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dc.contributor.authorDouthit, Stephanie Ann
dc.contributor.authorDlakic, Mensur
dc.contributor.authorOhman, Dennis E.
dc.contributor.authorFranklin, Michael J.
dc.date.accessioned2017-07-19T20:55:09Z
dc.date.available2017-07-19T20:55:09Z
dc.date.issued2005-06
dc.identifier.citationDouthit SA, Dlakic M, Ohman DE, Franklin MJ, "Epimerase active domain of Pseudomonas aeruginosa AlgG, a protein that contains a right-handed ß-helix," J Bacteriol, 2005 187(13):4573-4583en_US
dc.identifier.issn0021-9193
dc.identifier.urihttps://scholarworks.montana.edu/xmlui/handle/1/13337
dc.description.abstractThe polysaccharide alginate forms a protective capsule for Pseudomonas aeruginosa during chronic pulmonary infections. The structure of alginate, a linear polymer of ß1-4-linked O-acetylated D-mannuronate (M) and L-guluronate (G), is important for its activity as a virulence factor. Alginate structure is mediated by AlgG, a periplasmic C-5 mannuronan epimerase. AlgG also plays a role in protecting alginate from degradation by the periplasmic alginate lyase AlgL. Here, we show that the C-terminal region of AlgG contains a right-handed ß-helix (RHßH) fold, characteristic of proteins with the carbohydrate-binding and sugar hydrolase (CASH) domain. When modeled based on pectate lyase C of Erwinia chrysanthemi, the RHßH of AlgG has a long shallow groove that may accommodate alginate, similar to protein/polysaccharide interactions of other CASH domain proteins. The shallow groove contains a 324-DPHD motif that is conserved among AlgG and the extracellular mannuronan epimerases of Azotobacter vinelandii. Point mutations in this motif disrupt mannuronan epimerase activity but have no effect on alginate secretion. The D324A mutation has a dominant negative phenotype, suggesting that the shallow groove in AlgG contains the catalytic face for epimerization. Other conserved motifs of the epimerases, 361-NNRSYEN and 381-NLVAYN, are predicted to lie on the opposite side of the RHßH from the catalytic center. Point mutations N362A, N367A, and V383A result in proteins that do not protect alginate from AlgL, suggesting that these mutant proteins are not properly folded or not inserted into the alginate biosynthetic scaffold. These motifs are likely involved in asparagine and hydrophobic stacking, required for structural integrity of RHßH proteins, rather than for mannuronan catalysis. The results suggest that the AlgG RHßH protects alginate from degradation by AlgL by channeling the alginate polymer through the proposed alginate biosynthetic scaffold while epimerizing approximately every second D-mannuronate residue to L-guluronate along the epimerase catalytic face.en_US
dc.titleEpimerase active domain of Pseudomonas aeruginosa AlgG, a protein that contains a right-handed ß-helixen_US
dc.typeArticleen_US
mus.citation.extentfirstpage4573en_US
mus.citation.extentlastpage4583en_US
mus.citation.issue13en_US
mus.citation.journaltitleJournal of Bacteriologyen_US
mus.citation.volume187en_US
mus.identifier.categoryEngineering & Computer Scienceen_US
mus.identifier.doi10.1128/jb.187.13.4573-4583.2005en_US
mus.relation.collegeCollege of Engineeringen_US
mus.relation.departmentCenter for Biofilm Engineering.en_US
mus.relation.departmentChemical & Biological Engineering.en_US
mus.relation.departmentChemical Engineering.en_US
mus.relation.universityMontana State University - Bozemanen_US
mus.relation.researchgroupCenter for Biofilm Engineering.en_US
mus.data.thumbpage5en_US


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