RESUMO
Fragment-based lead discovery has proven to be a powerful method in the drug discovery process. The combinatorial output that is accessible by combining fragments is very attractive; however, identifying fragment pairs that bind synergistically and linking them productively can be challenging. Several technologies have now been established to prepare and screen nucleic acid-encoded libraries (ssDNA, dsDNA, PNA), and it has been shown that pairs of molecules combined by hybridization can bind synergistically to a target. Herein we apply this concept to combinatorially pair two libraries of small molecule fragments, use the fittest fragments supplemented with closely related analogs to build a focused library covalently linking the fragments with different spacers, and apply this strategy to the discovery of a potent ligand for Hsp70.
RESUMO
AIMS: To develop a strategy to increase the stability of transcripts of structural genes expressed under the control of sacR, the leader region of Bacillus subtilis levansucrase gene. METHODS AND RESULTS: Insertion of Shine Dalgarno like sequences in the 5'-untranslated sacR region controlling the expression of sacB. Depending on the number of stabilizing sequences inserted and the position of these sequences with respect to the translation start codon, it was observed that the mRNA stability and the final protein production could be increased or decreased. CONCLUSIONS: This mRNA stabilization can be used to increase exocellular protein production in the degU32 (Hy) mutant. SIGNIFICANCE AND IMPACT OF THE STUDY: This approach can be applied to the expression of heterologous genes of biotechnological interest.
