Massively parallel analysis of single-molecule dynamics on next-generation sequencing chips.

Single-molecule techniques are ideally poised to characterize complex dynamics but are typically limited to investigating a small number of different samples. However, a large sequence or chemical space often needs to be explored to derive a comprehensive understanding of complex biological processes. Here we describe multiplexed single-molecule characterization at the library scale (MUSCLE), a method that combines single-molecule fluorescence microscopy with next-generation sequencing to enable highly multiplexed observations of complex dynamics.

Spatiotemporally controlled generation of NTPs for single-molecule studies.

Many essential processes in the cell depend on proteins that use nucleoside triphosphates (NTPs). Methods that directly monitor the often-complex dynamics of these proteins at the single-molecule level have helped to uncover their mechanisms of action. However, the measurement throughput is typically limited for NTP-utilizing reactions, and the quantitative dissection of complex dynamics over multiple sequential turnovers remains challenging.

Direct Measurements of Erythromycin's Effect on Protein Synthesis Kinetics in Living Bacterial Cells.

Macrolide antibiotics, such as erythromycin, bind to the nascent peptide exit tunnel (NPET) of the bacterial ribosome and modulate protein synthesis depending on the nascent peptide sequence. Whereas in vitro biochemical and structural methods have been instrumental in dissecting and explaining the molecular details of macrolide-induced peptidyl-tRNA drop-off and ribosome stalling, the dynamic effects of the drugs on ongoing protein synthesis inside live bacterial cells are far less explored. In the present study, we used single-particle tracking of dye-labeled tRNAs to study the kinetics of mRNA translation in the presence of erythromycin, directly inside live Escherichia coli cells.

Real-time measurements of aminoglycoside effects on protein synthesis in live cells.

The spread of antibiotic resistance is turning many of the currently used antibiotics less effective against common infections. To address this public health challenge, it is critical to enhance our understanding of the mechanisms of action of these compounds. Aminoglycoside drugs bind the bacterial ribosome, and decades of results from in vitro biochemical and structural approaches suggest that these drugs disrupt protein synthesis by inhibiting the ribosome's translocation on the messenger RNA, as well as by inducing miscoding errors.

Effects of Agave Fructans, Inulin, and Starch on Metabolic Syndrome Aspects in Healthy Wistar Rats.

Healthy Wistar rats were supplemented during 20 weeks with commercial inulin (I) and fructans (CAT), experimental fructans from (EAT) and (AS) mature stems, rice starch 10% (RS), and standard feed for rodents (C). Feed intake was kept steady, but with I, body weight and abdominal adipose tissue (6.01 g) decreased at the end.