Synthetic biology

Engineering Escherichia coli with new functions

Wang et al. present multiplex automated genome engineering (MAGE), a method to rapidly generate combinatorial genetic modifications by repeatedly introducing synthetic DNA to cells. The approach is based on the concept that for a pool of degenerate oligonucleotides, those with greater homology to the chromosomal target will be incorporated with higher frequency via allelic replacement. They applied MAGE to program E. coli to overproduce the antioxidant lycopene by optimizing the biosynthetic pathway.

Wang, H. H. et al. Nature 460, 894–898 (2009).

Chemical biology

Multiplexed kinase activity profiling

Protein kinases have differential levels of activity under different biological conditions. Yu et al. describe an approach to profile the activation state of kinases in a cell lysate by monitoring the phosphorylation of 90 synthetic known peptide substrates via mass spectrometry, using heavy isotope–labeled versions of the peptides as internal quantitative standards. They applied the method to profile kinase activity during the cell cycle, in breast cancer cell lines and after applying kinase inhibitors.

Yu, Y. et al. Proc. Natl. Acad. Sci. USA 106, 11606–11611 (2009).

Nanotechnology

Fluorescent-plasmonic nanoparticles

Jin and Gao describe an approach to make multifunctional nanoparticles that are both fluorescent and plasmonic. This presented a challenge because gold, the plasmonic material, can quench fluorescence. Jin and Gao overcame this by precisely controlling the spacing between a quantum dot core and an ultrathin gold shell via layer-by-layer assembly.

Jin, Y. & Gao, X. Nat. Nanotechnol. advance online publication (26 July 2009).

Proteomics

The Dub interactome

Although the ubiquitin conjugation machinery is fairly well understood, the functions of deubiquitinating enzymes (Dubs), which catalyze the removal of ubiquitin from proteins, have not been vastly studied. Sowa et al. profiled the human Dub protein family using an affinity purification scheme followed by mass spectrometry to analyze Dub protein interaction partners. They also introduce CompPASS, a software platform for identifying high-confidence interactions.

Sowa, M.E. et al. Cell 138, 389–403 (2009).

Biophysics

Transcription initiation on a single-molecule scale

Single-molecule fluorescence resonance energy transfer can be used to monitor molecular motions over time, but multiplexing is limited. Sorokina et al. describe a single-molecule method that requires only a single fluorophore, using time correlated single-photon counting for monitoring fluorescence lifetime trajectories of an immobilized molecule. They used this approach to follow the complex process of transcription initiation by T7 RNA polymerase.

Sorokina, M. et al. J. Am. Chem. Soc. 131, 9630–9631 (2009).