Wednesday, August 1, 2007 - 8:05 AM
S125
A third alternative for protein expression: the methanogenic Archaea
Kevin R. Sowers and Sheridan R. MacAuley. Center of Marine Biotechnology, University of Maryland Biotechnology Institute, 701 E. Pratt St., Baltimore, MD 21202
The Archaea is a phylogenetic lineage of microorganisms that is genotypically distinct from both bacteria and eukaryotes. These microorganisms are morphologically similar to monocellular, non-nucleated bacteria and possess many biosynthesis pathways typically found in bacteria. In contrast nucleic acid processing including both transcription and translation generally occurs by mechanisms similar to those in eukaryotes. Several characteristics make the methanogenic Methanosarcina particularly uniquely useful for expression of recombinant bacterial and eukaryotic proteins including mesophilic growth, a neutral intercellular milieu, the presence of all four chaperone systems for protein folding and the availability of methods for mass culturing. Several characteristics of these microorganisms have largely precluded attempts to apply bacterial or eukaryotic genetic protocols to members of this phylum including unique cell wall structures that prevent the use of commonly used antibiotic genetic markers targeting cell wall synthesis; bacterial gene promoters associated with many of the commonly used genetic markers that are not recognized by the archaeal transcriptional apparatus; bacterial plasmids and phages that will not replicate in archaeal species. In recent years several laboratories have overcome these difficulties by developing effective plating techniques, identifying genetic markers that do not target cell wall synthesis, fusing archaeal promoters with recombinant genes and isolating native archaeal vectors and identifying promiscuous non-archaeal vectors. Gene transfer systems now exist for methanogenic species and despite varying degrees of difficulty applying these protocols they are routinely used by laboratories conducting research on archaeal genetics and can be mastered by anyone with a fundamental knowledge of microbial genetic techniques.
Web Page: www.umbi.umd.edu/~sowers/