Publications (7) View all
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Article: Amicyanin transfers electrons from methylamine dehydrogenase to cytochrome c-551i via a ping-pong mechanism, not a ternary complex.
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ABSTRACT: The first crystal structure of a ternary redox protein complex was comprised of the enzyme methylamine dehydrogenase (MADH) and two electron transfer proteins, amicyanin and cytochrome c-551i from Paracoccus denitrificans [Chen et al. Science 1994, 264, 86-90]. The arrangement of the proteins suggested possible electron transfer from the active site of MADH via the amicyanin copper ion to the cytochrome heme iron, although the distance between the metals is large. We studied the interactions between these proteins in solution. A titration followed by NMR spectroscopy shows that amicyanin binds cytochrome c-551i. The interface comprises the hydrophobic and positive patches of amicyanin, not the binding site observed in the ternary complex. NMR experiments further show that amicyanin binds tightly to MADH with an interface that matches the one observed in the crystal structure and that mostly overlaps with the binding site for cytochrome c-551i. Upon addition of cytochrome c-551i, no changes in the NMR spectrum of MADH-bound amicyanin are observed, suggesting that a possible interaction of the cytochrome with the binary complex must be very weak, with a dissociation constant higher than 2 mM. Reconstitution of the entire redox chain in vitro demonstrates that amicyanin can react rapidly with cytochrome c-551i, but that association of amicyanin with MADH inhibits this reaction. It is concluded that electron transfer from MADH to cytochrome c-551i does not involve a ternary complex but occurs via a ping-pong mechanism in which amicyanin uses the same interface for the reactions with MADH and cytochrome c-551i.Journal of the American Chemical Society 10/2010; 132(41):14537-45. · 9.91 Impact Factor -
Article: Nicotine determination in mushrooms by LC-MS/MS with preliminary studies on the impact of drying on nicotine formation.
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ABSTRACT: A problem concerning significant amounts of nicotine in dried wild mushrooms (mainly Boletus edulis from China) has been reported to the European Commission. As a consequence, the European Food Safety Authority (EFSA) proposed temporary maximum residue levels (MRLs) of 0.036 mg kg(-1) for fresh wild mushrooms and 1.17 mg kg(-1) for dried wild mushrooms (2.3 mg kg(-1) for dried ceps only). The EFSA also highlighted the necessity for a monitoring and testing programme to be launched by food business operators at the start of the 2009 harvest season. In the present study, a quick and sensitive analytical method for routine analysis of nicotine in fresh and dried mushrooms was developed and validated by a single-laboratory procedure. The method, which employs an LC-MS/MS system and (+/-)-nicotine-d(4) as internal standard, has a limit of quantification of 6 and 60 microg kg(-1) for fresh and dried product, respectively. Analyses of samples spiked with different levels of nicotine showed recoveries ranging from 107 to 122%, with relative standard deviations of 2.9-10.1% depending on the spiking level. The combined uncertainties, calculated at a low level for frozen (0.015 mg kg(-1)) and a high level for the dried (2 mg kg(-1)) matrix, were 13 and 10%, respectively. Application of the method to real samples of mushrooms purchased on the market or obtained from local producers showed nicotine levels ranging 0.01-0.04 and 0.1-4.5 mg kg(-1) in fresh/frozen and dried matrices, respectively. To establish reasons for the unexpectedly high levels of the nicotine in dried matrices, preliminary laboratory experiments involving drying mushrooms were performed under various conditions.Food Additives and Contaminants - Part A Chemistry, Analysis, Control, Exposure and Risk Assessment 04/2010; 27(4):473-7. -
Article: Rapid Screening of Alicyclobacillus acidoterrestris Spoilage of Fruit Juices by Electronic Nose: A Confirmation Study
Journal of Sensors. 01/2010; -
Article: Search for a more adequate test to predict the long-term migration from the PVC gaskets of metal lids into oily foods in glass jars.
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ABSTRACT: As shown previously, the conventional testing procedure for simulating long-term migration from the gaskets of metal closures into oily foods does not adequately reflect reality. It appears to be impossible to accelerate migration to the extent that the situation at the end of the shelf life of a product can be anticipated in a few days or weeks. Therefore, we investigated whether long-term migration could be extrapolated from migration rates determined for new lids. Jars were kept in the normal upright position. Since heat treatment may have a strong temporary impact, migration during the initial heating for pasteurization or sterilization and storage at ambient temperature were determined using different lids. Commercial products were recalled from sales points throughout Europe to determine the real migration over extended periods of time and for jars with differing histories. This migration was compared with data from the short-term testing to investigate whether an empirical relationship could be derived. The results show that the short-term test enables the comparison of lids and plasticizers in the initial phase of migration, but that long-term extrapolation presupposes more complex kinetic modeling. The results also demonstrate that the legal relevance of "official" testing methods should be reconsidered to avoid conflict when food contact materials comply with migration limits in the test but not in actual application.Food Additives and Contaminants - Part A Chemistry, Analysis, Control, Exposure and Risk Assessment 08/2009; 26(7):1113-22. -
Article: Structural Comparison of Crystal and Solution States of the 138 kDa Complex of Methylamine Dehydrogenase and Amicyanin from Paracoccus versutus.
Biochemistry 04/2009; · 3.42 Impact Factor
