Figure 1.

a, Initial characterization of DAG-125. In vitro translated dystroglycan fragments were used to probe the indicated membrane fractions from Torpedo electric organ. The subscripts indicate the amino acid numbering for the human sequence (see b for the domain structure of the dystroglycans and the position of these fragments). Lanes 1 and 2, A polydisperse membrane protein (DAG-125) binds to an extracellular portion of dystroglycan. The extracellular domain of dystroglycan (lane 1, DG_1-750) bound to DAG-125, whereas the intracellular portion of dystroglycan (lane 2, DG_776-891) did not. The region of negativity in the center of the polydisperse DAG-125 band is due to a high abundance, nonbinding protein that does not copurify with DAG-125 (see lanes 5–7). Lanes 3 and 4, DAG-125 is enriched in synaptic as compared with nonsynaptic membranes. Equivalent amounts of protein from each membrane fraction were loaded in both lanes. Lanes 5–7, DAG-125 can be extracted from the membrane by alkaline treatment. Synaptic membranes were extracted at pH 12 and the insoluble (lane 6) and soluble (lane 7) fractions were analyzed. Greater than 90% of DAG-125 is solubilized by pH 12.0 treatment. b, Mapping of the DAG-125–binding region of dystroglycan. A schematic diagram of the in vitro translated recombinant dystroglycan fragments used to probe DAG-125 by blot overlay is shown. The COOH-terminal one-third of α-dystroglycan binds DAG-125. A small contribution from the middle third of α-dystroglycan is also possible. β-Dystroglycan does not appear to contribute to binding of DAG-125.