title: A New Synthetic Pathway Towards Legionaminic Acid
Creators: name: Carter, James
Publisher: name: Griffith University
Year: 2018
Subjects: subject: Legionaminic acid
subject: Synthetic pathway
subject: Nonulosonic acids
subject: Flagellar glycans
subject: Leg derivatives

description: This thesis describes the results of a PhD project aimed at researching a new synthetic pathway towards legionaminic acid analogues. Legionaminic acid is a bacterial family of nonulosonic acids expressed in a terminal position on the flagellar glycans and are considered to be a virulence factor in the pathogenic bacteria that possess them. At the outset of this project several examples of Leg synthesis had been published. Whilst three syntheses of Leg derivatives have been published while the research described in this thesis was being carried out, the potential of all the published syntheses of Leg to efficiently generate C-7 analogues is very limited. There are a number of C-7 analogues of legionaminic acid that occur in nature, and additionally it is the functionality at C-7 which primarily separates the bacterial nonulosonic acids from the those found in animals (in addition to being 9-deoxy). It was therefore the goal of this project to develop a synthetic chemical pathway that is adaptable towards a number of C-7 analogues of legionaminic acid, and potentially other analogues that may be considered of value. It was also key to our approach that we were able to produce these analogues of legionaminic acid from a common precursor. Chapter one of this thesis introduces in detail the literature background of the nonulosonic acids, in particular the bacterial nonulosonic acids pseudaminic acid, legionaminic acid, and the recently discovered acinetaminic acid. It discusses the natural occurrence of these molecules and gives a complete overview of all existing synthetic pathways: chemical, enzymatic, and chemoenzymatic. Chapter two describes our initial approach towards the synthesis of legionaminic acid and 7-epi-legionaminic acid. N-Acetylneuraminic acid was selected as the starting material due to its commercial availability and structural similarities to legionaminic acid. The transformations required to convert Neu5Ac to legionaminic acid were the deoxygenation of carbons seven and nine, and the introduction of an acetamide to carbon seven. We based our initial approach on the synthesis of 8-epi-pseudaminic acid previously completed in our group. To this end, Neu5Ac was selectively protected to leave the only free hydroxyl group at carbon seven, which was activated and displaced with a nitrogen nucleophile. The remainder of the approach consisted of the removal of the protecting groups, acylation of the C-7 nitrogen, and deoxygenation of carbon nine, however the deoxygenation step provided us with insurmountable difficulties. In chapter three, the investigation into the synthesis of Leg2en analogues is discussed. Analogues of nonulosonic acids with a double bond between carbons two and three have proven to be of value in the past, and with the discovery of a putative glycohydrolase enzyme for pseudaminic acid, Leg2en and Pse2en analogues could well prove to be just as valuable. An analogous approach to that discussed in chapter 2 is taken, with the difference being that Neu5Ac is converted to Neu5Ac2en prior to selective protection and the introduction of the C-7 nitrogen. Once again, the acetamido functionality was successfully introduced to carbon seven, but the deoxygenation of carbon nine could not be accomplished. In chapter four, a revised approach towards legionaminic acid and 7-epi-legionaminic acid is described. This approach avoided the previously encountered problems with C-9 deoxygenation by performing it early in the synthesis and introducing the nitrogen functionality to C-7 of 9-deoxy-Neu5Ac. With this new approach, 7-epi-legionaminic acid and legionaminic acid were both successfully synthesised, with the functionalisation of the C-7 nitrogen step being performed at the very end of the synthesis, prior only to the removal of protecting groups. This therefore allows for the investigation into a number of different functionalisation steps that could be performed on the protected 7-amino-legionaminic acid substrate. The final chapter, chapter five, details our investigations into an interesting and important by-product that we isolated during the activation of the C-7 hydroxyl on multiple substrates. This by-product was identified as a dihydro-1,3-oxazine derivative, which had formed between carbon seven and the acetamide of carbon five. We explored the different conditions under which this by-product formed and found that we could control relatively well whether the C-7 activated Neu5Ac would convert to the oxazine by changing the base used, and in what proportions. We also investigated opening this oxazine by-product, with the hope that we could use its inverted stereochemistry as an avenue towards legionaminic acid stereochemistry, which we were eventually able to do.