Fig. 5

Mutant Myh protein constitutes a small proportion of the total cardiac myosin

Heart homogenates were prepared from control, 131 line, and 140 line mice, separated on a 6% acrylamide gel, transferred to nitrocellulose, and probed with an anti-α Myh-specific monoclonal antibody (BAG5 [22]). The position of the wild-type (Wt) and transgene (Tg) proteins are indicated, as well as a closely migrating breakdown product (bdp). The distinct mobilities of the transgene protein and the breakdown product can be seen clearly in the samples from the 131 line as well as in an artificial mixture of control and 140 line homogenates (mix). Laser densitometry indicates that the mutant species comprises less than 12% of the total myosin (0.6–2.5%, 131 line; 10–12%, 140 line). However, this may be an overestimate due to the impossibility of simultaneously maintaining the wild-type and transgene protein in both the detectable and linear range of the assay. Nevertheless, the data indicate that the relative levels of transgene and endogenous Myh mRNAs do not reflect the levels of mutant protein (Fig. 2).