Regulation of α-Helical Coiled-coil Dimerization in Chicken Skeletal Muscle Light Meromyosin

Abstract

The dimerization specificity of the light meromyosin (LMM) domain of chicken neonatal and adult myosin isoforms was analyzed by metal chelation chromatography. Our results show that neonatal and adult LMMs associate preferentially, although not exclusively, as homodimeric coiled-coils. Using chimeric LMM constructs combining neonatal and adult sequences, we observed that a stretch of 183 amino acids of sequence identity at the N terminus of the LMM was sufficient to allow the adult LMM to dimerize in a non-selective manner. In contrast, sequence identity in the remaining C-terminal 465 amino acids had only a modest effect on the dimerization selectivity of the adult isoform. Sequence identity at the N terminus also promoted dimerization of the neonatal LMM to a greater degree than sequence identity at the C terminus. However, the N terminus had only a partial effect on the dimerization specificity of the neonatal sequence, and residues distributed throughout the LMM were capable of affecting dimerization selectivity of this isoform. These results indicated that dimerization preference of the neonatal and adult isoforms was affected to a different extent by sequence identity at a given region of the LMM.

The carboxyl-terminal isoforms of smooth muscle myosin heavy chain determine thick filament assembly properties

“…Homodimeric myosins are also preferentially formed in invertebrates such as Caenorhabditis elegans ( Miller et al, 1983 ) and in vertebrate skeletal muscles, where different developmental heavy chain isoforms are the products of separate genes in the multinucleated cells ( Lowey et al, 1991 ). When bacterially expressed LMM fragments of the adult and neonatal isoforms were denatured and renatured in vitro, they preferentially formed homodimers, indicating that the information for skeletal myosin formation probably is inherent in the amino acid sequence ( Arrizubieta and Bandman, 1999 ). In contrast, mammalian cardiac muscle contains amounts of heterodimeric V2 myosin comparable to those of the homodimeric V1 and V3 forms during certain stages of development ( Hoh et al, 1978 ; Lompre et al, 1979 ).…”

Dimerization Specificity of Adult and Neonatal Chicken Skeletal Muscle Myosin Heavy Chain Rods

“…Ni-NTA Column Chromatography. The use of Ni-NTA column chromatography to separate homodimers and heterodimers on the basis of the different affinity of zero, one, or two histidine-tagged LMM molecules for the Ni-NTA resin has been established before (29). A similar approach was taken here to study the amount of heterodimers or homodimers formed during the exchange experiments with His-tagged and FLAG-tagged or untagged MyHC rod.…”

“…Subsequently, the amount of homodimers and heterodimers was determined. The results indicated that neonatal and adult LMMs preferentially formed homodimers but ∼25% of heterodimers were also detected (29). However, in another study using chymotryptic full-length rods the amount of heterodimers formed was significantly less (9).…”

“…This suggests that the sequences responsible for the dimerization specificity are not restricted to just the LMM region of the MyHC but amino acid residues in the S2 region, consisting of the N-terminal 448 residues of the MyHC, play a role in the process as well. In the present study the dimerization specificity of a recombinantly expressed and purified full-length myosin rod produced in the same manner as LMM in our previous study (29) has been investigated. The contribution made toward the dimerization specificity by the S2 region of the MyHC has been addressed by making a chimeric protein in which the S2 region of the adult isoform was combined with the LMM region of the neonatal isoform.…”

Myosin Assembly, Maintenance and Degradation in Muscle: Role of the Chaperone UNC-45 in Myosin Thick Filament Dynamics

“…The ACD allows the tails of myosin dimers to self-assemble into antiparallel arrays that form the centre H-band of the thick filament. Filaments are rarely composed of only one form of type II myosin and often differences in the distribution of myosins within the thick filament are a result of subtle yet unidentified sequence differences in the tail domain [ 32 , 33 ]. For example, nematode body wall muscle thick filaments are composed of two different myosin heavy chains (MHC), A and B [ 34 ].…”