Identification of the glutamine residue that may be involved in the transglutaminase-mediated intramolecular crosslinking of carp and walleye pollack myosin

Abstract

In order to elucidate the molecular mechanism of transglutaminase-mediated myosin crosslinking, a fluorescent monodansylcadaverine (MDC) was incorporated into carp Cyprinus carpio myosin and the reactive Gln residues were analyzed by cyanogen bromide cleavage. The fluorescence was predominantly detected in a 10.5 kDa BrCN fragment, assumed to be located in subfragment 2 of the myosin heavy chain. Furthermore, lysyl endopeptidase digestion of the 10.5 kDa fragment revealed that MDC was specifically incorporated into the 520th Gln residue of the subfragment 2 domain. When meat paste prepared from frozen walleye pollack (Theragra chalcogramma) surimi was incubated with MDC, the fluorescence was mostly observed in a 16 kDa BrCN fragment and also slightly detected in other three bands. By digesting the 16 kDa fragment with lysyl endopeptidase, it was elucidated that MDC was incorporated specifically into Gln-520 of myosin subfragment 2, as also detected in carp. This domain around Gln-520 is likely to be a critical region for the formation of myosin heavy chain dimers that both fish species have in common. In walleye pollack, other reactive Gln residues are presumed to exist at the C-terminus of the light meromyosin. This slight difference may have a significant effect on the capacity of myosin to form tetramers or even larger multimers.