THE SUPRAMOLECULAR ARCHITECTURE OF THE POLYHYDROXYALKANOATE INCLUSIONS IN PSEUDOMONAS-OLEOVORANS

A study has been initiated to further clarify the role of H2O and protein in maintaining a functional amorphous state of poly-beta-hydroxyalkanoates (PHAs) in the living cell. It has been clearly demonstrated that the ordered hydrophobic hydrocarbon chains of these materials crystallize at varying rates, depending on repeating unit heterogeneity, subsequent to the removal of the polymer granule from the cell and after solvent extraction. Both isolation and extraction procedures remove water and some protein from the polymer granule, and the polymer crystallizes. We have developed a procedure for the isolation of poly-beta-hydroxyoctanoate (PHO) containing granules that maintain their 'in vivo' fluid, amorphous and functional state with water and proteins intact. These purified granules have been observed and their purity determined by 'in vivo' Nomarski computer-enhanced imaging light microscopy. Both this technique and low temperature rapid freeze-fracture electron microscopy have demonstrated an organized para-crystalline network of proteins on the surface of the granule. The structural role of these proteins in maintaining granule integrity and function both in the cell and after granule isolation is discussed. It is suggested that these procedures may permit in vitro studies of the mechanisms of synthesis and degradation of PHAs in a variety of polyester producing microbial systems.