Destabilizing the Super Relaxed State of Skeletal Muscle Myosin to Treat Obesity and Type 2 Diabetes

In resting skeletal muscle, myosin is found in 2 states, the super-relaxed state (SRX) and the disordered relaxed state (DRX). In the SRX the ATPase activity is strongly inhibited by binding of the myosin heads to the core of the thick filament; in the DRX the myosin heads are not bound to the core of the thick filament and have an ATPase rate that is an order of magnitude greater. We made a series of single cysteine mutants of the myosin regulatory light chain, placed fluorescent probes on them, and exchanged them into skinned rabbit fast skeletal muscle fibers. We found fluorescent probes that reported on the relative populations of the SRX and DRX, and used them to carry out a high throughput screen. The screen identified one compound, piperine, which destabilized the SRX as measured by probe fluorescence. The kd for inhibition was ∼3 μM with a maximum inhibition of ∼50% at high concentations. The compound was also shown to destabilize the SRX using single nucleotide turnover measurements. The effect was only seen in fast twitch fibers. Piperine was shown to increase the ATPase activity of relaxed fibers by 40%. It had no effect on the mechanics of either active or resting muscle fibers. Previous work has shown that piperine can attenuate both obesity and type 2 diabetes in rodent models of these conditions. We propose that these effects are due to the up-regulation of resting muscle metabolism. The results described here suggest that piperine would provide a useful lead compound for the development of therapies to treat obesity and type 2 diabetes in humans.