Escherichia coli glutamine synthetase (GS) is a dodecamer of identical subunits which are arranged as two face-to-face hexameric rings. In the presence of 10% ammonium sulfate, wild type GS exhibits a pH-dependent "salting out' with a pKa of 4.51. Electron micrographs indicate that the pH-dependent aggregation corresponds to a highly specific self-assembly of GS tubules, which result from stacking of individual dodecamers. This stacking of dodecamers is similar to the metal ion-induced GS tubule formation previously described. Site-directed mutagenesis experiments indicate that the N-terminal helix of each subunit is involved in the salting out reaction, as it is in the metal-induced stacking. A single substitution of alanine for His4 completely abolishes the (NH4)2SO4-induced aggregation. However, the H4C mutant protein does nearly completely precipitate under the same salting out conditions. Mutations at other residues within the helix have no effect on the stacking reaction. Differential catalytic activity of unadenylylated GS versus adenylylated GS has been used to determine whether wild type dodecamers "complement' the H4A mutant in the stacking reaction. The complementation experiments indicate that His4 residues on both sides of the dodecamer-dodecamer interfaces are not absolutely required for salting out, although the wild type dodecamers clearly stack preferentially with other wild type dodecamers. Approximately 20% of the protein precipitated from the mixtures containing the wild type GS and the H4A mutant is the mutant. The implications of these results for protein engineering are discussed.