D-arabitol metabolism in Candida albicans: studies of the biosynthetic pathway and the gene that encodes NAD-dependent D-arabitol dehydrogenase

Abstract

Candida albicans produces large amounts of the pentitol D-arabitol in culture and in infected mammalian hosts, but the functional and pathogenic significance of D-arabitol in C. albicans is not known. In this study, we sought to elucidate the pathway by which C. albicans synthesizes D-arabitol and to identify and characterize key enzymes in this pathway. C. albicans B311 produced D-[14C-l_arabitol from [14C-21glucose; Acyclic polyols are produced in large amounts by many fungi, and several functions have been ascribed to these compounds. For example, it has been proposed that fungi utilize polyols as storage or transport carbohydrates, as intracellular osmoregulatory solutes, and as metabolic reducing equivalents. Also, it has been suggested that fungal polyol metabolic pathways participate in the regulation of intracellular NAD(P)/NAD(P)H ratios and pH levels (14, 16). In 1979, Kiehn et al. (19) showed that cultures of the human pathogen Candida albicans produced large amounts of the acyclic pentitol D-arabitol and that people with invasive candidiasis had higher concentrations of arabitol in the blood than did normal or superficially colonized controls. Subsequent studies established that tissue and body fluid arabitol concentrations rose directly in proportion to fungal load in animals with C. albicans infection (42), that the excess arabitol in infected animals and humans was D-arabitol (the enantiomer produced by Candida spp.) (2, 43), that most humans with invasive candidiasis had high serum D-arabitol levels (11,44), and that these levels declined with effective therapy (40). Thus, D-arabitol is a quantitative diagnostic marker for invasive Candida infections.Several groups have investigated the clinical diagnostic implications of D-arabitol metabolism by C. albicans (9, 31, 36), but more basic aspects of D-arabitol metabolism have not been studied. Consequently, the metabolic pathway by which C. albicans synthesizes D-arabitol is not known, nor have the functional and pathogenetic significances of D-arabitol metabolism in C. albicans been examined. The goals of this study were to elucidate the D-arabitol biosynthetic pathway in C. albicans and to identify and characterize key * Corresponding author. enzymes in this pathway. This report describes (i) metabolic labeling studies that suggest that the pentose pathway intermediate D-ribulose-5-PO4 is the major metabolic precursor of D-arabitol in C. albicans, (ii) the presence in C. albicans lysates of a previously unknown NAD-dependent D-arabitol dehydrogenase (ArDH), and (iii) molecular cloning and analysis of the gene that encodes ArDH. Saccharomyces cerevisiae BWG 1-7A (MA4Ta his4-119 leu2-112 ura3-52 adel-100; from J. Loper, University of Cincinnati) was cultured in YPD or in 0.5% yeast extract-1% peptone-1% D-arabitol (YP-D-arab). Uracil prototrophs were selected on synthetic complete medium lacking uracil (12). MATERIALS AND METHODS StrainsEscherichia coli JM109 (45) and DH5-a (GIBCO BRL, Gaithersburg, Md.) were grown in LB medium (31) supplemented...

Monosomy of a specific chromosome determines l -sorbose utilization: A novel regulatory mechanism in Candida albicans

“…In particular, the conserved residues Gly43, Gly47, and Gly49 apparently correspond to the highly conserved NAD-binding site of many dehydrogenases (23). Comparisons of Sou1p revealed 57% similarities and 39.5% identities with C. albicans arabinitol dehydrogenase (24), and 54.5% similarities and 37.4% identities with Candida tropicalis arabinitol dehydrogenase (25). Thus, we suggest that Sou1p corresponds to L-sorbose dehydrogenase.…”

Saccharide composition in atmospheric particulate matter in the southwest US and estimates of source contributions

“…Although trehalose resembles sucrose both structurally and functionally as a carrier of energy and carbon (Wiemken, 1990), these two sugars originate differently. Trehalose is widely present in bacteria, fungi, lower plants and invertebrates (Bieleski, 1982), serving as a protectant against stressful environmental conditions (Chaturvedi, Bartiss, & Wong, 1997;Eleutherio et al, 1993;Wong, Murray, Castellanos, & Croen, 1993). Sucrose, on the other hand, is prevalent in the phloem of plants and has been found to be enriched in pollen grains (Bieleski, 1995;Pacini, 2000).…”

D-arabitol metabolism in Candida albicans: construction and analysis of mutants lacking D-arabitol dehydrogenase

“…C. albicans lysates were tested for ArDH catalytic activity as previously described (28). Briefly, reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) was measured spectrophotometrically (A 578 ) in cuvettes containing 12.5 mM D-arabitol, 35 mM HEPES (N-2-hydroxyethylpiperazine-NЈ-2-ethanesulfonic acid; pH 8.5), 0.2 mM NAD or NADP, 0.2 mM phenazine methosulfate, 0.4 mM MTT, and the diluted cell supernatant.…”