Mapping Interactions Between The Chaperone Domain Of Unc-45b And Myosin

Force generation in striated muscle comes from the myosin motor domain, an 110kDa globular protein that allows conversion of the chemical potential energy in ATP into mechanical work. This complex protein is incapable of self-folding and assembly. Instead, molecular chaperones work in a precise network to allow a nascent myosin polypeptide to be protected from aggregation and folded into its native and functional conformation. The UNC-45B chaperone is associated with the proper folding and function of the sarcomeric myosin. Hence, deciphering the precise mechanisms by which the chaperone UNC-45B helps to fold myosin heads is a challenge at the core of muscle development and function. Which are the key regions in UNC-45B mediate its myosin-chaperonin function? Guided by the available crystal structures, biophysical, biochemical data we have mapped putative key amino acid regions in UNC-45B that may mediate its myosin-binding and chaperoning functions. We are using a combination of complementary assays that probe effect of mutations on UNC-45B: i) structure (by circular dichroism and Bis-ANS fluorescence); ii) binding to myosin (by using a myosin-chaperone fluorescence assay) and iii) chaperone function (by using a light scattering assay for myosin aggregation). Together, these biophysical studies have started to shed light upon the critical centers for UNC-45B function, which may serve as target sites for future drug development.