WD-repeat proteins are found in all eukaryotes and are implicated in a variety of regulatory functions as a result of protein-protein interactions. PkwA from Thermomonospora curvata CCM3352 is a first potential example of a WD-repeat protein in a prokaryotic actinomycete. A mAb (3G2) was generated against the carboxy terminus of PkwA and was used to analyse the expression of PkwA in T. curvata. PkwA was detected in exponential growth phase following inoculation with spores, but could not be found at any stage of growth following inoculation with vegetative mycelium. PkwA and its WD domain were expressed in Escherichia coli as His-tag derivatives and purified on a Talon metal affinity matrix. The WD domain was phosphorylated by Pkg2, a membrane-spanning protein Ser/Thr kinase from 'Streptomyces granaticolor'. A membrane fraction from an exponential, spore-derived culture of T. curvata was found to phosphorylate the WD domain specifically in the presence of Mn(2+). These data confirm that PkwA is expressed in spore-derived exponential growth phase of T. curvata and could play a role as a molecular switch in a signalling pathway.

Laboratory of Physiology and Genetics of Actinomycetes Institute of Microbiology AS CR Videnska 1083 142 20 Prague 4 Czech Republic1

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WD-repeat protein encoding genes among prokaryotes of the Streptomyces genus