Cross‐hybridization of light chains of cardiac myosin isozymes: Atrial and ventricular myosins

Hollósi, G; Srivastava, Sudhir; Wikman‐Coffelt, Joan

doi:10.1016/0014-5793(80)80297-3

Cited by 11 publications

(1 citation statement)

References 14 publications

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“…4) is either the result of the Ca2+-binding light chain not reassociating with the S-1 subfragment lacking the (S-1)-(S-2) junction or partial reassociation occurred but the Ca2+-binding properties were no longer influenced by the heavy chains. Native myosin, conversely reassociated under these conditions Higuchi et al, 1978;Wikman-Coffelt, 1980;Hollosi et al, 1980) and the high Ca2+-binding affinity value was restored. The Ca2+-binding light chain has a different affinity for Ca2+ when complexed to the heavy chains as compared with when it is in the dissociated state (Wikman-Coffelt et al, 1979) and only binds at Ca2+ concentrations required to induce a conformational change in free light chains (Ca2+ 10-510-3 M) (Morakovcic et al, 1979;Yamamoto etal., 1980).…”

Section: Resultsmentioning

confidence: 94%

Influence of myosin heavy chains on the Ca2+-binding properties of light chain, LC2

Srivastava

Mühlrád

Wikman‐Coffelt

1981

Biochemical Journal

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The association of myosin light chains with heavy chains, i.e. the intact oligomeric structure, profoundly affects the Ca(2+)-binding properties of the light chains. The Ca(2+)-binding affinity of the light chains is more than two magnitudes higher in the presence of heavy chains than in its absence. Modification of the reactive SH(2) thiol of myosin results in an alteration in the conformation of heavy chains of the molecule that influences the Ca(2+)-binding properties of light chains and generation of tension. When the SH(2) moiety is blocked with N-ethylmaleimide the influence of the heavy chains on the Ca(2+)-binding properties of light chain LC(2) is lost; under these conditions the Ca(2+)-binding affinity value of SH(2)-N-ethylmaleimide-blocked myosin (3.3x10(4)m(-1)) decreases to near that expressed with the dissociated light chain LC(2) (0.7x10(4)m(-1)). Conversely, the presence of actin, nucleotides or modification of either the reactive lysyl residue or SH(2) thiol does not affect Ca(2+) binding. The native secondary and tertiary structure of myosin seem to be required for Ca(2+) binding; binding does not occur in the presence of 6m-urea with either native myosin or the dissociated light chains. With SH(2)-N-ethylmaleimide-blocked myosin normal Ca(2+)- and (Mg(2+)+actin)-stimulated ATPase activities are expressed; however, there is a loss in K(+)-stimulated ATPase activity and the synthetic actomyosin threads of such myosin express no isometric tension. There are also variances in the binding of Ca(2+) with alterations in pH values. In the absence of Ca(2+)/EGTA buffer the biphasic Ca(2+)-binding affinity of myosin is twice as high at pH7.4 (site one: 1.2x10(6)m(-1) and site two: 0.4x10(6)m(-1)) as compared with values obtained at pH6.5 (site one: 0.64x10(6)m(-1) and site two: 0.2x10(6)m(-1)). The Ca(2+)-binding affinity of light chain LC(2) and S(1), where the (S-1)-(S-2) junction was absent, were not influenced by changes in pH values. Both expressed a low Ca(2+)-binding affinity, approx. 0.7x10(4)m(-1), whereas heavy meromyosin, where both (S-1) and (S-2) myosin subfragments were present, expressed a Ca(2+)-binding affinity value similar to that of native myosin, but was not biphasic. However, it is important to point out than in preparation of S(1) myosin subfragment light chain LC(2) was lost and thus was added back to the purified S(1) fraction. Light chain LC(2) was not, however, added to the heavy meromyosin fraction because it was not lost during preparation of the heavy meromyosin subfragment. In conclusion, it appears that the (S-1)-(S-2) junction is needed for the positioning of light chain LC(2) and thus influences its essential conformation for Ca(2+) binding.

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Section: Resultsmentioning

confidence: 94%

Influence of myosin heavy chains on the Ca2+-binding properties of light chain, LC2

Srivastava

Mühlrád

Wikman‐Coffelt

1981

Biochemical Journal

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Functional effects of LC1-reassociation with cardiac papain Mg � S1

Margossian

White

Lefford

et al. 1993

J Muscle Res Cell Motil

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The effect of LC1 on cardiac myosin structure and activity was investigated using as a model S1 prepared by papain digestion in the presence of Mg2+. The resulting S1 contained LC2 but a part of the N-terminal region of LC1 was cleaved. Sequencing the N-terminal part of the band migrating below LC1 on SDS gels revealed it to consist of alternating alanyl and prolyl residues thus establishing LC1 as the origin of this band. However, Western blots did not reveal any LC1 while radioimmunoassays indicated it to be present at the 5% level suggesting the anti-LC1 antibody used in these experiments did not recognize the C-terminal portion of LC1 still attached to Mg.S1. Mixing a 10-15 M excess of isolated light chains with Mg.S1 in the presence of 10 mM ATP, 12 mM MgCl2, 4.7 M NH4Cl allowed LC1 to recombine with LC1-deficient Mg.S1. Equilibrium ultracentrifugation analysis revealed a highly heterogeneous LC1-deficient S1 which upon recombination with intact LC1 became monodisperse as indicated by the superimposition of molecular weight averages all across the centrifuge cell. LC1-deficient Mg.S1 had a Vm of 0.4 s-1, Ka of 30 microM and a Kbind of 28 microM. In the presence of intact LC1, Vm rose to 0.8 s-1 while Ka and Kbind were reduced to 7.5 and 12 microM, respectively. The fourfold decrease in Ka strongly indicated an increased affinity for actin by Mg.S1 in the presence of uncleaved LC1. Also, Ca(2+)-regulation of dog heart myofibrils was suppressed when Ca(2+)-activated MgATPase assays, as a function of Ca2+, were performed in the presence of anti-LC1 antibodies. These observations suggest the presence of intact, uncleaved LC1 in S1 is required for the stability of S1 heavy chains and proper Ca(2+)-regulation.

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Myosin light chain compositions of the interatrial and interventricular septa of sheep heart

1985

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The myosin light chain composition of sheep interatrial and interventricular septa were analysed by one- and two-dimensional polyacrylamide gel. The interventricular septum has myosin light chain composition indistinguishable from that of ventricular myosin. Myosin from the interatrial septum contains three light chains, two of which co-migrated with the two atrial light chains (ALC1 and ALC2), while the third co-migrated with ventricular light chain 2 (VLC2). ALC1 are more abundant than ALC2 or VLC2 suggesting a mixed myosin population. Myosin with ALC1 and VLC2 light chain composition may be present, and its possible relationship with cardiac "conducting" cells is discussed.

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Cross‐hybridization of light chains of cardiac myosin isozymes: Atrial and ventricular myosins

Cited by 11 publications

References 14 publications

Influence of myosin heavy chains on the Ca2+-binding properties of light chain, LC2

Influence of myosin heavy chains on the Ca2+-binding properties of light chain, LC2

Functional effects of LC1-reassociation with cardiac papain Mg � S1

Myosin light chain compositions of the interatrial and interventricular septa of sheep heart

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