Mechanism of Inhibition of Calcium Uptake into Sarcoplasmic Reticulum by Notexin, a Neurotoxic and Myotoxic Polypeptide

Abstract

Notexin belongs to a class of snake venom neurotoxins and myotoxins that have phospholipase A₂ activity. Previous studies have shown that these toxins affect target cells differently from phospholipases that are not neurotoxic or myotoxic. Notexin inhibited the Ca²⁺ uptake into fragmented sarcoplasmic reticulum from rabbit skeletal muscle, but it did not cause an efflux of previously accumulated Ca²⁺ or inhibit the Ca²⁺–ATPase activity. It is suggested that notexin specifically binds to and decreases the conductance for Ca²⁺ of the Ca²⁺ pump and/or the conductance of a channel for an ion that facilitates Ca²⁺ transport. The K⁺ ionophore valinomycin reversed the notexin-induced inhibition of Ca²⁺ uptake into sarcoplasmic reticulum, suggesting that the molecular target of notexin could be a K⁺ channel. Two types of reconstitution experiments make it unlikely that notexin acts by degrading a minor lipid that is resistant to hydrolysis by nontoxic phospholipases A₂. Notexininactivated sarcoplasmic reticulum vesicles were reactivated (with respect to Ca²⁺ uptake) by simple solubilization with detergent and subsequent reconstitution by detergent removal. Second, notexin was still active on sarcoplasmic reticulum vesicles after >94% of the lipids were replaced by soybean phosphoglycerides during the reconstitution procedure.

Key Words:

• Notexin
• phospholipase A₂
• sarcoplasmic reticulum
• calcium uptake