A calcium-sensitive preparation from Physarum polycephalum

Abstract

Differential ultracentrifugation of an extract of the plasmodium of P. polycephalum yields a high-speed fraction which exhibits Ca sensitive ATP activity at low ionic strength. The rate of inorganic phosphate production increased from 2- to 25-fold in different preparations when the Ca concentration was increased from about 10-8-10-5 M. Complement fixation using specific antibody to Physarum myosin showed the fraction to contain 3% myosin. By EM actin-like microfilaments 50-150 nm long were present. Addition of pure rabbit F-actin or myosin to this fraction activated the ATPase measured in EGTA [ethylene glycol bis(.beta.-aminoethyl ether)N,N,N'',N tetraacetic acid] and so partially reversed the Ca sensitivity. If muscle myosin was added to the supernatant from which the fraction was centrifuged, a hybrid complex was obtained which included actin and additional protein from the plasmodium, and this hybrid was also Ca sensitive. Over 85% of the Ca-sensitive, Mg-activated ATPase was precipitated by sequential hybrid formation. The Ca sensitivity of the hybrid was maximal when formed at the lowest ratios of added myosin to Physarum proteins. The results do not allow a simple interpretation along the lines of either actin-linked or myosin-linked sensitivity. Evidence consistent with both a form of actin-linked and myosin-linked sensitivity is present in our results.