Molecular insights into substrate recognition and catalysis by phthalate dioxygenase from Comamonas testosteroni

Phthalate, a plasticizer, endocrine disruptor, and potential carcinogen, is degraded by a variety of bacteria. This degradation is initiated by phthalate dioxygenase (PDO), a Rieske oxygenase (RO) that catalyzes the dihydroxylation of phthalate to a dihydrodiol. PDO has long served as a model for understanding ROs despite a lack of structural data. Here we purified PDO_KF1 from Comamonas testosteroni KF1 and found that it had an apparent k_cat/K_m for phthalate of 0.58 ± 0.09 μM⁻¹s⁻¹, over 25-fold greater than for terephthalate. The crystal structure of the enzyme at 2.1 Å resolution revealed that it is a hexamer comprising two stacked α₃ trimers, a configuration not previously observed in RO crystal structures. We show that within each trimer, the protomers adopt a head-to-tail configuration typical of ROs. The stacking of the trimers is stabilized by two extended helices, which make the catalytic domain of PDO_KF1 larger than that of other characterized ROs. Complexes of PDO_KF1 with phthalate and terephthalate revealed that Arg207 and Arg244, two residues on one face of the active site, position these substrates for regiospecific hydroxylation. Consistent with their roles as determinants of substrate specificity, substitution of either residue with alanine yielded variants that did not detectably turnover phthalate. Together, these results provide critical insights into a pollutant-degrading enzyme that has served as a paradigm for ROs and facilitate the engineering of this enzyme for bioremediation and biocatalytic applications.     

Decomposition and nitrogen dynamics of turtle grass (<Emphasis Type="Italic">Thalassia testudinum</Emphasis>) in a subtropical estuarine system

Seagrass beds are pivotal in the functioning of coastal ecosystems in terms of productivity, organic matter turnover and nutrient cycling. Aiming to document decay and nitrogen (N) dynamics of turtle grass (Thalassia testudinum) in a subtropical estuarine system, decomposition patterns of leaves and rhizomes were characterized and compared. Nitrogen usage during decomposition of tissues, and of live tissues and epiphytes growing on live leaves, was also quantified and compared. Stable isotope ratios allowed tracing N within the seagrass bed, following N incorporation into seagrass tissues from the surrounding media (water, sediment). Leaves had a higher N content and decomposed at a faster rate (~6.4 times) compared to rhizomes. Leaching of soluble materials explain the rapid (0–3 days) initial mass loss of leaves (20%) and rhizomes (18%); with a loss of 85 and 56%, respectively, by the end of the study (77 days). Overall, leaves released N while rhizomes immobilized it. Nitrogen concentration was significantly different among live tissues. The main source of N for both seagrass tissues was the sediment, and water column for epiphytes. Differences in decomposition rates among seagrass tissues can be explained by the quality of the substrate and its susceptibility to microbial use. Seagrass leaves and rhizomes are equally important in taking up nutrients from either the water column or the sediments. This study provides a platform to study energy and matter transfers through detrital foodwebs linked to seagrass meadows.     

Long-Term GABA Administration Induces Alpha Cell-Mediated Beta-like Cell Neogenesis

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Mutagens induced chromosomal damage in <Emphasis Type="Italic">Lablab purpureus</Emphasis> (L.) Sweet var. <Emphasis Type="Italic">typicus</Emphasis>

Cytological analysis with respect to meiotic behaviour is considered to be the one of the most dependable indices to estimate the potency of mutagens and to elucidate the response of various genotypes to a particular mutagen. Seeds of Lablab purpureus (L.) Sweet var. typicus cv. CO(Gb)14 were subjected to different doses/concentrations of gamma rays and EMS. The effects of different mutagenic treatments on meiosis were studied on treated and control plants. Various types of meiotic aberrations such as stickiness, clumping of chromosomes, laggards, ring chromosomes and precocious movements were observed in the mutagenic treatments. As increase in the concentration, the frequency of cells showing chromosomal aberrations shows a linear increase up to a certain level. However, the EMS treatments proved to be more effective in inducing meiotic aberrations as compared to gamma rays.