REGULATION OF NUCLEOSIDE DIPHOSPHATE KINASE AND AN ALTERNATIVE KINASEIN ESCHERICHIA-COLI - ROLE OF THE SSPA AND RNK GENES IN NUCLEOSIDE TRIPHOSPHATE FORMATION

We have previously reported that two genes cloned from a cosmid librar y of Escherichia coli can restore mucoidy to an algR2 mutant of Pseudo monas aeruginosa. AlgR2 is a protein involved in the regulation of nuc leoside diphosphate kinase (Ndk) as well as alginate synthesis in P. a eruginosa. One of the E. coli genes, rnk, encodes a 14.9 kDa protein w ith no homology to any other proteins. The other gene, sspA, encodes t he stringent starvation protein, a regulatory protein involved in stat ionary-phase regulation and the stringent response of E. coli. While b oth rnk and sspA restored alginate production to the P. aeroginosa alg R2 mutant, only rnk restored Ndk activity to the mutant. In this repor t, we have examined the effect of mutations in rnk and sspA on the lev els of Ndk in E. coli. We find that a mutation in rnk drastically redu ces the level of Ndk in E. coli. A mutation in sspA, however, affects the level of another nucleoside diphosphate kinase distinct from Ndk. The proteins can be easily distinguished from each other by their diff erent affinities for nucleoside diphosphates (NDPs) and also by the di fferential effect of anti-Ndk antibodies on the reactions they catalys e. The ability of either of these two proteins to restore alginate syn thesis in the algR2 mutant of P. aeruginosa demonstrates the importanc e of nucleoside triphosphate synthesis and energy metabolism for algin ate synthesis. Additionally, a role for the stringent starvation prote in (SspA) in the modulation of nucleoside triphosphate (NTP) levels in E. coli is also suggested from these experiments.