Why labelling the ‘fast-reacting thiols’ of myosin reduces force output: re-assessing a year at the University of California San Francisco (1987-8)

Abstract The fast-reacting thiol in the SH1 helix of myosin was often used to attach spin or fluorescent probes to the motor domain, to readout myosin orientation during the 1980s. However, at this time, we found that labelling this residue (Cys707) in myosin in skinned rabbit psoas fibres, using iodoacetacetamidotetramethyl rhodamine (IATR), reduced force output during active contraction. With access to recent pre- and post-powerstroke actomyosin structures, we can now explain these results. We modelled IATR onto the equivalent SH1 cysteine residue in the pre-power and post-power structures of myosin 5. This revealed that labelling of the fast-reacting thiol would interfere with the structural changes in the relay and SH1 helices required to generate the power stroke, explaining why force output reduces.