Oligomerization of the cytoplasmic fragment from the aspartate receptor of Escherichia coli.
We have observed that a 31-kDa cloned fragment from the Escherichia coli aspartate receptor exhibits a reversible monomer-oligomer reaction. The fragment, derived from the cytoplasmic region of the receptor (c-fragment), contains the signaling functions of the receptor. The wild-type and nine missense mutant fragments were analyzed. The latter were selected by the effect of the mutations on the signaling properties of the intact receptor, which induced either persistent smooth swimming or tumbling in bacteria [Mutoh, N., Oosawa, K., & Simon, M. I. (1986) J. Bacteriol. 167, 992-998]. In pH 7.0 buffer, the mutations caused five out of the six smooth mutant c-fragments to form oligomers, while neither the three tumble mutant nor wild-type fragments exhibited significant oligomer formation. At a lower pH (5.5), all of the fragments displayed some tendency to form oligomers. The equilibria between the monomer and the oligomers were monitored by gel permeation chromatography (GPC) which resolved two to three forms with apparent molecular weights between 110,000 and 270,000. The proportions of the different forms depended on concentration, indicating an association-dissociation reaction. Static light scattering (SLS) was used to demonstrate that the solution molecular mass of the wild-type c-fragment was 31 kDa and not 110 kDa as indicated by chromatography. One oligomer-forming c-fragment (S461L) eluted as the monomer and one other form, which was determined to be a dimer by SLS. The weight-average molecular weights, calculated from GPC data as a function of protein concentration, agreed well with the weight-average molecular weights obtained by SLS for this mutant.(ABSTRACT TRUNCATED AT 250 WORDS)
Long
D G
DG
Department of Chemistry, University of Massachusetts, Amherst 01003.
Weis
R M
RM
D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003593:Cytoplasm / Q000737:chemistry*; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D008970:Molecular Weight; D016296:Mutagenesis; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D017459:Receptors, Amino Acid / Q000737:chemistry* / Q000235:genetics; D015398:Signal Transduction
D046911:Macromolecular Substances; D010446:Peptide Fragments; D017459:Receptors, Amino Acid; C031959:aspartic acid receptor