Discovery of Reldesemtiv, a Fast Skeletal Muscle Troponin Activator for the Treatment of Impaired Muscle Function.

The discovery of reldesemtiv, a second-generation fast skeletal muscle troponin activator (FSTA) that increases force production at submaximal stimulation frequencies, is reported. Property-based optimization of high throughput screening hit led to compounds with improved free exposure and muscle activation potency compared to the first-generation FSTA, tirasemtiv. Reldesemtiv demonstrated increased muscle force generation in a phase 1 clinical trial and is currently being evaluated in clinical trials for the treatment of amyotrophic lateral sclerosis.

Middle-down mass spectrometry enables characterization of branched ubiquitin chains.

Protein ubiquitylation, one of the most prevalent post-translational modifications in eukaryotes, is involved in regulating nearly every cellular signaling pathway. The vast functional range of ubiquitylation has largely been attributed to the formation of a diverse array of polymeric ubiquitin (polyUb) chains. Methods that enable the characterization of these diverse chains are necessary to fully understand how differences in structure relate to function.

Nonenzymatic polymerization of ubiquitin: single-step synthesis and isolation of discrete ubiquitin oligomers.

Linked: a method based on thiol-ene chemistry enables the synthesis and purification of ubiquitin oligomers with ≥4 units. This approach, which employs free-radical polymerization, can be applied towards the synthesis of homogeneous Lys6-linked ubiquitin oligomers currently inaccessible by enzymatic methods. By using these chains, one can study their roles in the ubiquitin proteasome system and the DNA damage response pathway.

Forging isopeptide bonds using thiol-ene chemistry: site-specific coupling of ubiquitin molecules for studying the activity of isopeptidases.

Chemical methods for modifying proteins can enable studies aimed at uncovering biochemical function. Herein, we describe the use of thiol-ene coupling (TEC) chemistry to report on the function of branched (also referred to as forked) ubiquitin trimers. We show how site-specific isopeptide (Nε-Gly-L-homothiaLys) bonds are forged between two molecules of Ub, demonstrating the power of TEC in protein conjugation.