Membrane morphogenesis is a universal aspect of biological cell division, intracellular organellogenesis and mult cellular development. We wish to demonstrate how specific proteins cause shape changes in biological membranes. Bacteriophage PM2 is a useful model because of its simple lipid bilayer membrane. Proteins coded by bacteriophage PM2 cause a bud to form in the membrane of its host, Alteromonas espejiana. We hope to describe the molecular aspects of this lipid-protein interaction by isolation of the gene(s) responsible for synthesis of a morphogenic protein(s). We have demonstrated coupled transcription-translation directed by restriction endonuclease fragments of PM2 DNA. One of these fragments codes for the synthesis of sp6.6, a viral structural protein implicated in membrane morphogenesis. This sp6.6 restriction fragment and others will be inserted into plasmid pBR322 for amplification in E. coli. In transformed cells, we hope to see extreme membrane morphogenesis caused by overproduction of sp6.6. We will also determine the amino acid sequence of sp6.6 by sequencing its DNA restriction fragment. Stretches of hydrophobic amino acids will suggest a model for lipid-protein interaction. The model will be tested for the effect on membrane morphogenesis of site-specific mutagenesis of the sp6.6 gene. These fragments will also be subjected to in vitro protein synthesis in the presence of membranes to determine whether the membrane morphogenic protein is inserted co-translationally or post-translationally. Thus, we expect to be able to describe the genetic, biochemical and physical principles of membrane morphogenesis.
| Armour, G A; Brewer, G J (1990) Membrane morphogenesis from cloned fragments of bacteriophage PM2 DNA that contain the sp6.6 gene. FASEB J 4:1488-93 |
