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Inhibition of the chitin biosynthetic enzyme, glucosamine synthase (EC., L-glutamine:D-Fructose-6-phosphate amidotransferase) has been shown to be a promising approach to the development of antifungal agents targeting the cell wall. The best known inhibitors of glucosamine synthase possess epoxy-ketones and {dollar}\\alpha,\\beta{dollar} unsaturated carbonyls. In vivo, these inherently reactive moieties are easily deactivated by nucleophiles like glutathione. Hence we decided to develop inhibitors lacking such highly reactive groups. Development of multisubstrate analogs is a logical approach to inhibition of multisubstrate enzymes. The compounds investigated in our study possess structural modifications of both glutamine and fructose-6-phosphate so as to bind to both binding sites on the enzyme leading to a potent inhibition of the enzyme. IC{dollar}\\sb{lcub}50{rcub}{dollar} data indicated AP3 to be a potent inhibitor of glucosamine synthase from Candida albicans. The correlation between the distance from the {dollar}\\alpha{dollar}-carbon of the amino acid to the phosphonate phosphorus indicates that shorter chain lengths provide better inhibitors. CGS19755, a structural analog of AP5 showed a higher potency than AP4, indicating that the rigidity and hydrophobicity enhances binding to this site. Our data indicates that the distance between the phosphate binding site and the amino acid binding site is {dollar}\\rm{lcub}\\approx{rcub}3\\ A\\sp\\circ{dollar}. However AP3 does not inhibit the C. albicans enzyme when a protease inhibitor like PMSF was used in the isolation procedure. One explanation for this observation could be that proteases released during previous isolation procedure cleave some portion on the enzyme allowing AP3 to access the active site and inhibit the enzyme. In the presence of PMSF, this is prevented and AP3 cannot reach the active site. AP3 did not inhibit glucosamine synthase isolated from S. cereviseae with or without using PMSF. This reinforces existing evidence that significant differences exist between the enzyme protein from S. cereviseae and C. albicans. The unstable nature of the partially purified enzyme extract however did not allow us to determine a K{dollar}\\rm\\sb{lcub}i\\ app{rcub}{dollar} for these compounds. Hence we cannot state whether the inhibition is multisubstrate or selectively competitive with respect to either substrate. Repeating this study with the homogeneously purified enzyme obtainable from recently developed clones would clarify these areas.