Proteins with CHADs (Conserved Histidine α-Helical Domains) Are Attached to Polyphosphate GranulesIn Vivoand Constitute a Novel Family of Polyphosphate-Associated Proteins (Phosins)
Authors: Tumlirsch Tony , Jendrossek Dieter 
Published: January 2017

On the basis of bioinformatic evidence, we suspected that proteins with a CYTH (CyaBthiamine triphosphatase) domain and/or a CHAD (conservedhistidineα-helicaldomain) motif might represent polyphosphate (polyP) granule-associated proteins. We found no evidence of polyP targeting by proteins with CYTH domains. In contrast, two CHAD motif-containing proteins fromRalstonia eutrophaH16 (A0104 and B1017) that were expressed as fusions with enhanced yellow fluorescent protein (eYFP) colocalized with polyP granules. While the expression of B1017 was not detectable, the A0104 protein was specifically identified in an isolated polyP granule fraction by proteome analysis. Moreover, eYFP fusions with the CHAD motif-containing proteins MGMSRV2-1987 fromMagnetospirillum gryphiswaldenseand PP2307 fromPseudomonas putidaalso colocalized with polyP granules in a transspecies-specific manner. These data indicated that CHAD-containing proteins are generally attached to polyP granules. Together with the findings from four previously polyP-attached proteins (polyP kinases), the results of this study raised the number of polyP-associated proteins inR. eutrophato six. We suggest designating polyP granule-bound proteins with CHAD motifs as phosins (phosphate), analogous tophasins andoleosins that are specifically bound to the surface of polyhydroxyalkanoate (PHA) granules in PHA-accumulating bacteria and to oil droplets in oil seed plants, respectively.

IMPORTANCEThe importance of polyphosphate (polyP) for life is evident from the ubiquitous presence of polyP in all species on earth. In unicellular eukaryotic microorganisms, polyP is located in specific membrane-enclosed organelles, called acidocalcisomes. However, in most prokaryotes, polyP is present as insoluble granules that have been designated previously as volutin granules. Almost nothing is known regarding the macromolecular composition of polyP granules. Particularly, the absence or presence of cellular compounds on the surface of polyP granules has not yet been investigated. In this study, we identified a novel class of proteins that are attached to the surface of polyP granules in three model species ofAlphaproteobacteria,Betaproteobacteria, andGammaproteobacteria. These proteins are characterized by the presence of a CHAD (conservedhistidineα-helicaldomain) motif that functions as a polyP granule-targeting signal. We suggest designating CHAD motif-containing proteins as phosins [analogous to phasins for poly(3-hydroxybutyrate)-associated proteins and to oleosins for oil droplet-associated proteins in oil seed plants]. The expression of phosins in different species confirmed their polyP-targeting function in a transspecies-specific manner. We postulate that polyP granules in prokaryotic species generally have a complex surface structure that consists of one to several polyP kinases and phosin proteins. We suggest differentiating polyP granules from acidocalcisomes by designating them as polyphosphatosomes.