SEQUENCING OF HEAT-SHOCK-PROTEIN-70 (DNAK) HOMOLOGS FROM DEINOCOCCUS-PROTEOLYTICUS AND THERMOMICROBIUM-ROSEUM AND THEIR INTEGRATION IN A PROTEIN-BASED PHYLOGENY OF PROKARYOTES

The 70-kDa heat shock protein (hsp70) sequences define one of the most conserved proteins known to date. The hsp70 genes from Deinococcus pr oteolyticus and Thermomicrobium roseum, which were chosen as represent atives of two of the most deeply branching divisions in the 16S rRNA t rees, were cloned and sequenced. hsp70 from both these species as well as Thermus aquaticus contained a large insert in the N-terminal quadr ant, which has been observed before as a unique characteristic of gram -negative eubacteria and eukaryotes and is not found in any gram-posit ive bacteria or archaebacteria. Phylogenetic analysis of hsp70 sequenc es shows that all of the gram-negative eubacterial species examined to date (which includes members from the genera Deinococcus and Thermus, green nonsulfur bacteria, cyanobacteria, chlamydiae, spirochetes, and alpha-, beta-, and gamma-subdivisions of proteobacteria) form a monop hyletic group (excluding eukaryotic homologs which are derived from th is group via endosybitic means) strongly supported by the bootstrap sc ores. A closer affinity of the Deinococcus and Thermus species to the cyanobacteria than to the other available gram-negative sequences is a lso observed in the present work. In the hsp70 trees, D. proteolyticus and T. aquaticus were found to be the most deeply branching species w ithin the gram-negative eubacteria. The hsp70 homologs from gram-posit ive bacteria branched separately from gram-negative bacteria and exhib ited a closer relationship to and shared sequence signatures with the archaebacteria. A polyphyletic branching of archaebacteria within gram -positive bacteria is strongly favored by different phylogenetic metho ds. These observations differ from the rRNA-based phylogenies where bo th gram-negative and gram-positive species are indicated to be polyphy letic. While it remains unclear whether parts of the genome may have v ariant evolutionary histories, these results call into question the ge neral validity of the currently favored three-domain dogma.