Significance of the N-terminal fragment of myosin regulatory light chain for myosin-actin interaction.
The influence of myosin regulatory light chains (LC2s) lacking the 2kD N-terminal portions of these chains, on internal organization of myosin heads and on actin-myosin interaction was studied with limited proteolysis and polarized fluorescence methods. For these studies heavy meromyosin (HMM) preparations were used: HMM containing intact LC2s (phosphorylated or dephosphorylated) and HMM containing LC2s lacking the 2kD N-terminal portions (including serine which can be phosphorylated). It was found that the susceptibility of the heavy chain cleavage site to trypsin and the alkali light chain (LC1) site to papain of myosin containing shortened regulatory LC2s, is not dependent on saturation of LC2s with Ca2+ or Mg2+ ions. This is in contrast to the myosin containing intact LC2s where Ca2+ or Mg2+ ion saturation does demonstrate a dependence. Similarly, in spectroscopic experiments, dephosphorylated HMM containing intact LC2s causes decrease or increase of actin filament flexibility depending on whether Mg2+ or Ca2+ are bound to LC2s. Correspondingly, HMM with shortened LC2s induces only increase of actin flexibility despite cations being bound. We conclude that the N-terminal fragment of LC2 is important for ensuring a proper Ca2+ dependent conformation of myosin head in the course of its actin-activated ATP hydrolysis.
Stepkowski
D
D
Department of Cellular Biochemistry, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Warszawa.
Szczesna
D
D
Babiychuk
E B
EB
Borovikov
Y S
YS
Kakol
I
I
D000199:Actins / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D005454:Fluorescence Polarization; D008274:Magnesium / Q000378:metabolism; D018482:Muscle, Skeletal / Q000737:chemistry; D015879:Myosin Subfragments / Q000378:metabolism; D009218:Myosins / Q000737:chemistry* / Q000378:metabolism*; D010206:Papain / Q000378:metabolism; D010766:Phosphorylation; D011817:Rabbits; D012045:Regulatory Sequences, Nucleic Acid; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000378:metabolism
D000199:Actins; D015879:Myosin Subfragments; D000255:Adenosine Triphosphate; D014357:Trypsin; D010206:Papain; D009218:Myosins; D008274:Magnesium; D002118:Calcium