Methylobacterium extorquens RSH Enzyme Synthesizes (p)ppGpp and pppApp in vitro and in vivo, and Leads to Discovery of pppApp Synthesis in Escherichia coli

This study of the RSHMex enzyme activity is the first direct biochemical demonstration that an RSH enzyme is capable of synthesizing a nucleotide derivative other than (p)ppGpp, i.e., pppApp. This is shown in vitro with the RSHMex1-352 protein (Figures 3, 4), as well as in vivo with E. coli cells induced to express full-length RSHMex and with wild type M. extorquens AM1 cells (Figure 6). Moreover, we demonstrate pppApp accumulation in wild type E. coli cells, which points to a possible new player in bacterial stringent response. In this work, it became evident to us that there may be a need to re-evaluate RSH nomenclature, changing it from the traditional standard of Rel(Species-name) to RSH(Species-name) and therefore in this case we use RSHMex. We suspect the problem arose when the Cashel-Mechold lab began studies on what they then thought was a relA-like monofunctional protein from a S. equisimilis strain. This led to calling it RelSeq as the RelA protein from S. equisimilis (Mechold et al., 1996). It was later discovered to be a bifunctional RSH protein but the name was not changed. The E. coli RelA is a monofunctional enzyme capable only of (p)ppGpp synthesis, but not its hydrolysis. We trust that the RSHMex full-length enzyme is bifunctional and capable of both, (p)ppNpp synthesis and hydrolysis (see below). Also, as already mentioned in the section “Results,” this enzyme has a higher homology to SpoT and RelSeq (both are bifunctional enzymes) than RelA. ...
Source: Frontiers in Microbiology - Category: Microbiology Source Type: research