Computer entry and verification of nuclei acid sequences

The identification of mutant genes in plasmid clones of synthetic DNAs.How to avoid errors that are frequent in synthetic DNAs and RNAs.Supported by an extramural research grant AI10257 from the National Institutes of Health, USA. in synthetic DNA gene sequences

The structure of an AspRS–tRNA(Asp) complex reveals a tRNA-dependent control mechanism

The 2.6 Å resolution crystal structure of an inactive complex between yeast tRNA(Asp) and Escherichia coli aspartyl-tRNA synthetase reveals the molecular details of a tRNA-induced mechanism that controls the specificity of the reaction. The dimer is asymmetric, with only one of the two bound tRNAs entering the active site cleft of its subunit. However, the flipping loop, which controls the proper positioning of the amino acid substrate, acts as a lid and prevents the correct positioning of the terminal adenosine. The structure suggests that the acceptor stem regulates the loop movement through sugar phosphate backbone– protein interactions. Solution and cellular studies on mutant tRNAs confirm the crucial role of the tRNA three-dimensional structure versus a specific recognition of bases in the control mechanism

Orientational control of fimE expression in Escherichia coli.

Phase-variable expression of type 1 fimbriae is, in part, controlled by site-specific DNA inversion of the fim switch in Escherichia coli. Of the two fim recombinases (FimB and FimE) that catalyse the inversion reaction, FimE exhibits a strong bias for phase switching from the ON to the OFF orientation. The specificity associated with fimE is the result of two different mechanisms: (i) FimE exhibits a preference for the invertible element in the ON orientation as substrate for recombination; (ii) the invertible element in the OFF orientation acts in cis to inhibit recombinase activity (orientational control). We show here that the invertible element negatively regulates fimE, even though expression of a fimE-lacZYA transcriptional fusion is unaffected by orientational control. The fimE transcript extends into the invertible region and hence switch ON-specific and switch OFF-specific mRNA contain different sequences. Furthermore, we show that orientational control is suppressed by the insertion of a structured RNA (tRNA(Gly)) between fimE and the fim switch, indicating that the switch OFF-specific mRNA is inactivated by 3' to 5' degradation. Analysis of the fim switch reveals that it contains two inhibitory elements that exert orientational control independently