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The trimethylamine dehydrogenase-electron transferring flavoprotein (TMADH⅐ETF) electron transfer complex has been studied by fluorescence and absorption spectroscopies. These studies indicate that a series of conformational changes occur during the assembly of the TMADH⅐ETF electron transfer complex and that the kinetics of assembly observed with mutant TMADH (Y442F/L/G) or ETF (␣R237A) complexes are much slower than are the corresponding rates of electron transfer in these complexes. This suggests that electron transfer does not occur in the thermodynamically most favorable state (which takes too long to form), but that one or more metastable states (which are formed more rapidly) are competent in transferring electrons from TMADH to ETF. Additionally, fluorescence spectroscopy studies of the TMADH⅐ETF complex indicate that ETF undergoes a stable conformational change (termed structural imprinting) when it interacts transiently with TMADH to form a second, distinct, structural form. The mutant complexes compromise imprinting of ETF, indicating a dependence on the native interactions present in the wild-type complex. The imprinted form of semiquinone ETF exhibits an enhanced rate of electron transfer to the artificial electron acceptor, ferricenium. Overall molecular conformations as probed by smallangle x-ray scattering studies are indistinguishable for imprinted and non-imprinted ETF, suggesting that changes in structure likely involve confined reorganizations within the vicinity of the FAD. Our results indicate a series of conformational events occur during the assembly of the TMADH⅐ETF electron transfer complex, and that the properties of electron transfer proteins can be affected lastingly by transient interaction with their physiological redox partners. This may have significant implications for our understanding of biological electron transfer reactions in vivo, because ETF encounters TMADH at all times in the cell. Our studies suggest that caution needs to be exercised in extrapolating the properties of in vitro interprotein electron transfer reactions to those occurring in vivo.Trimethylamine dehydrogenase (TMADH, 1 EC 1.5.99.7) from the methylotrophic bacterium Methylophilus methylotrophus (sp. W 3 A 1 ) is a homodimeric iron-sulfur flavoprotein that forms an electron transfer complex with electron transferring flavoprotein (ETF (1)). Each subunit of TMADH (molecular mass ϳ 83 kDa) contains a covalently linked 6-S-cysteinyl FMN cofactor, a 4Fe-4S center, and a tightly bound ADP of unknown function (2-8). TMADH catalyzes the oxidative demethylation of trimethylamine to form dimethylamine and formaldehyde (Reaction 1),The mechanism of enzyme reduction by trimethylamine has been addressed by stopped-flow spectroscopy in wild-type (9 -11) and mutant (12-15) TMADH enzymes and has recently been reviewed (16). Following substrate reduction of TMADH, electrons are transferred sequentially from the FMN cofactor to the 4Fe-4S center and, subsequently, in an interprotein electron transfer reaction to the FAD of ...

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An improved model of the screening effect is suggested. Mutual ≪shielding≫ of chromophores from light due to competition for the incident photon can take place in molecular aggregates and macromolecules. From a common point of view, it could be interpreted as in ≪interaction≫ of the absorption dipol moment transitions. Screening leads to a decrease in the extinction coefficient. The largest decrease is observed in the maximum of the absorption band. This is demonstrated with chromophores of adenine, tyrosine, tryptophan, retinol, porphyrin and anthracene. The model enables prediction of hypochromic spectra or evaluation of the quantity of chromophores in an aggregate or macromolecule.