Cloning of the major protein of the Caulobacter crescentus periodic surface layer: detection and characterization of the cloned peptide by protein expression assays

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Abstract
A precisely ordered crystalline array is found on the surface of the bacterium Caulobacter crescentus CB15. Using an immunological assay, we identified recombinant bacteriophage clones expressing the predominant protein of this structure from a lambda 1059 library of C. crescentus CB15 DNA. A single 4.4-kilobase HindIII fragment encoded a polypeptide whose antigenic determinants, molecular weight, and peculiar solubilization properties were identical with those of the authentic predominant polypeptide (130K) of the surface array. The 130K protein was produced as a discrete product as a result of gene transcription initiated from a lambda promoter; several experiments suggested that the Caulobacter promoter for this gene is not efficiently recognized by the Escherichia coli transcription machinery. Genomic Southern analysis revealed a single copy of the 130K protein gene per genome. The 130K protein gene was hybridized with DNA of two closely related laboratory strains of C. crescentus which have lost their ability to produce a surface array. One of these strains, CB2, possesses an homologous copy of the 130K gene, whereas DNA from the other strain, CB13B1a, showed a lesser degree of hybridization to the 130K gene probe; genomic fragments which did hybridize were of different sizes in CB13 as compared with those of CB15. These findings are discussed in relation to studies of the surface array function and its role in cellular morphogenesis in this stalk-forming bacterium.