Oligomeric Status and Nucleotide Binding Properties of the Plastid ATP/ADP Transporter 1: Toward a Molecular Understanding of the Transport Mechanism

Background: Chloroplast ATP/ADP transporters are essential to energy homeostasis in plant cells. However, their molecular mechanism remains poorly understood, primarily due to the difficulty of producing and purifying functional recombinant forms of these transporters. Methodology/Principal Findings: In this work, we describe an expression and purification protocol providing good yields and efficient solubilization of NTT1 protein from Arabidopsis thaliana. By biochemical and biophysical analyses, we identified the best detergent for solubilization and purification of functional proteins, LAPAO. Purified NTT1 was found to accumulate as two independent pools of well folded, stable monomers and dimers. ATP and ADP binding properties were determined, and Pi, a co-substrate of ADP, was confirmed to be essential for nucleotide steady-state transport. Nucleotide binding studies and analysis of NTT1 mutants lead us to suggest the existence of two distinct and probably inter-dependent binding sites. Finally, fusion and deletion experiments demonstrated that the C-terminus of NTT1 is not essential for multimerization, but probably plays a regulatory role, controlling the nucleotide exchange rate. Conclusions/Significance: Taken together, these data provide a comprehensive molecular characterization of a chloroplas

Characterization of aerosols released during handling or processing operations of MWCNTs

The goal of this study was to characterize the release at the source and potential for exposure to multi-walled carbon nanotubes (MWCNTs) in an industrial pilot plant. Various operations had been investigated: cleaning of the gas phase reactor, filling bins with MWCNTs powder, hot compounding of MWCNT-containing polymer composites and dry cutting of the composites into mm-sized pellets. In addition, manual transferring and sieving of MWCNTs have been examined with the intention of investigating a worst case potential exposure scenario. The activities were operated in normal conditions, with the exception of the simulation of the worst case scenario of manual transferring and weighing for which extreme precautionary principles were adopted. Background conditions were determined in parallel. Overall particle concentration levels, size distributions were not significantly different for the reactor cleanout and filling bins operations from the background aerosol that could be attributed to the outside air that fed the workplace atmosphere. The hot compounding and dry cutting processes resulted in particle concentration levels and size distributions significantly different from the background. Besides polymer fragments and carbonaceous aggregates, few possible polymer fragments containing MWCNTs were observed on EM micrographs of aerosol samples collected close to the source. The results obtained during the manual transferring and sieving as a worst case potential exposure scenario demonstrate an association between the activity and the increase of the particle concentration and the modification of the size distribution. EM micrographs indicated that MWCNTs existed mainly as bundles whose size ranged from approximately a few 100 nm to 10 µm