Comparison of the structure and activity of glycosylated and asglycosylated human carboxylesterase 1

Human Carboxylesterase 1 (hCES1) is the key liver microsomal enzyme responsible for detoxification and metabolism of a variety of clinical drugs. To analyse the role of the single N-linked glycan on the structure and activity of the enzyme, authentically glycosylated and aglycosylated hCES1, generated by mutating asparagine 79 to glutamine, were produced in human embryonic kidney cells. Purified enzymes were shown to be predominantly trimeric in solution by analytical ultracentrifugation. The purified aglycosylated enzyme was found to be more active than glycosylated hCES1 and analysis of enzyme kinetics revealed that both enzymes exhibit positive cooperativity. Crystal structures of hCES1 a catalytically inactive mutant (S221A) and the aglycosylated enzyme were determined in the absence of any ligand or substrate to high resolutions (1.86 Å, 1.48 Å and 2.01 Å, respectively). Superposition of all three structures showed only minor conformational differences with a root mean square deviations of around 0.5 Å over all Cα positions. Comparison of the active sites of these un-liganded enzymes with the structures of hCES1-ligand complexes showed that side-chains of the catalytic triad were pre-disposed for substrate binding. Overall the results indicate that preventing N-glycosylation of hCES1 does not significantly affect the structure or activity of the enzyme

Toxic Element Concentrations in the Razorbill Alca torda (Charadriiformes, Alcidae) in Portugal

The present study provides the first data on inorganic element levels (As, Cd, Co, Cr, Cu, Hg, Mn, Pb, Se, and Zn) in juvenile, immature, and adult razorbills (Alca torda) collected along the central coast of Portugal. Element concentrations were assessed by ICP-MS in kidney, liver, muscle, and feathers of 28 razorbills, including 4 juveniles, 17 subadults, and 7 adults. The effect of age and tissue on element accumulation was also assessed. The detected levels in razorbills may indicate a possible contamination risk by Hg and Cr. With respect to bird tissues, higher accumulation of Se and Cd was detected in kidney, Zn and Pb in feathers, and As and Mn in liver. Age was found to affect the accumulation of Cd, Cr, Cu, Hg, and Mn, juveniles presenting higher levels of Cu and Mn than older individuals. In razorbill kidney, Zn-Hg constituted the most significant relationship among metal concentrations. Liver presented the highest number of significant relationships (mostly involving Zn and Co). With regard to feathers, the most significant relationships involved Se, Zn, Cr and Cu concentrations. Positive linear relationships were detected among kidney, liver, and muscle, with emphasis on relationships involving Se and Hg, which may be indicative of similar accumulation/regulation mechanisms in those organs. Element concentrations are discussed in view of possible detoxification mechanisms in seabirds.SEID/CRUP - project HP 2005-0011 (ACIN)FCT/MCTES - SFRH/BPD/27014/200