On the alkali light chains of vertebrate skeletal myosin. Nucleotide binding and salt-induced conformational changes.
The interaction of alkali light chains of vertebrate skeletal myosin with nucleotides and KCl has been examined by chemical modifications of these proteins, by direct binding measurements, and in circular dichroism studies. The reactivities of the single thiol groups in the isolated alkali light chains A1 and A2 have been studied by reacting these proteins with 5,5'-dithiobis(2-nitrobenzoic acid) (Nbs2). MgATP and MgADP reduced the reactivities of thiol groups while high concentrations of KCl increased them. Subsequent equilibrium dialysis experiments verified the presence of a low-affinity nucleotide binding site per each alkali subunit. Circular dichroism measurements revealed that KCl induced local (around phenylalanines) and overall (alpha-helical content) conformational changes of equal magnitude in the two alkali light chains. However, salt induced different conformational changes in the subfragment 1 isoenzymes, S-1(A1) and S-1(A2). This differential salt effect on the S-1 isoenzymes was confirmed by comparing their thermal stability in different salt conditions. AT low KCl concentrations (5 mM), S-1(A1) was found to be considerably more heat labile than S-1(A2); at higher salt levels (50 mM KCl) the stability of S-1(A1) approached that of S-1(A2). These experiments are discussed in terms of the relationship between the alkali subunits and the ATP and the actin-binding sites of myosin.
D000244:Adenosine Diphosphate / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002942:Circular Dichroism; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009132:Muscles / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D011189:Potassium Chloride / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013056:Spectrophotometry, Ultraviolet
D000244:Adenosine Diphosphate; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; D004228:Dithionitrobenzoic Acid; D009218:Myosins; D008274:Magnesium