Comparative electron microscopy and image analysis of oxy- and deoxy-hemocyanin from the spiny lobster Panulirus interruptus

Structural differences between oxy-hemocyanin and deoxy-hemocyanin from the spiny lobster P. interruptus were studied by electron microscopy and image analysis of negatively stained preparations. Projections of the hexameric P. interruptus hemocyanin from electron microscopy were compared with simulated projections of the high-resolution structure determined by X-ray crystallography previously. Image analysis was focused on the hexagonal views. Four independent data sets of hexagons (two of oxy- and two of deoxy-preparations) were analyzed and compared. Despite the small size of the projections, a resolution in the average images of better than 1.5 nm was obtained. A significant difference in stain modulation could be noticed between the oxy- and deoxy-preparations in two areas of the projections. The first difference in stain exclusion occurred in the center of the hexagonal views. The second is the shape of the domain 3 region. In the deoxy-images this region has a triangular shape, in the oxy-images it appears to be more rounded. The overall differences suggest a "breathing" of the hexamer upon oxygenation/deoxygenation.