| Abstract: | Transverse tubule (TT) membrane vesicles contain a very active Mg-ATPase (EC 3.6.1.3). Concanavalin A (ConA) and other lectins were found to activate the TT Mg-ATPase from chicken skeletal muscle up to 25-fold yielding specific activities greater than 800 mumol/h/mg. The sarcoplasmic reticulum Ca-ATPase and the sarcolemma Na,K-ATPase were unaffected by ConA. 125I-Labeled lectin binding to the TT membrane Mr 102,000 glycoprotein supports the contention that this protein is identical with or is intimately associated with the TT Mg-ATPase. The ATPase exhibited non-Michaelis-Menton kinetics with both apparent negative cooperativity (n = 0.723; S0.5, Mg-ATP = 14 microM) and substrate inhibition (Ki, Mg-ATP = 10.2 mM), both of which were eliminated in the presence of ConA. Under the same conditions, ConA also abolished the unusual temperature dependence and potent Triton X-100 inhibition. The similarities in ConA suppression of both Triton and substrate inhibition suggest that these ligands may be interacting through a non-catalytic site and that Triton is serving as a nucleotide-mimetic agent. The unique kinetic responses are consistent with a homotropic substrate modifier mechanism wherein the enzyme can be viewed as possessing a single catalytic and a single regulatory site on a single polypeptide chain. It is proposed that ConA interferes either with ligand interaction at a putative regulatory site or blocks communication between a regulatory site and the catalytic site. The possible nature of the regulatory site and its modulation by a ConA-like, endogenous, skeletal muscle lectin and their combined role in excitation-contraction coupling is discussed. |
