The long term objectives for this project are to understand how living cells regulate, synthesize and assemble biological membranes. This is of particular significance when considering all cells are separated for their external environment by intact membranes and that, within cells, compartmentalization of specific functions, e.g. respiratory, secretion, nuclear replication, synthesis, digestion, etc., requires unique membrane systems. Likewise, internalization of materials from outside to the inside of cells, receptors, cell-cell interactions, viral attachment, all involve membrane structure and function. Numerous human diseases involve malformations in structure and function of a variety of these unique cellular membrane systems. Thus, we have chosen for study a unique experimental system where the presence and amount of an intact, functional membrane system can be easily regulated and is gratuitous for cell function and growth. This membrane system is found within the facultative photoheterotrophic bacterium, Rhodobacter sphaeroides. This inducible membrane system has unique electron transport capabilities which are readily studied, as well as specialized spectral properties which make it relatively easy to follow the intact membrane or membranous subfractions through purification and isolation procedures. Three specific subfractions or complexes, comprising 60% of the membrane protein, are readily isolated and have been the subject of intense biophysical, crystallographic, and protein studies. Through a combination of physiological, molecular biological, genetic and DNA cloning procedures it has become possible to address a series of specific questions which are intended to investigate the synthesis, regulation and assembly of each of these three integral membrane complexes. Recent studies have demonstrated the existence of transcriptional, post-transcriptional and translational and post- translational control mechanisms required to provide for the orderly synthesis of this membrane system as well as cellular adaptation from one growth condition to another. The existence of unique regulatory mechanisms involving codon usage, protein turnover and accessory assembly factors make this experimental system unique, as does the demonstration of overlapping transcriptional units. The use of site-system directed mutagenesis, measurements of mRNA expression, abundance and stability; the use of reporter groups; and, measurements of amounts of localization of gene protein products will all be employed in order to investigate membrane synthesis and assembly.
NIH ExPorter
NIH ExPorter