Aldoheptose Biosynthesis. Gram-negative bacteria (pathogens and nonpathogens) have a unique outer membrane and are more refractory to therapeutic attack than their gram-positive counterparts, save mycobacteria. Lipopolysaccharide (LPS) forms all of the lipid of the inner core aldoheptose (i.e., L-glycero-D-mannoheptose) are hypersensitive to a number of antibiotics. The molecular genetics and biology of the E. coli K-12 rfaD gene have been reported by this laboratory. The purified rfaD gene product has been characterized both kinetically and physically. Crystals of rfaD gene product (ADP-L- glycero-D-mannoheptose 6-epimerase) have been obtained. The ubiquity of the rafD gene among enteric and nonenteric gram-negatives is under study. For example, we have demonstrated structural and functional homology of the rafD gene of nonenteric pathogen Pseudomonas aeruginosa. The molecular genetics and biology of a second mutation, recently designated rfaC (formerly rfa-2), clearly defines the rfaC gene and its product (heptosyl transferase 1). The opening frames for both the rfaF and rfaL genes and their chromosomal relationship to rfaC and rfaD genes have been determined. An affinity chromatography based purification protocol for phosphoheptose isomerase has been developed. New mini-Tn10dCAM insertion rfa novobiocin hypersensitive or barrier mutants have been isolated and are now subjects of investigation.
