Spiralin polymorphism in strains of Spiroplasma citri is not due to differences in posttranslational palmitoylation

Abstract

Spiralin is defined as the major membrane protein of the helical mollicute Spiroplasma citri. According to the S. citri strain used, spiralin shows polymorphism in its electrophoretic mobility. The spiralin gene sequences of eight S. citri strains were determined by direct sequencing of the PCR-amplified genes. All spiralins were found to be 241 amino acids long, except for the spiralin of strain Palmyre, which is 242 amino acids long. The molecular masses calculated from these sequences did not explain the differences observed in the electrophoretic mobilities. In all of the spiralins examined, the first 24 N-terminal amino acids were conserved, including a cysteine at position 24, and had the features of typical signal peptides of procaryotic lipoproteins. When S. citri strains were grown in the presence of [ ]palmitic acid labeled, and it had almost the same electrophoretic mobility irrespective of the spiralins used. Samples of the unlabeled peptide fragments from the four representative strains had slightly different electrophoretic mobilities (⌬DaХ800 Da); however, these were much smaller than those of the whole spiralins before formic acid hydrolysis (⌬DaХ8,000 Da). These results suggest that spiralin polymorphism in S. citri is not due to differences in posttranslational modification by palmitic acid and is certainly a structural property of the whole protein or could result from an unidentified posttranslational modification of spiralin.Among the mollicutes (i.e., wall-less eubacteria with low GϩC contents and small genomes), spiroplasmas are characterized by their helical morphology and motility. Spiroplasma citri, the first spiroplasma to have been cultured, is the causal agent of ''Stubborn'' disease of citrus (24) and affects many other plant species. S. citri also multiplies in various leafhoppers, which act as insect vectors. The spiroplasmas belong to the recently established order Entomoplasmatales, which also contains Entomoplasma and Mesoplasma species (31). All of these mollicutes are associated with arthropods, mainly insects. The spiroplasmas, like the mycoplasmas, ureaplasmas, mesoplasmas, and entomoplasmas, but unlike the acholeplasmas, use the triplet UGA, usually a stop codon, as a tryptophan codon (8,21).Spiralin is the most abundant membrane protein in S. citri (36). Its amphiphilic and antigenic properties (34, 38) and its membrane location and arrangement (30, 35) have been extensively studied. The spiralin gene has been identified, cloned, and expressed in Escherichia coli cells (19). Several forms of spiralin were produced in E. coli, including a major form with a slightly larger molecular mass than that of spiralin in S. citri. S. citri spiralin is free of tryptophan, and its gene has no UGA codon (6). This feature explains why the gene could be efficiently translated in E. coli at a time when it was not yet known that in spiroplasmas, UGA is not read as a stop codon but instead is read as a tryptophan codon. However, the function of spiralin in the spiroplasma cell remains undet...

Versatile Use oforiCPlasmids for Functional Genomics ofMycoplasma capricolumsubsp.capricolum

“…capricolum/pSPI. A polypeptide of 29 kDa (a mass close to that of spiralin) (13,37), specific to the M. capricolum subsp. capricolum/pSPI transformant ( Fig.…”

Entry of Spiroplasma citri into Circulifer haematoceps Cells Involves Interaction between Spiroplasma Phosphoglycerate Kinase and Leafhopper Actin

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Molecular Biology and Pathogenicity of Mycoplasmas

“…Furthermore, membrane lipoproteins are among the most dominant antigens in mollicutes, and a majority of the mycoplasma cell surface antigens known to undergo antigenic and/or size variation are lipoproteins (see "Antigenic variation" below). Cultivation of mollicutes in the presence of labeled palmitate or myristate labels a significant number of membrane proteins (75,126,132,323,453).…”