Microtubule-associated protein 1B (MAP1B), an abundant developmentally regulated neuronal protein, is a stoichiometric complex of a heavy chain and two light chains (light chain 1 and light chain 3). We find that light chain 1 is encoded within the 3' end of a previously reported MAP1B heavy chain cDNA. Amino acid sequencing, epitope mapping, Northern blotting, and Southern blotting indicate that the light chain and heavy chain are encoded by the same mRNA within the same open reading frame. In addition, amino acid sequencing of a 120 kd microtubule-binding and light chain-binding fragment of the heavy chain reveals that light chain 1 binds near the heavy chain N-terminus. Together these data indicate that the heavy chain and light chain 1 are produced by proteolytic processing of a MAP1B polyprotein and form a complex microtubule-binding domain.