Microbial Sterolomics as a Chemical Biology Tool

Abstract

Metabolomics has become a powerful tool in chemical biology. Profiling the human sterolome has resulted in the discovery of noncanonical sterols, including oxysterols and meiosis-activating sterols. They are important to immune responses and development, and have been reviewed extensively. The triterpenoid metabolite fusidic acid has developed clinical relevance, and many steroidal metabolites from microbial sources possess varying bioactivities. Beyond the prospect of pharmacognostical agents, the profiling of minor metabolites can provide insight into an organism's biosynthesis and phylogeny, as well as inform drug discovery about infectious diseases. This review aims to highlight recent discoveries from detailed sterolomic profiling in microorganisms and their phylogenic and pharmacological implications.

Keywords: algal sterols; ergosterol biosynthesis; infectious disease; lipidomics; oxyphytosterol; pharmacognosy; phytosterol; sterolomics.

Figure 1

Structure and numbering systems of sterols and steroids. (a) Δ⁵ end product inserts from mammals, fungi, and vascular plants, respectively, cholesterol 1, ergosterol 2, and sitosterol 3. (b) Examples of steroidal metabolites important in human biology for F-MAS 4, TT-MAS 5, 25-hydroxycholesterol 6. (c) Examples of steroidal metabolites from nonhuman sources with bioactivity, fusidic acid 7, ergosterol peroxide 8, and squalamine 9. The numbering system shown here, and used in this manuscript, is the conventional system [1]. Designations of α and β within the sterol nucleus signify below and above the plane. Unrelated to nucleus α and β, substituents on C24 are also designated α and β to reflect the C24 stereochemistries of sitosterol and ergosterol, respectively, as drawn above. Carbon numbering is provided on 1–4, and stereochemistries at C8, C9, C14, and C16 on structure 1 are hereafter implied on structures, unless otherwise annotated as in fusidic acid. Molecular features for each structure are provided relative to 5α-cholestanol for clarity. For a complete list of systematic names of compounds, see Table A1.

Figure 2

Truncated hypothetical pathway of fungal ergosterol 2 biosynthesis from squalene 10. Inhibitor targets of squalene epoxidase (SqE) by allylamines, e.g., terbinafine 19, sterol C14-demethylase (14-SDM = CYP51) by azoles, e.g., voriconazole 20, fluconazole 21, itraconazole 22, and posaconazole 23, sterol C14-reductase (14-SR) and sterol C8(7)-isomerase (8(7)-SI) by morpholines, e.g., fenpropimorph 24, and sterol C24-methyltransferase (24-SMT) by 25-azalanosterol 25 or 24(R,S),25-epiminolanosterol 26 are highlighted at the biosynthetic steps they block. 3-SR; sterol C3 reductase, 24-SR, sterol C24 reductase.

Figure 3

Comparative phytosterol biosynthesis in the photosynthetic lineage from the protosterol cycloartenol 27. In algae, 24-methyl and 24-ethyl sterols arise from a bifurcation of products of biomethylation by sterol methyltransferase (SMT); In higher plants, they arise from alternate pathways from the intermediate 24(28)-methylene lophenol 30, which can be methylated again or metabolized to campesterol 36. Red methyl groups from SMT co-substrate S-adenosyl methionine (AdoMet) are annotated to show hypothetical labeling patterns of Δ⁵ sterols as discussed in [45,50]. An additional 15 algal sterols were reported in [45]. Truncated fungal phytosterol biosynthesis from protosterol lanosterol 12 is illustrated in Figure 2.

Figure 4

Molecular structures of algal sterols.

Figure 5

Comparative cholesterol biosynthesis between humans and arthropods. (a) Late-stage cholesterol biosynthesis in humans from de novo zymosterol 15. (b) Proposed synthesis of cholesterol in herbivorous insects via dealkylation of dietary plant sterols (sitosterol) [58]. (c) Amphipod Gammarus roeselii can dealkylate the side chain of Δ⁷ algal sterols, such as fungisterol and chondrillasterol, but cannot produce cholesterol [56].

Figure 6

Sterol structures from various dinoflagellates.

Figure 7

Abbreviated biosynthetic sterol pathway and composition in T. brucei. In T. brucei, C4 is demethylated before C14, contrary to mammalian and fungal pathways (cf. Figure 2). Values are percentage sterol composition reported by Zhou et al. [43]. Dietary cholesterol 1 accounted for 20.0 %, and other components were 16 (0.1%), 30 (1.0%), 48 (1.0%), 57 (8.0%), and others (0.2%). 24,24-Dimethylcholesta-5,7,25(27)-trienol and 86 and protothecasterol 87 were not detected in this composition, but have been reported in subsequent studies [42,66], respectively.

Figure 8

26-Fluorinated sterol analogues. (a) Fluorinated inhibitors of T. brucei 24-SMT and growth. (b) Metabolites of 88 identified from T. brucei and HEK cells [67].

Figure 9

Structures of amebasterols.

Figure 10

Growth-phase dependence of predominant sterols in A. castellanii. R = Me and Et. Adapted from [36].

Figure 11

Sterolomic identification of ergosterol biosynthesis inhibitors (EBIs) in fungi. Red arrows signify increase or decrease in sterols within the profile of inhibited cultures relative to non-inhibited cultures. (a) Oxazole amidoester-treated cultures of C. albicans decrease in ergosterol and increase in lanosterol and by-products obtusifoliol and eburicol, indicating disruption of 14-SDM activity [77,78]. (b) Posaconazole-treated cultures of Rhizopus arrhizus decrease in ergosterol and ergosta-5,7-dienol and increase in lanosterol, obtusifoliol, and eburicol, and produce toxic 14-methylergosta-8,24(28)-dien-3β,6α-diol [79]. (c) ent-Isoalantolactone-treated cultures of C. albicans decrease in ergosterol and increase in lanosterol and zymosterol, indicating disruption of 24-SMT activity [80].

Figure 12

EBIs FR171456 105 and VT-1129 106, confirmed by sterolomic analysis.

Figure 13

Steryl peroxides discussed in text.

Figure 14

Steryl acetates discussed in text.

Figure 15

Sterols bearing a 3-membered ring.

Figure 16

Bioactive sterols and steroids. Activities are given in Table 2.