Identification of the active site residues in ATP‐citrate lyase's carboxy‐terminal portion

ATP‐citrate lyase (ACLY) catalyzes production of acetyl‐CoA and oxaloacetate from CoA and citrate using ATP. In humans, this cytoplasmic enzyme connects energy metabolism from carbohydrates to the production of lipids. In certain bacteria, ACLY is used to fix carbon in the reductive tricarboxylic acid cycle. The carboxy(C)‐terminal portion of ACLY shows sequence similarity to citrate synthase of the tricarboxylic acid cycle. To investigate the roles of residues of ACLY equivalent to active site residues of citrate synthase, these residues in ACLY from Chlorobium limicola were mutated, and the proteins were investigated using kinetics assays and biophysical techniques. To obtain the crystal structure of the C‐terminal portion of ACLY, full‐length C. limicola ACLY was cleaved, first non‐specifically with chymotrypsin and subsequently with Tobacco Etch Virus protease. Crystals of the C‐terminal portion diffracted to high resolution, providing structures that show the positions of active site residues and how ACLY tetramerizes.     

The effects of temperature, pH and sulphide on the community structure of hyperthermophilic streamers in hot springs of northern Thailand

Hyperthermophilic community diversity was assessed in hot-spring streamers along gradients of temperature, pH and sulphide in northern Thailand. A hierarchical sampling design was employed to obtain biomass for culture-independent estimates of 16S rRNA gene-defined prokaryotic diversity. All springs supported several archaeal and bacterial phylotypes, including novel phylotypes that expand the known phylogenetic diversity of terrestrial hyperthermophiles. Diversity appeared significantly greater than that observed for several other geographic locations. Phylotypes belonging to the Aquificales were ubiquitous, further supporting the hypothesis that these chemolithoautotrophs are key members of all hyperthermophilic communities. The chemoorganotrophic genus Thermus was also represented by phylotypes in all springs. Other bacterial taxa represented by environmental sequences included Bacillus, Thermotoga and various unidentified Alphaproteobacteria and Betaproteobacteria. Archaeal phylotypes included the Crenarchaea Desulfurococcus, Pyrobaculum, plus several unidentified hyperthermophilic lineages. A Methanothermococcus-like Euryarchaeon was also identified, with this genus not previously known from streamer communities. A multivariate approach to the analysis of biotic and abiotic data revealed that diversity patterns were best explained by a combination of temperature and sulphide rather than by any other abiotic variable either individually or in combination.     

Synthesis and structure evaluation of a novel cantharimide and its cytotoxicity on SK-Hep-1 hepatoma cells

A remarkable control of the potency of cantharimide is described based on the electronic properties of functional group and it exhibits a relatively less toxic effect to the non-malignant hematological disorder bone marrow cells.     

The problem of assessing landmark error in geometric morphometrics: theory, methods, and modifications

Geometric morphometric methods rely on the accurate identification and quantification of landmarks on biological specimens. As in any empirical analysis, the assessment of inter- and intra-observer error is desirable. A review of methods currently being employed to assess measurement error in geometric morphometrics was conducted and three general approaches to the problem were identified. One such approach employs Generalized Procrustes Analysis to superimpose repeatedly digitized landmark configurations, thereby establishing whether repeat measures fall within an acceptable range of variation. The potential problem of this error assessment method (the "Pinocchio effect") is demonstrated and its effect on error studies discussed. An alternative approach involves employing Euclidean distances between the configuration centroid and repeat measures of a landmark to assess the relative repeatability of individual landmarks. This method is also potentially problematic as the inherent geometric properties of the specimen can result in misleading estimates of measurement error. A third approach involved the repeated digitization of landmarks with the specimen held in a constant orientation to assess individual landmark precision. This latter approach is an ideal method for assessing individual landmark precision, but is restrictive in that it does not allow for the incorporation of instrumentally defined or Type III landmarks. Hence, a revised method for assessing landmark error is proposed and described with the aid of worked empirical examples.     

Combinatorial mixture synthesis and biological evaluation of dihydrophenyl triazine antifolates.

The traditional 'one-pot' three component synthesis was adapted successfully for combinatorial mixtures synthesis of dihydrophenyl triazines, which are nonclassical, dihydrofolate reductase (DHFR) inhibitors. Each library was designed to comprise eight reaction mixture pots and in every pot there were three dihydrophenyl triazines. A total of three libraries were synthesized and the final number of compounds harvested was 64. The products precipitated out of the reaction mixture and could be collected easily and cleansed by washing. Solid supports and further purification processes were not required. The reactions were monitored by TLC and a HPLC method was developed to determine the number of products in each pot. All 24 pots were screened for inhibitory activity against the rat liver DHFR. Two pots showed good inhibitory activity and the products in them were individually synthesized, characterized and biologically tested again. One lead compound was identified amongst all the compounds synthesized, and would be further optimized.