Recently we have identified mRNA encoding a hitherto unknown mammalian X,K-ATPase beta -subunit expressed predominantly in muscle tissue (Pestov, N. B. ef al. (1999) FEES Lett. 456, 243-248), Here we demonstrate the existence of the predicted protein, designated as beta (m) (beta (muscle)), in human adult skeletal muscle membranes using immunoblotting with beta (m)-specific antibodies generated against recombinant polypeptide formed by extramembrane beta (m) domains. The electrophoretic mobility of beta (m) was shown to be abnormally low due to the presence of Glu-rich sequences, In contrast to mature forms of other known X,K-ATPase beta -subunits, carbohydrate moiety of beta (m) is sensitive to endoglycosidase H and appears to be composed of short high-mannose or hybrid N-glycans, This finding argues in favor of an intracellular location of beta (m) in human skeletal muscle.