Conserved transmembrane glycine residues in the Shigella flexneri polysaccharide co-polymerase protein WzzB influence protein-protein interactions
Authors: Papadopoulos, Magdalene; Elizabeth Ngoc Hoa Tran; Murray, Gerald Laurence; Morona, Renato
The O antigen (Oag) component of lipopolysaccharides (LPS) is crucial for virulence and Oag chain-length regulation is controlled by the polysaccharide co-polymerase class 1 (PCP1) proteins. Crystal structure analyses indicate that structural conservation among PCP1 proteins is highly maintained, however the mechanism of Oag modal-chain-length control remains to be fully elucidated. Shigella flexneri PCP1 protein WzzB(SF) confers a modal-chain length of 10-17 Oag repeat units (RUs), whereas the Salmonella enterica Typhimurium PCP1 protein WZZB(ST) confers a modal-chain length of similar to 16-28 Oag RUs. Both proteins share >70% overall sequence identity and contain two transmembrane (TM1 and TM2) regions, whereby a conserved prolineglycine-rich motif overlapping the TM2 region is identical in both proteins. Conserved glycine residues within TM2 are functionally important, as glycine to alanine substitutions at positions 305 and 311 confer very short Oag modal-chain length (similar to 2-6 Oag RUs). In this study, WzzB(SF) was co-expressed with WzzB(ST) in S. flexneri and a single intermediate modal-chain length of similar to 11-21 Oag RUs was observed, suggesting the presence of Wzz:Wzz interactions. Interestingly, co-expression of WzzB(SF) with WzzB(G305A/G311A) conferred a bimodal LPS Oag chain length (despite over 99% protein sequence identity), and we hypothesized that the proteins fail to interact. Co-purification assays detected His(6)-WzzB(SF) co-purifying with FLAG-tagged WzzB(ST) but not with FLAG-tagged WzzB(G305A/G311A), supporting our hypothesis. These data indicate that the conserved glycine residues in TM2 are involved in Wzz:Wzz interactions, and provide insight into key interactions that drive Oag modal length control.
Survival of foodborne bacteria on strawberries and antibacterial activities of Hibiscus sabdariffa extracts and chemical sanitizers on strawberries
JOURNAL OF FOOD SAFETY
Authors: Gomez-Aldapa, Carlos A.; Portillo-Torres, Lizbeth A.; Villagomez-Ibarra, Jose R.; Rangel-Vargas, Esmeralda; Tellez-Jurado, Alejandro; Cruz-Galvez, Andres M.; Castro-Rosas, Javier
The potential of 13 foodborne bacteria to attach and survival on strawberries and the antimicrobial activities of Hibiscus sabdariffa calyx extracts and chemical sanitizers against foodborne bacteria on strawberries were investigated. Strawberries were inoculated with thirteen foodborne bacteria: Listeria monocytogenes, Shigella flexneri, Staphylococcus aureus, Salmonella Typhimurium, S. Typhi, S. Montevideo, Vibrio cholerae O1, and six pathotypes of Escherichia coli: O157:H7, Non-O157:H7-Shiga toxin-producing, enteropathogenic, enterotoxigenic, enteroinvasive, and enteroaggregative. The antibacterial effects of four roselle calyx extracts (in water, methanol, acetone, and ethyl acetate), sodium hypochlorite, colloidal silver, and acetic acid against all pathogenic bacteria were evaluated on contaminated strawberries. All foodborne bacteria attached and survived on strawberries at least 15 days. Acetonic and methanolic roselle calyx extracts caused a greater reduction in the concentration (until 3.8-log(10) reduction in some cases) of all foodborne bacteria than the sodium hypochlorite, colloidal silver and acetic acid on contaminated strawberries. Practical applicationsThe strawberry is an important crop in United States, Spain, and Mexico. However, fresh strawberries have been associated with foodborne disease caused by E. coli O157:H7 and Salmonella. There is little information about the attachment and survival of foodborne bacteria on strawberries. A number of studies have shown that chemical agents used to eliminate pathogenic bacteria on fruits have a limited antimicrobial effect. Roselle calyx extracts are reported to have an antimicrobial effect. This study provides information about the attachment and survival of different foodborne bacteria on strawberries and information on the significant impact of dry roselle calyx extracts may be potentially useful in disinfection procedures of strawberries.