Isolation of a Ca2+-protease resistant high Mr microtubule binding protein from mammalian brain: Characterization of properties partially expected for a dynein-like molecule

Abstract

A novel microtubule binding protein was isolated from cell extracts of pig brain after selective destruction of high Mr MAPs by Ca2+-activation of endogenous proteases. The protein had an apparent Mr of 300,000, but several criteria, including peptide maps, immunological crossreactivities, resistance to Ca2+-activated proteolysis, and inability to induce microtubule assembly, distinguished this protein from the major high Mr microtubule associated proteins MAP-1 and MAP-2. Certain molecular properties of the protein resembled those of dynein: its size under denaturing conditions, its crossreactivity with antibodies toTetrahymena dynein, the ATP-dependence of its microtubule binding, the erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA)—sensitivity of its ATPase activity and its resistance to Ca2+-activated proteolysis. However, in peptide maps and the insensitivity of its ATPase activity to vanadate the protein could be differentiated fromTetrahymena dynein heavy chain polypeptides. Based on the properties characterized so far, the protein seems qualified to function as a mechanochemical enzyme in the cytoplasm of mammalian brain cells and could represent a cytoplasmic dynein variant.