PARTIAL CLONING OF A SQUID MICROTUBULE-ASSOCIATED PROTEIN (MAP-H1) AND THE IDENTIFICATION OF THE MICROTUBULE BINDING DOMAIN

An approximately 420-kDa ATP binding protein, referred to as MAP H1, has previously been shown to be involved in microtubule-dependent vesicle motility in the squid giant axon. To gain further insight into the structure and function of this protein, partially overlapping cDNA clones encoding approximately a quarter of the MAP Hl molecule were identified from two squid optic lobe libraries using affinity-purified antibodies to squid MAP H 1. One clone in particular (KS18), which hybridizes to an approximately 13-kb message, encodes a series of almost identical repeats of a 16-amino-acid sequence that is tandemly repeated. The sequence of clone KS18 is unique and does not correspond to any nucleotide or amino acid sequence in the data base. The presence of repeated elements within the microtubule binding domain of several other MAPs prompted us to investigate whether the MAP HI repeats are involved in microtubule binding. In-vitro-synthesized polypeptides containing these repeats sediment with taxol-stabilized microtubules in a microtubule binding assay. The predicted secondary structure of the 16-amino-acid repeat region of MAP H1 contains alternating beta-sheets and turns and could form the globular domain seen in negative-stain electron micrographs of MAP Hl.