1. |
- Balonova, Lucie, et al.
(författare)
-
Characterization of protein glycosylation in Francisella tularensis subsp holarctica
- 2012
-
Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 11:7
-
Tidskriftsartikel (refereegranskat)abstract
- FTH_0069 is a previously uncharacterized strongly immunoreactive protein that has been proposed to be a novel virulence factor in Francisella tularensis. Here, the glycan structure modifying two C-terminal peptides of FTH_0069 was identified utilizing high resolution, high mass accuracy mass spectrometry, combined with in-source CID tandem MS experiments. The glycan observed at m/z 1156 was determined to be a hexasaccharide, consisting of two hexoses, three N-acetylhexosamines, and an unknown monosaccharide containing a phosphate group. The monosaccharide sequence of the glycan is tentatively proposed as X-P-HexNAc-HexNAc-Hex-Hex-HexNAc, where X denotes the unknown monosaccharide. The glycan is identical to that of DsbA glycoprotein, as well as to one of the multiple glycan structures modifying the type IV pilin PilA, suggesting a common biosynthetic pathway for the protein modification. Here, we demonstrate that the glycosylation of FTH_0069, DsbA, and PilA was affected in an isogenic mutant with a disrupted wbtDEF gene cluster encoding O-antigen synthesis and in a mutant with a deleted pglA gene encoding pilin oligosaccharyltransferase PglA. Based on our findings, we propose that PglA is involved in both pilin and general F. tularensis protein glycosylation, and we further suggest an inter-relationship between the O-antigen and the glycan synthesis in the early steps in their biosynthetic pathways. Molecular & Cellular Proteomics 11: 10.1074/mcp.M111.015016, 1-12, 2012.
|
|
2. |
- Syrén, Per-Olof, 1981-
(författare)
-
On electrostatic effects, minimal motion and other catalytic strategies used by enzymes
- 2011
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Enzymes are powerful biocatalysts that provide rate accelerations of up to 1019 fold compared to the corresponding uncatalyzed reaction in solution. The origin of the remarkable performance displayed by enzymes has fascinated and puzzled researchers for over a hundred years. It is clear that the catalytic effect is a consequence of the higher degree of transition state stabilization for the enzyme catalyzed reaction compared to the corresponding uncatalyzed reaction. It is still not well understood exactly how this transition state stabilization occurs and the relative importance of various catalytic effects are discussed. Catalytic effects involving electrostatics, near attack conformers, dynamic effects and an economy in atomic motion are discussed in this thesis. The importance of electrostatic effects is corroborated in this thesis. A single hydrogen bond in transition state constitutes an important difference between amidases and esterases. A hydrogen bond in transition state is found in all sixteen analyzed amidases representing ten different reaction mechanisms and eleven different folding families. The hydrogen bond is shown to be either substrate assisted or enzyme assisted. The role of this hydrogen bond is to assist nitrogen inversion in amidases. Esterases lack this interaction in transition state and therefore they are very poor catalysts in the hydrolysis of amides. Electrostatic interactions are found to facilitate proton transfer that enhances the rate of lipase catalyzed N-acylation of amino alcohols. In this thesis electrostatic effects in the substrate are shown to be important for the lipase catalyzed transacylation of acrylates The α,β-double bond present in acrylates introduce electronic effects that has the consequence of restricting the conformational freedom of the substrate in its ground state to two flat conformations, s-cis and s-trans. It is shown that acrylates form near attack conformers (NACs) from their ground state s-cis/s-trans planar conformations. The ability of the enzyme to accommodate such apparent s-cis/s-trans substrate conformations dictates the probability to form productive transition states and thus the reaction rate. Dynamic effects are important in enzymes. In this thesis it is found that a point mutation increases the flexibility of a neighbouring residue in Candida antarctica lipase B. This allows the mutated enzyme to explore conformations not accessible for the wild-type enzyme. The dynamics has the effect to decrease steric interactions in transition state with concomitant rate increase for the transacylation of methyl methacrylate. In this thesis an economy of atomic motion during enzyme catalysis is observed. Nitrogen inversion in amidases constitutes an interesting example. A rotation as part of the reaction mechanism for amide bond hydrolysis would involve much more motion.
|
|
3. |
- Uppgård, Lise-Lott, 1970-
(författare)
-
Nonionic surfactants : A multivariate study
- 2002
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In this thesis technical nonionic surfactants are studied using multivariate techniques. The surfactants studied were alkyl ethoxylates (AEOs) and alkyl polyglucosides (APGs). The aquatic toxicity of the surfactants towards two organisms, a shrimp and a rotifer, was examined. The specified effect was lethality, LC50, as indicated by immobilisation. In a comparative study, the LC50 values obtained were used to develop two different types of model. In the log P model the toxicity was correlated to log P alone, while in the multivariate model several physicochemical variables, including log P, were correlated to the toxicity. The multivariate model gave smaller prediction errors than the log P model. Further, the change in reactivity when a surfactant mixture was added to dissolving pulp under alkaline conditions was studied, using the amount of residual cellulose as a measure of the reactivity. Ten AEO/APG mixtures were tested, and the mixture with greatest potential was studied in more detail. An optimum in the amount of added surfactant was found that seems to coincide, according to surface tension measurements, with the CMC.
|
|