Identification of a Microtubule-binding Domain in a Cytoplasmic Dynein Heavy Chain

• Published in: The Journal of biological chemistry 1997-08, Vol.272 (32), p.19714-19718
• Main Author: Koonce, Michael P.
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - metabolism; Animals; BINDING SITES; Cytoplasm - enzymology; DELETIONS; DICTYOSTELIUM; DICTYOSTELIUM DISCOIDEUM; DYNEIN ATPASE; Dyneins - chemistry; Dyneins - genetics; Dyneins - metabolism; GENE; GENES; HIDROLASAS; HYDROLASE; HYDROLASES; Hydrolysis; MICROTUBULE; MICROTUBULES; Microtubules - metabolism; MICROTUBULOS; MOLECULAR CONFORMATION; Molecular Sequence Data; Open Reading Frames; PARTIAL DELETIONS; PEPTIDE; PEPTIDES; PEPTIDOS; POLYPEPTIDES; Protein Binding; Protein Biosynthesis; Protein Conformation; Protein Folding; PYROPHOSPHATASES; SECONDARY STRUCTURE; Sequence Deletion; Transcription, Genetic; Ultraviolet Rays; Vanadates - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.272.32.19714

As a molecular motor, dynein must coordinate ATP hydrolysis with conformational changes that lead to processive interactions with a microtubule and generate force. To understand how these processes occur, we have begun to map functional domains of a dynein heavy chain from Dictyostelium. The carboxyl-terminal 10-kilobase region of the heavy chain encodes a 380-kDa polypeptide that approximates the globular head domain. Attempts to further truncate this region fail to produce polypeptides that either bind microtubules or UV-vanadate cleave, indicating that the entire 10-kilobase fragment is necessary to produce a properly folded functional dynein head. We have further identified a region just downstream from the fourth P-loop that appears to constitute at least part of the microtubule-binding domain (amino acids 3182–3818). When deleted, the resulting head domain polypeptide no longer binds microtubules; when the excised region is expressed in vitro, it cosediments with added tubulin polymer. This microtubule-binding domain falls within an area of the molecule predicted to form extended α-helices. At least four discrete sites appear to coordinate activities required to bind the tubulin polymer, indicating that the interaction of dynein with microtubules is complex.