Evaluation of small probes through in silico docking & screening against GDP-mannose dehydrogenase (GMD)c Alice J. C. Wahart *1,2 , Jonathan P. Dolan 1,2 , Sanaz Ahmadipour 3 , Simone Dedola 3 , Jóhannes Reynisson 2,4 ,Suat Sari 5 , Chatchakorn Eurtivong 6 , Rob A. Field 3 and Gavin J. Miller 1,2 1 Lennard-Jones Laboratory, School of Chemical and Physical Sciences, UK 2 Centre for Glycosciences, Keele University, UK 3 Department of Chemistry & Manchester Institute of Biotechnology, The University of Manchester, UK 4 School of Pharmacy and Bioengineering, Keele University, 5 Hacettepe University,Turkey. 6 Program in Chemical Sciences, Chulabhorn Graduate Institute,Thailand. In the UK cystic fibrosis (CF) is detected in 1 every 2,500 babies. CF is an inherited condition caused by mutations in the gene that expresses the cystic fibrosis transmembrane conductance regulator (CFTR) protein and characterised by the build-up of thick mucus in the lungs and digestive system. Over the lifetime of one CF patient one bacterial strain, the opportunistic gram-negative human pathogen Pseudomonas aeruginosa (PA), becomes the dominant pathogen infecting over 80% of CF patients. 1 PA under goes a phenomenon known as mucoid conversion, whereby mutations which facilitate long-term survival within the lung are favoured. Mucoid PA secretes copious amounts of the carbohydrate exopolysaccharide alginate, reinforcing the PA biofilm environment and the resultant deleterious bacterial resistance to antibiotic treatments. 2 Alginate is therefore a major factor for CF lung infections and contributes deleteriously to patient life expectancy. 3 The biosynthesis of alginate relies on a critical building block of sugar nucleotide GDP-mannuronic acid (GDP- ManA, 5 ); the action of GDP-mannose dehydrogenase (GMD) which oxidises GDP-mannose 1 to 5 is the limiting step in alginate production. As GMD does not exist in humans, the discovery of selective inhibitors that could disrupt alginate biosynthesis is an exciting area of investigation. Here we present the identification of a panel of potential inhibitors of GMD through in silico docking from the Known Drug Space Libary. 4 Of the 38 small molecules hits identified, six molecules when incubated with recombinant GMD (including 6 – 9 ) reduced NADPH production in a time dependant manner; one molecule ( 6 ) demonstrated inhibition of GMD with an of IC 50 = 16.7 ± 2.1 µM. Therefore, incubation of the six molecules individually with GMD overnight, showed evidence of one adduct between 6 and GMD by ESI-MS, two adducts with 7 and none in presence of 8 and 9 .
References 1. Li, Z. et al. JAMA. 293 (5), 581-588 (2005). 2. Pier, G.B. et al. Infect Immun. 69 (3), 1895-1901 (2001). 3. Pedersen SS, Høiby N, Espersen F, et al. Thorax 47 ,6-13 (1992). 4. Bade R, Chan HF, Reynisson J. Eur J Med Chem . 45 (12):5646-5652 (2010).
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