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We have partially characterized the biochemical parameters of glutamine synthetase from Klebsiella pneumoniae and have shown that the differential affinity of adenylylated and unadenylylated glutamine synthetase for adenosine diphosphate provides a convenient means of determining the adenylylation state. Using this assay procedure, we examined the relationship between the adenylylation state and the expression of other genes involved in nitrogen assimilation. We observed no correlation between the adenylylation state and the expression of histidase, glutamine synthetase, glutamate synthase, glutamate dehydrogenase, and urease in aerobic cultures.

We have used P1 transduction to create intergeneric hybrid strains of enteric bacteria by moving the genA and hut genes between Klebsiella aerogenes, Escherichia coli and Salmonella typhimurium. The use of E. coli as the recipient in such transductions permits the construction of episomes and specialized transducing phage containing non-E. coli material. The effect of host restriction modification and deoxyribonucleic acid homology on the frequency of intergeneric transduction of these loci has been examined.

Regulation of the expression of the histidase coded by hutk of Klebsiella aerogenes in Salmonella typhimurium and in Escherichia coli and of the expression of the histidase coded by huts of S. typhimurium in E. coli was investigated. The hutk histidase was found to be sensitive to catabolite repression in K. aerogenes and in E. coli, but insensitive to catabolite repression in S. typhimurium; huts histidase has previously been shown to be catabolite sensitive in all three organisms. The expression of both hutk and huts histidase in E. coli was activated by nitrogen starvation. Apparently, the glutamine synthetase of E. coli may activate the formation of some glutamate- and ammonia-producing enzymes.

P1-sensitive mutants of Klebsiella aerogenes were isolated and the gene order of the hut region was then determined using P1-mediated transduction. The genes are located in the Klebsiella chromosome between gal and bio as in Salmonella typhimurium. The gene order, gal, hutI, hutG, hutC, huU, hutH, bio is also the same as that observed in S. typhimurium.

We examined the regulation of nitrogen metabolism in four classes (glnA, glnB, glnF, and glnG) of Gln- auxotrophs of Klebsiella pneumoniae. These studies indicate that glutamine synthetase does not directly mediate the physiological response to NH4+ in this organism. We present evidence suggesting that the effect of NH4+ on the expression of genes involved in nitrogen metabolism involves the products of the glnF and glnG genes.

We investigated the regulation of genes concerned with nitrogen metabolism by oxygen in the facultative anaerobe Klebsiella pneumoniae. We found oxygen to be required for the expression of the hut operons; the effect of O2 on the glutamine synthetase and urease was less pronounced than on the hut operons. Glutamine synthetase was transiently repressed during the transition from an aerobic to an anaerobic environment. Regulation of hut by O2 suppressed the effect of nitrogen limitation on the expression of these genes.