STRUCTURAL AND GEOMETRICAL CONSTRAINTS ON THE OUTER DYNEIN ARM IN-SITU

This study considers the relationship between two structural forms of the 22S dynein arm of Tetrahymena thermophila: the bouquet and the com pact arm. The compact arm differs from the bouquet and from other prop osed forms (e.g., the ''toadstool'') in that the globular domains are situated transversely across the interdoublet gap with one globular su bunit, the head, proximal to the adjacent doublet microtubule. The oth er models place all three globular domains proximal to the neighboring doublet microtubule. When sliding of an isolated axoneme is induced, at least 57% of total attached arms on exposed doublets are in the com pact form within dimensions of 24 X 24 X 12 nm, and only about 2% of t he arms are bouquets. Toadstools are incompatible with the images seen . Bouquets are not found in regions of the doublet protected by a neig hboring doublet. When axonemes with exposed doublets are treated with 0.5 M KCl for 30 min, the compact arms and the dynein heavy (H)-chains disappear, while isolated bouquets and dynein H-chains appear in the medium, suggesting that the compact arms give rise to the bouquets as they are solubilized. The bouquet is the predominant form of isolated 22S dynein molecules, which are found in two apparently enantiomorphic forms, within dimensions 45 X 39 X 13 nm; bouquets attached to double ts have dimensions similar to those of isolated bouquets. Computer mod eling indicates that in an intact standard-diameter axoneme, these dim ensions are incompatible with the interdoublet volume available for an arm; the bouquet therefore represents an unfolded compact arm. A plau sible sequence of changes can be modeled to illustrate the conversion of an attached compact arm to an attached and then free bouquet. The t oadstool is probably an artifact that arises after unfolding. Consiste nt with the conformational difference, H-chains of attached compact ar ms differ from those of isolated bouquets in their susceptibility to l imited proteolysis. These results suggest that the compact arm, rather than the unfolded bouquet or the toadstool, is the functional form of the outer arm in the intact axoneme. (C) 1994 Wiley-Liss, Inc.