GLYCEROL-3-PHOSPHATE TRANSPORT IN HAEMOPHILUS-INFLUENZAE - CLONING, SEQUENCING, AND TRANSCRIPTION ANALYSIS OF THE GLPT GENE

The presence of a functional glpT gene in Haemophilus influenzae could be questioned, since there is only what appears to be a truncated glp T (HI0686, 143 nt in the 5'-end) available in the H. influenzae Rd gen ome database (Fleischmann et al., 1995). For cloning of the glpT gene from H. influenzae type b strain Eagan, an isogenic glpT, rec-1 double mutant and a selective medium for detection of the glpT mutant strain s were constructed. The recombinant plasmid carrying glpT was able to complement the isogenic glpT mutant to wild-type levels of G3P uptake and permitted growth on a selective medium with G3P as a major carbon source. The nucleotide sequences of the glpT gene were determined both directly from PCR products and from the cloned DNA insert of strain E agan. An identical 1440 bp open reading frame with 480 deduced amino a cids, highly homologous to other bacterial G3P permeases, was identifi ed. A Northern blot analysis showed that the glpT genes in both Eagan and Rd strains were transcribed on a RNA of approximately 1.4 kb in si ze. Thus, it is likely that HI0686 sequence originates from a mutated glpT clone in Escherichia coli. (C) 1998 Elsevier Science B.V. All rig hts reserved.