The outer cell-wall layer ofSpirillum serpensVHA, composed of a hexagonal array of macromolecules, was dissociated from 'cleaned' cell-wall fragments with 1.5 Mguanidine hydrochloride, pH 7.0. The soluble material contained 98% protein, 2% carbohydrate, and no ethanolamine, phosphate, RNA, or DNA. Evidence for the homogeneity of the isolated cell-wall protein was obtained by sedimentation velocity, polyacrylamide disc gel electrophoresis, molecular sieve chromatography on Sephadex G-200, and sucrose density gradients. The protein was acidic and had a minimum molecular weight of about 48 000 daltons calculated from the amino acid analysis. A molecular weight of between 125 000 and 150 000 daltons was obtained for the protein by polyacrylamide disc-gel electrophoresis, sucrose density gradients, and molecular sieve chromatography. The sedimentation rate of the protein was considerably less in 1.5 Mguanidine hydrochloride, pH 7.0, than in its absence although the protein remained homogenous under both conditions. Self-assembly of the purified protein into its original hexagonal layer on a template of wall fragments from which the protein had been removed previously was obtained when the mixture was incubated in the presence, but not in the absence of Ca2+. It seems likely that the particles (hexagons) seen on the surface of the cell are an assembly of trimers.