An alternative PII protein in the regulation of glutamine synthetase in Escherichia coli

The P_II protein has been considered pivotal to the dual cascade regulating ammonia assimilation through glutamine synthetase activity. Here we show that P_II, encoded by the glnB gene, is not always essential; for instance upon ammonia deprivation of a glnB deletion strain, glutamine synthetase can be deadenylylated as effectively as in the wild-type strain. We describe a new operon, glnK amtB , which encodes a homologue of P_II and a putative ammonia transporter. We cloned and overexpressed glnK and found that the expressed protein had almost the same molecular weight as P_II, reacted with polyclonal P_II antibody, and was 67% identical in terms of amino acid sequence with Escherichia coli P_II. Like P_II, purified GlnK can activate the adenylylation of glutamine synthetase in vitro , and, in vivo , the GlnK protein is uridylylated in a glnD -dependent fashion. Unlike P_II, however, the expression of glnK depends on the presence of UTase, nitrogen regulator I (NRI), and absence of ammonia. Because of a NRI and a σ^N (σ⁵⁴) RNA polymerase-binding consensus sequence upstream from the glnK gene, this suggests that glnK is regulated through the NRI/NRII two-component regulatory system. Indeed, in cells grown in the presence of ammonia, glutamine synthetase deadenylylation upon ammonia depletion depended on P_II. Possible regulatory implications of this conditional redundancy of P_II are discussed.