Antibody decoration of neurofilaments

Abstract

We have decorated neurofilaments with antibodies against three polypeptides (designated here as H [mol wt=195,000], 45[mol wt = 145,000], and 46 [mol wt=73,000]) in an effort to understand the arrangement of these poypeptides within neurofilaments. The three polypeptides were purified and antibodies were raised against each. The cross-reactivity of the antibodies suggested that each polypeptide contains both shared and unique antigenic determinants. The differential reactivities of each antibody preparation were enhanced by adsorption with the two heterologous polypeptides, and the resulting preparations were used to decorate purified neurofilaments, which were then negatively stained and examined in an electron microscope. The appearance of the antibody-decorated structures led to the following conclusions: All three polypeptides are physically associated with the same neurofilament. However, the disposition of H and 46 within a filament is different; 46 antigens appear to be associated with a 'central core' of the filament, whereas H antigens compose a structure more loosely and peripherally attached to the central core and periodically arranged along its axis. The antibody-decorated H-containing structure assumes variable configurations; in some cases it appears as a bridge connecting two filaments; in other cases it appears as a helix wrapping the central core with a period of ~1,000 Å and a apparent unit length of ~1.5 periods. These configurations suggest several functional implications, including the possibility that H is a component of the cross-bridges observed between filaments in situ. We also note that the central core-helix relationship could be used in the design of an intracellular transport motor.