Chymotryptic digestion of Tetrahymena 22S dynein. I. Decomposition of three-headed 22S dynein to one- and two-headed particles

Abstract

Molecular composition of Tetrahymena ciliary dynein has been examined by electron microscopy and gel electrophoresis. SDS-urea gel electrophoresis revealed that Tetrahymena 22S dynein contains three (Aα, Aβ, and Aγ) heavy chains whereas 14S dynein contains only one. The molecular masses of 22S and 14S dynein heavy chains were estimated to be ~490 and 460 kD, respectively. Electron microscopy of negatively stained specimens showed 22S dynein has three globular heads and thin stalks, whereas 14S dynein consists of a single head. Chymotrypsin digested each of the three 22S dynein heavy chains into large fragments with different time courses. Sucrose density gradient centrifugation separated the digestion products as two peaks. The one with a larger sedimentation coefficient mainly consisted of two-headed particles having binding ability to doublet microtubules, whereas the other with a smaller sedimentation coefficient consisted of only isolated globular particles. Both fractions had ATPase activities. Thus, one active head of 22S dynein can be isolated by chymotrypsin digestion.