The de novo engineering of pyrrolysyl-tRNA synthetase for genetic incorporation of l-phenylalanine and its derivatives

2011 ◽  
Vol 7 (3) ◽  
pp. 714 ◽  
Author(s):  
Yane-Shih Wang ◽  
William K. Russell ◽  
Zhiyong Wang ◽  
Wei Wan ◽  
Lindsey E. Dodd ◽  
...  
eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Harrison Tudor Evans ◽  
Liviu-Gabriel Bodea ◽  
Jürgen Götz

The formation of spatial long-term memory (LTM) requires the de novo synthesis of distinct sets of proteins; however, a non-biased examination of the de novo proteome in this process is lacking. Here, we generated a novel mouse strain, which enables cell-type-specific labelling of newly synthesised proteins with non-canonical amino acids (NCAAs) by genetically restricting the expression of the mutant tRNA synthetase, NLL-MetRS, to hippocampal neurons. By combining this labelling technique with an accelerated version of the active place avoidance task and bio-orthogonal non-canonical amino acid tagging (BONCAT) followed by SWATH quantitative mass spectrometry, we identified 156 proteins that were altered in synthesis in hippocampal neurons during spatial memory formation. In addition to observing increased synthesis of known proteins important in memory-related processes, such as glutamate receptor recycling, we also identified altered synthesis of proteins associated with mRNA splicing as a potential mechanism involved in spatial LTM formation.


2014 ◽  
Vol 9 (5) ◽  
pp. 1092-1096 ◽  
Author(s):  
Han Xiao ◽  
Francis B. Peters ◽  
Peng-Yu Yang ◽  
Sean Reed ◽  
Johnathan R. Chittuluru ◽  
...  

2019 ◽  
Author(s):  
Thomas A. Scott ◽  
Sibyl F. Batey ◽  
Patrick Wiencek ◽  
Govind Chandra ◽  
Silke Alt ◽  
...  

AbstractTo meet the ever-growing demand of antibiotic discovery, new chemical matter and antibiotic targets are urgently needed. Many potent natural product antibiotics which were previously discarded can also provide lead molecules and drug targets. One such example is the structurally unique β-lactone obafluorin, produced by Pseudomonas fluorescens ATCC 39502. Obafluorin is active against both Grampositive and -negative pathogens, however the biological target was unknown. We now report that obafluorin targets threonyl-tRNA-synthetase and we identify a homologue, ObaO, which confers self-immunity to the obafluorin producer. Disruption of obaO in P. fluorescens ATCC 39502 results in obafluorin sensitivity, whereas expression in sensitive E. coli strains confers resistance. Enzyme assays demonstrate that E. coli threonyl-tRNA synthetase is fully inhibited by obafluorin, whereas ObaO is only partly susceptible, exhibiting a very unusual partial inhibition mechanism. Altogether, our data highlight the utility of a self-immunity guided approach for the identification of an antibiotic target de novo and will ultimately enable the generation of improved obafluorin variants.


2012 ◽  
Vol 8 (2) ◽  
pp. 405-415 ◽  
Author(s):  
Yane-Shih Wang ◽  
Xinqiang Fang ◽  
Hsueh-Ying Chen ◽  
Bo Wu ◽  
Zhiyong U. Wang ◽  
...  

2021 ◽  
Author(s):  
Chloe M. Jones ◽  
D. Miklos Robkis ◽  
Robert J. Blizzard ◽  
Mika Munari ◽  
Yarra Venkatesh ◽  
...  

Acridonylalanine (Acd) is photostable, with a high quantum yield and long fluorescence lifetime in water. An evolved tRNA synthetase (RS) enables genetic incorporation of Acd in mammalian cells and its use in fluorescence lifetime imaging microscopy.


2010 ◽  
Vol 76 (20) ◽  
pp. 6769-6777 ◽  
Author(s):  
Anna M. Kielak ◽  
Johannes A. van Veen ◽  
George A. Kowalchuk

ABSTRACT The bacterial phylum Acidobacteria has a widespread distribution and is one of the most common and diverse phyla in soil habitats. However, members of this phylum have often been recalcitrant to cultivation methods, hampering the study of this presumably important bacterial group. In this study, we used a cultivation-independent metagenomic approach to recover genomic information from soilborne members of this phylum. A soil metagenomic fosmid library was screened by PCR targeting acidobacterial 16S rRNA genes, facilitating the recovery of 17 positive clones. Recovered inserts appeared to originate from a range of Acidobacteria subdivisions, with dominance of subdivision 6 (10 clones). Upon full-length insert sequencing, gene annotation identified a total of 350 open reading frames (ORFs), representing a broad range of functions. Remarkably, six inserts from subdivision 6 contained a region of gene synteny, containing genes involved in purine de novo biosynthesis and encoding tRNA synthetase and conserved hypothetical proteins. Similar genomic regions had previously been observed in several environmental clones recovered from soil and marine sediments, facilitating comparisons with respect to gene organization and evolution. Comparative analyses revealed a general dichotomy between marine and terrestrial genes in both phylogeny and G+C content. Although the significance of this homologous gene cluster across subdivision 6 members is not known, it appears to be a common feature within a large percentage of all acidobacterial genomic fragments recovered from both of these environments.


2021 ◽  
Vol 65 (5) ◽  
Author(s):  
Uday S. Ganapathy ◽  
Rubén González del Rio ◽  
Mónica Cacho-Izquierdo ◽  
Fátima Ortega ◽  
Joël Lelièvre ◽  
...  

ABSTRACT Global infections by nontuberculous mycobacteria (NTM) are steadily rising. New drugs are needed to treat NTM infections, but the NTM drug pipeline remains poorly populated and focused on repurposing or reformulating approved antibiotics. We sought to accelerate de novo NTM drug discovery by testing advanced compounds with established activity against Mycobacterium tuberculosis. 3-Aminomethyl 4-halogen benzoxaboroles, a novel class of leucyl-tRNA synthetase inhibitors, were recently discovered as active against M. tuberculosis. Here, we report that the benzoxaborole EC/11770 is not only a potent antitubercular agent but is active against the M. abscessus and M. avium complexes. Focusing on M. abscessus, which causes the most-difficult-to-cure NTM disease, we show that EC/11770 retained potency against drug-tolerant biofilms in vitro and was effective in a mouse lung infection model. Resistant mutant selection experiments showed a low frequency of resistance and confirmed leucyl-tRNA synthetase as the target. This work establishes the benzoxaborole EC/11770 as a novel preclinical candidate for the treatment of NTM lung disease and tuberculosis and validates leucyl-tRNA synthetase as an attractive target for the development of broad-spectrum antimycobacterials.


Sign in / Sign up

Export Citation Format

Share Document