Evolution of the bacterial phosphotransferase system: from carriers and enzymes to group translocators.

The bacterial phosphotransferase system (PTS) is a structurally and functionally complex system with a surprising evolutionary history. The substrate-recognizing protein constituents of the PTS (Enzymes II) derive from at least four independent sources. Some of the non-PTS precursor constituents have been identified, and evolutionary pathways taken have been proposed. Our analyses suggest that two of these independently evolving systems are still in transition, not yet having acquired the full-fledged characteristics of PTS Enzyme II complexes. The work described provides detailed insight into the process of catalytic protein evolution.

Sequence and phylogenetic analyses of 4 TMS junctional proteins of animals: connexins, innexins, claudins and occludins.

Connexins and probably innexins are the principal constituents of gap junctions, while claudins and occludins are principal tight junctional constituents. All have similar topologies with four alpha-helical transmembrane segments (TMSs), and all exhibit well-conserved extracytoplasmic cysteines that either are known to or potentially can form disulfide bridges. We have conducted sequence, topological and phylogenetic analyses of the proteins that comprise the connexin, innexin, claudin and occludin families. A multiple alignment of the sequences of each family was used to derive average hydropathy and similarity plots as well as phylogenetic trees. Analyses of the data generated led to the following evolutionary and functional suggestions: (1) In all four families, the most conserved regions of the proteins from each family are the four TMSs although the extracytoplasmic loops between TMSs 1 and 2, and TMSs 3 and 4 are usually well conserved. (2) The phylogenetic trees revealed sets of orthologues except for the innexins where phylogeny primarily reflects organismal source, probably due to a lack of relevant organismal sequence data. (3) The two halves of the connexins exhibit similarities suggesting that they were derived from a common origin by an internal gene duplication event. (4) Conserved cysteyl residues in the connexins and innexins may point to a similar extracellular structure involved in the docking of hemichannels to create intercellular communication channels. (5) We suggest a similar role in homomeric interactions for conserved extracellular residues in the claudins and occludins. The lack of sequence or motif similarity between the four different families indicates that, if they did evolve from a common ancestral gene, they have diverged considerably to fulfill separate, novel functions. We suggest that internal duplication was a general evolutionary strategy used to generate new families of channels and junctions with unique functions. These findings and suggestions should serve as guides for future studies concerning the structures, functions and evolutionary origins of junctional proteins.

Influence of threonine exporters on threonine production in Escherichia coli.

Threonine production in Escherichia coli threonine producer strains is enhanced by overexpression of the E. coli rhtB and rhtC genes or by heterologous overexpression of the gene encoding the Corynebacterium glutamicum threonine excretion carrier, thrE. Both E. coli genes give rise to a threonine-resistant phenotype when overexpressed, and they decrease the accumulation of radioactive metabolites derived from [(14)C] L-threonine. The evidence presented supports the conclusion that both RhtB and RhtC catalyze efflux of L-threonine and other structurally related neutral amino acids, but that the specificities of these two carriers differ substantially.

Phylogenetic and structural analyses of the oxa1 family of protein translocases.

Mitochondrial Oxa1p homologs have been shown to function in protein export and membrane insertion in bacteria, mitochondria and chloroplasts, but their mode of action, organismal distribution and evolutionary origins are poorly understood. All sequenced homologs of Oxa1p were retrieved from the databases and multiply aligned. All organisms with a fully sequenced genome possess at least one Oxa1p homolog showing that the family is truly ubiquitous. Most prokaryotes possess just one Oxa1p homolog, but several Gram-positive bacteria and one archaeon possess two, and eukaryotes may have as many as six. Although these proteins vary in length over a 5-fold range, they exhibit a common hydrophobic core region of about 200 residues. Multiple sequence alignments reveal conserved residues and provide the basis for structural and phylogenetic analyses that serve to characterize the Oxa1 family.