CHARACTERIZATION AND MUTATION OF IODOTYROSINE DEIODINASE FROM Haliscomenobacter hydrossis FOR DETOXIFICATION OF IODOPHENOLS

Iodotyrosine deiodinase (IYD) catalyzes deiodination of mono- and diiodotyrosines (I-Tyr and I2-Tyr) to recycle iodide for proper thyroid function in mammals. IYD is also present in all animals and even some bacteria, although their function in lower organisms is not clear. In this study, a representative set of IYD from eukaryotes and prokaryotes was selected to assess their deiodination of iodophenols for bioremediation. Human IYD, bacterial IYD (hhIYD) and archaeal IYD (pfuIYD) exhibit different substrate binding recognition but their catalytic specificity is surprisingly conserved for iodotyrosines. Greater affinity of hhIYD for 2-iodophenol (2IP) compared to human IYD (~20 fold increased in affinity) indicated that interactions from the phenolate anion of 2IP can compensate for the lack of interactions established by the zwitterion of I-Tyr. Deiodination rates of 2IP by hhIYD are still very slow compared to that of I-Tyr suggesting that affinity is not diagnostic of catalytic efficiency. A crystal structure of hhIYD•I-Tyr shows the formation of an active site lid induced by interactions with the zwitterion of I-Tyr. However, the active site lid is not ordered in a structure of hhIYD•2IP indicating that 2IP cannot trigger closure of the active site lid. Reduction of hhIYD in the alternative presence of a substrate analog F-Tyr and 2IP also indicates differences in the ability of the zwitterionic substrate and 2IP to initiate IYD catalysis. A flavin semiquinone (FMNsq) was detected during the reduction of hhIYD in the presence of F-Tyr but accumulation of this same intermediate was not observed in the presence of 2IP. Crystallographic and redox studies demonstrated that 2IP lacks an ability to initiate formation of the active site lid and cannot stabilize the one-electron chemistry for IYD catalysis, which explain its slow deiodination rates. To enable IYD for bioremediation of iodophenols, three hhIYD mutants were generated in hopes of improving the kcat/Km values for iodophenol turnover. However, these mutants did not provide much increase in the catalytic efficiency for deiodination of iodopehnols as their kcat/Km values are ~3 – 17 fold lower than that of the wild-type hhIYD. However, the studies demonstrated a repulsive interaction between Glu91 and the carboxylate of 4-hydroxy-3-iodobenzoate (2IPCOOH) can significantly increase the Km for deiodination of 2IPCOOH. Removal of this repulsive interaction can significantly decrease the Km as evident by a ~4 fold lower Km for deiodination of 2IPCOOH by an E91R mutant compared to the wild type. This result suggests E91 is a potential mutation site for fine-tuning the Km for deiodination of iodophenols. In addition to hhIYD, two IYD-related enzymes (BluB and 3EO8) were selected to examine their native affinity for 2IP. Neither BluB nor 3EO8 bound 2IP with measurable affinity. Further structural analysis of 3EO8 indicated that its active site was relatively small to accommodate 2IP. 3EO8 mutants were therefore generated to enlarge its active site for 2IP coordination. However, the 3EO8 mutants did not improve the affinity for 2IP. A mutant with the highest affinity with 2IP demonstrated only a 20 % increase in affinity compared to the wild-type 3EO8

New Permo-Carboniferous geochemical data from central Thailand: implication for a volcanic arc model

Current ideas and models of geotectonic reconstructions of Southeast Asia are reviewed and new data on Late Carboniferous through Middle Permian tuffites and sills from central Thailand are presented in the light of the problems of Southeast Asian palaeogeography. The volcanic rocks of quartz-keratophyric to spilitic composition are associated with platform carbonates and deep basin sediments. Their geochemistry and the character of the accompanying sediments suggest the existence of a Late Palaeozoic volcanic arc separating a subduction zone in the west from a back arc basin to the east. The geotectonic frame of Southeast Asia is explained in terms of repeated accretion of volcanic arcs by the Late Palaeozoic subduction zone along the northern Tethys margin

The lower jaw of Sunosuchus thailandicus, a mesosuchian crocodilian from the Jurassic of Thailand

Volume: 27Start Page: 199End Page: 20

Sponge assemblage of some Upper Permian reef limestones from Phrae province (Northern Thailand)

The sponge fauna of uppermost Permian reef or reefal limestones of the Phrae province in northern Thailand include representatives of hexactinellida, sclerospongea,"sphinctozoans", and "inozoans". The "sphinctozoans" and "inozoans"are described in detail. Following taxa are new:"Sphinctozoans": Phraethalamia tubulara n. gen., n. sp., Ambithalamia pérmican. gen., n. sp."Inozoans": Bisiphonella tubulara n. sp., Solutossaspongia crassimuralis n.gen., n. sp.The genus name Belyaevaspongia nom. nov. is proposed for PolysiphonellaBelyaeva, 1991 (in Boiko et al., 1991), non Polysiphonella Russo, 1981

Tracing the disrupted outer margin of the Paleoeurasian continent through the Union of Myanmar

Based on stratigraphy, facies distribution and paleontology of upper Paleozoic and Triassic strata in Malaysia, Thailand, Myanmar and Yunnan (China), the location of division between the outer margin of the disrupted Paleoeurasian continent and possible Gondwana-derived terranes is discussed. It is proposed that this division is located much further to the west than that has usually been maintained

Active Site Binding Is Not Sufficient for Reductive Deiodination by Iodotyrosine Deiodinase

The minimal requirements for substrate recognition and turnover by iodotyrosine deiodinase were examined to learn the basis for its catalytic specificity. This enzyme is crucial for iodide homeostasis and the generation of thyroid hormone in chordates. 2-Iodophenol binds only very weakly to the human enzyme and is dehalogenated with a <i>k</i>_cat/<i>K</i>_m that is more than 4 orders of magnitude lower than that for iodotyrosine. This discrimination likely protects against a futile cycle of iodinating and deiodinating precursors of thyroid hormone biosynthesis. Surprisingly, a very similar catalytic selectivity was expressed by a bacterial homologue from <i>Haliscomenobacter hydrossis</i>. In this example, discrimination was not based on affinity since 4-cyano-2-iodophenol bound to the bacterial deiodinase with a <i>K</i>_d lower than that of iodotyrosine and yet was not detectably deiodinated. Other phenols including 2-iodophenol were deiodinated but only very inefficiently. Crystal structures of the bacterial enzyme with and without bound iodotyrosine are nearly superimposable and quite similar to the corresponding structures of the human enzyme. Likewise, the bacterial enzyme is activated for single electron transfer after binding to the substrate analogue fluorotyrosine as previously observed with the human enzyme. A cocrystal structure of bacterial deiodinase and 2-iodophenol indicates that this ligand stacks on the active site flavin mononucleotide (FMN) in a orientation analogous to that of bound iodotyrosine. However, 2-iodophenol association is not sufficient to activate the FMN chemistry required for catalysis, and thus the bacterial enzyme appears to share a similar specificity for halotyrosines even though their physiological roles are likely very different from those in humans