Specific substitution into the anticodon loop of yeast tyrosine transfer RNA

The aminoacylation kinetics of 19 different variants of yeast tRNATyr with nucleotide substitutions in positions 33-35 were determined. Substitution of the conserved uridine-33 does not alter the rate of aminoacylation. However, substitution of the anticodon position 34 or position 35 reduces Km from 2- to 10-fold and Vmax as much as 2-fold, depending on the nucleotide inserted. The ochre and amber suppressor tRNAsTyr both showed about a 7-fold reduction in Vmax/Km. Data from tRNATyr with different modified nucleotides at position 35 suggest that specific hydrogen bonds form between the synthetase and both the N1 and N3 hydrogens of psi-35. The effect of simultaneous substitutions at positions 34 and 35 can be predicted reasonably well by combining the effects of single substitutions. These data suggest that yeast tyrosyl-tRNA synthetase interacts with positions 34 and 35 of the anticodon of tRNATyr and opens the possibility that nonsense suppressor efficiency may be mediated by the level of aminoacylation.     

Comparison of the emulsion characteristics of Rhodococcus erythropolis and Escherichia coli SOXC-5 cells expressing biodesulfurization genes

Biodesulfurization of fuel oils is a two-phase (oil/water) process which may offer an interesting alternative to conventional hydrodesulfurization due to the mild operating conditions and reaction specificity afforded by the biocatalyst. For biodesulfurization to realize commercial success, a variety of process considerations must be addressed including reaction rate, emulsion formation and breakage, biocatalyst recovery, and both gas and liquid mass transport. This study evaluates emulsion formation and breakage using two biocatalysts with differing hydrophobic characteristics. A Gram-positive (Rhodococcus erythropolis) biocatalyst, expressing the complete 4S desulfurization pathway, and a Gram-negative biocatalyst (Escherichia coli), expressing only the gene for conversion of dibenzothiophene (DBT) to DBT sulfone, are compared relative to their ability to convert DBT and the ease of phase separation as well as biocatalyst recovery following desulfurization.