A biochemical correlate of dimorphism in a zygomycete Benjaminiella poitrasii: characterization of purified NAD-dependent glutamate dehydrogenase, a target for antifungal agents

Abstract

The fungal organisms, especially pathogens, change their vegetative (Y, unicellular yeast and H, hypha) morphology reversibly for survival and proliferation in the host environment. NAD-dependent glutamate dehydrogenase (NAD-GDH, EC 1.4.1.2) from a non-pathogenic dimorphic zygomycete Benjaminiella poitrasii was previously reported to be an important biochemical correlate of the transition process. The enzyme was purified to homogeneity and characterized. It is a 371 kDa native molecular weight protein made up of four identical subunits. Kinetic studies showed that unlike other NAD-GDHs, it may act as an anabolic enzyme and has more affinity towards 2-oxoglutarate than L-glutamate. Chemical modifications revealed the involvement of single histidine and lysine residues in the catalytic activity of the enzyme. The phosphorylation and dephosphorylation study showed that the NAD-GDH is present in active phosphorylated form in hyphal cells of B. poitrasii. Two of the 1,2,3 triazole linked β-lactam-bile acid conjugates synthesized in the laboratory (B18, B20) were found to be potent inhibitors of purified NAD-GDH which also significantly affected Y-H transition in B. poitrasii. Furthermore, the compound B20 inhibited germ tube formation during Y-H transition in Candida albicans strains and Yarrowia lipolytica. The possible use of NAD-GDH as a target for antifungal agents is discussed.

Multiple Forms of Glutamate Dehydrogenase in Animals: Structural Determinants and Physiological Implications

“…Reactivation via dephosphorylation is catalyzed by protein phosphatases. In contrast, phosphorylation of NADP + -specific GDH in the lower fungi Benjaminiella poitrasii was shown to activate the enzyme [130,131]. Differences in the GDH phosphorylation level were shown under nitrogen starvation (NAD + -specific GDH in C. utilis and S. cerevisiae ) or in yeast and mycelium states (NADP + -specific GDH in B. poitrasii) .…”

Inhibitors of amino acids biosynthesis as antifungal agents

“…NAD(P)-dependent glutamate dehydrogenase was also reported as one of the points of metabolic control of fungal morphological transformation (Joshi et al 2013 ; Peters and Sypherd 1979 ). 1,2,3-Triazole-linked β-lactam-bile acid conjugates: B18 and B20 (Fig.…”

“…10 d, e) were found to be the potent inhibitors of NAD-dependent glutamate dehydrogenase from Benjaminiella poitrasii , with K i = 27.38 and 18.28 μM, respectively and significantly affected yeast-to-mycelia transition of this fungus. Furthermore, the compound B20 inhibited germ tube formation during Y → M transition of Candida albicans and Yarrowia lipolytica (Joshi et al 2013 ).…”

Possible mechanism of antifungal phenazine-1-carboxamide fromPseudomonassp. against dimorphic fungiBenjaminiella poitrasiiand human pathogenCandida albicans

“…), 1,2,3 triazole‐linked β ‐lactam bile acid conjugates and bile acid dimers linked with triazole and bis β ‐lactam (Joshi et al . ). In this study, investigations on bioassay‐guided isolation and characterization of the active compound from Pseudomonas sp., its antifungal activity, mode of action studies in B. poitrasii and validation in C. albicans are presented.…”