 2-(hydroxyethyl)-ThDP also results in a reduction of the flavin. These data provide evidence for the first time that the key catalytic intermediate 2-(hydroxyethyl)ThDP in the carbanionic/enamine form is not only subject to covalent addition of 2-keto acids and an oxygenase side reaction but also transfers electrons to the adjacent FAD in an intramolecular redox reaction yielding 2-acetyl-ThDP and reduced FAD. The detection of the electron transfer supports the idea of a common ancestor of acetohydroxy acid synthase and pyruvate oxidase, a homologous ThDP- and FAD-dependent enzyme that, in contrast to AHASs, catalyzes a reaction that relies on intercofactor electron transfer.

Coherent charge and spin transfer in a quantum dot coupled to a mesoscopic ring

We study the effect of coherent charge and spin fluctuations in a mesoscopic device composed of a quantum dot and an Aharonov-Bohm ring. We show that, while the charge fluctuations suppress the persistent current algebraically as a function of the level spacing of the ring, the spin fluctuations give rise to a completely different behavior. We discuss the origin of this difference in relation to the peculiar nature of the ground state in the Kondo limit. (C) 2003 Elsevier B.V. All rights reserved.

Molecular basis of intramolecular electron transfer in sulfite-oxidizing enzymes is revealed by high resolution structure of a heterodimeric complex of the catalytic molybdopterin subunit and a c-type cytochrome subunit

Sulfite-oxidizing molybdoenzymes convert the highly reactive and therefore toxic sulfite to sulfate and have been identified in insects, animals, plants, and bacteria. Although the well studied enzymes from higher animals serve to detoxify sulfite that arises from the catabolism of sulfur-containing amino acids, the bacterial enzymes have a central role in converting sulfite formed during dissimilatory oxidation of reduced sulfur compounds. Here we describe the structure of the Starkeya novella sulfite dehydrogenase, a heterodimeric complex of the catalytic molybdopterin subunit and a c-type cytochrome subunit, that reveals the molecular mechanism of intramolecular electron transfer in sulfite-oxidizing enzymes. The close approach of the two redox centers in the protein complex (Mo-Fe distance 16.6 angstrom) allows for rapid electron transfer via tunnelling or aided by the protein environment. The high resolution structure of the complex has allowed the identification of potential through-bond pathways for electron transfer including a direct link via Arg-55A and/or an aromatic-mediated pathway. A potential site of electron transfer to an external acceptor cytochrome c was also identified on the SorB subunit on the opposite side to the interaction with the catalytic SorA subunit.

Pressure and temperature effects on metal-to-metal charge transfer in cyano-bridged Co-III-Fe-II complexes

The effects of pressure and temperature on the energy (E-op) of the metal-to-metal charge transfer (MMCT, Fe-II --> Co-III) transition of the cyano-bridged complexes trans - [(LCoNCFe)-Co-14(CN)(5)](-) and cis-[(LCoNCFe)-Co-14(CN)(5)](-) (where L-14 = 6-methyl-1,4,8,11-tetraazacyclotetradecan-6-amine) were examined. The changes in the redox potentials of the cobalt and iron metal centres with pressure and temperature were also examined and the results interpreted with Marcus Hush theory. The observed redox reaction volumes can mainly be accounted for in terms of localised electrostriction effects. The shifts in E-op due to both pressure and temperature were found to be less than the shifts in the energy difference (E degrees) between the Co-III-Fe-II and Co-II-Fe-III redox isomers. The pressure and temperature dependence of the reorganisational energy, as well as contributions arising from the different spin states of Co-II, are discussed in order to account for this trend. To study the effect of pressure on Co-III electronic absorption bands, a new cyano-bridged complex, trans - [(LCoNCCo)-Co-14(CN)(5)], was prepared and characterised spectroscopically and structurally. X-Ray crystallography revealed this complex to be isostructural with trans -[(LCoNCFe)-Co-14(CN)(5)] center dot 5H(2)O.

Experimental and analytical study of the effect of contact angle on liquid convective heat transfer in microchannels

In this paper we examine the effect of contact angle (or surface wettability) on the convective heat transfer coefficient in microchannels. Slip flow, where the fluid velocity at the wall is non-zero, is most likely to occur in microchannels due to its dependence on shear rate or wall shear stress. We show analytically that for a constant pressure drop, the presence of slip increases the Nusselt number. In a microchannel heat exchanger we modified the surface wettability from a contact angle of 20 degrees-120 degrees using thin film coating technology. Apparent slip flow is implied from pressure and flow rate measurements with a departure from classical laminar friction coefficients above a critical shear rate of approximately 10,000 s(-1). The magnitude of this departure is dependant on the contact angle with higher contact angles surfaces exhibiting larger pressure drop decreases. Similarly, the non-dimensional heat flux is found to decrease relative to laminar non-slip theory, and this decrease is also a function of the contact angle. Depending on the contact angle and the wall shear rate, variations in the heat transfer rate exceeding 10% can be expected. Thus the contact angle is an important consideration in the design of micro, and even more so, nano heat exchangers. (c) 2006 Elsevier Ltd. All rights reserved.

Multidimensional modelling to investigate interspecies hydrogen transfer in anaerobic biofilms

Anaerobic digestion is a multistep process, mediated by a functionally and phylogenetically diverse microbial population. One of the crucial steps is oxidation of organic acids, with electron transfer via hydrogen or formate from acetogenic bacteria to methanogens. This syntrophic microbiological process is strongly restricted by a thermodynamic limitation on the allowable hydrogen or formate concentration. In order to study this process in more detail, we developed an individual-based biofilm model which enables to describe the processes at a microbial resolution. The biochemical model is the ADM1, implemented in a multidimensional domain. With this model, we evaluated three important issues for the syntrophic relationship: (i) is there a fundamental difference in using hydrogen or formate as electron carrier? (ii) Does a thermodynamic-based inhibition function produced substantially different results from an empirical function? and; (iii) Does the physical colocation of acetogens and methanogens follow directly from a general model. Hydrogen or formate as electron carrier had no substantial impact on model results. Standard inhibition functions or thermodynamic inhibition function gave similar results at larger substrate field grid sizes (> 10 mu m), but at smaller grid sizes, the thermodynamic-based function reduced the number of cells with long interspecies distances (> 2.5 mu m). Therefore, a very fine grid resolution is needed to reflect differences between the thermodynamic function, and a more generic inhibition form. The co-location of syntrophic bacteria was well predicted without a need to assume a microbiological based mechanism (e.g., through chemotaxis) of biofilm formation.

Identification of success and failure factors for technology transfer in biotechnology: case studies and surveys from Academic Sinica in Taiwan

Academia Sinica is the leading research institute in Taiwan founded in 1928. Its Office of Technology Transfer (OTT) was established in 1998. It made great efforts to dramatically turn around the technology transfer activity of Academia Sinica, especially in biotechnology. Academia Sinica has more than 80 cases of experience in biotechnology transfer with companies in Taiwanese industry in the past five years. The purpose of this study is to identify potential success and failure factors for biotechnology transfer in Taiwan. Eight cases were studied through in-depth interview. The results of the analysis were used to design two surveys to further investigate 81 cases (48 successful and 33 failure cases) of biotechnology transfer in Academia Sinica from 1999–2003. The results indicated that 10 of the 14 success factors were cited in more than 40% of the cases as contributing to the success of technology transfer. By contrast, only 5 out of 16 key factors were present in more than 30% of the failure cases.

Multilevel phenomena and the implications for method: An example of knowledge transfer in sports

Multilevel theories integrate individual-level processes with those occurring at the level of the firm and above to generate richer and more complete explanations of IB phenomena than the traditional specification of IB relationships as single-level and parsimonious allows. Case study methods permit the timely collection of multiple sources of data, in context, from multiple individuals and multiple organizational units. Further, because the definitions for each level emerge from case data rather than being imposed a priori, case analysis promotes an understanding of deeper structures and cross-level processes. This paper considers the example of sport as an internationalized service to illustrate how the case method might be used to illuminate the multilevel phenomena of knowledge.