Specific interaction of cobaltic complexes with myosin

Abstract

The protein myosin is rather unique as its molecules consist of both globular and rodlike parts. The former contains two ‘heads’ (HMM S-l) each of which can bind actin and split ATP apparently at the same rate [l--23 . Preparations of isolated heads have recently been found to be capable of generating a mechanochemical force by interacting with actin filaments in the presence of ATP. It was therefore concluded that neither the aggregation of myosin molecules into filaments (via the water-insoluble rods of light meromyosin, @MM)) nor the concerted action of two bound-together heads is indispensable for mechanochemical energy transduction [3-6]. These findings do not, however, solve the problem ‘why two heads’ or exclude the possibility that the two heads differ from each other in some respects and/or that cooperative effects between the two heads play a role in motile systems under physiological conditions. It has recently been demonstrated [7,8] that a myosin-ATP-Mg*+ complex is stabilized in an activated conformation before the products of ATP hydrolysis, ADP and inorganic phosphate, are released. The process of contraction which involves the actin filaments is likely to take place during this activation step. It makes sense to believe that the rodlike parts of the myosin molecule i.e., HMM subfragmentand LMM are informed of the state of the catalytic site and, vice versa, that the ‘contractile complex’ is affected by the state of the rodlike parts. The latter could differ according to whether LMM is present or absent (as in HMM or HMM S-1) and/or according to the level of aggregation of myosin. An indication for a possible effect of LMM on the catalytic activity of myosin was recently found. We have shown [9] that affinity labeling of one of myosin’s active sites with Co3+-phenanthroline-ATP, [Co~phen)-ATPJ while abolishing the ATPase activity of that site, enhances remarkably the activity of the second site; moreover, the rate of labeling of the latter is much smaller than that of the first. These effects were, however, absent upon labeling of heavy meromyosin (HMM). Since the labeling was performed at a low ionic strength, it was interesting to repeat these experiments under conditions at which myosin does not aggregate into fdaments (i.e. at a high ionic strength). The idea was to find out whether the difference in behavior between myosin and HMM originates from the very existence of LMM in the former or is associated with aggregation. In addition, we tested the ability of this cobaltic complex to label HMM S-l. In our previous study we have also found that several other cobaltic complexes could inhibit , myosin ATPase. Since cobaltihexammine very often acts as a Ca2+ analog ([ 1 O]; also Danchin, unpublished data) we have investigated the influence of this complex on the Ca’ +and the EDTA-ATPase activities of myosin.