Likui Zhang

The chemical diversity of antifungal sorbicillinoids produced by the marine-derived fungus Trichoderma reesei HK1-12 was investigated by using bioassay-guided separation combined with molecular networking technology. A novel sorbicillinoid monomer, trichocillin A (1), along with four known analogues (4-5), was isolated from the fungal cultures. Furthermore, 12 monomeric, dimeric, trimeric, or hybrid sorbicillinoids (6-17) were tentatively identified from the molecular networks, six of which (7, 8, 11-13, and 15) represent previously unreported structures. Vertinolide (2) exhibited antifungal activity against Rhizoctonia solani with an inhibition rate of 90.9% at 50 μg/mL in vitro and comparable in vivo curative effects at 400 μg/mL (66.5%) to those of the spore suspension of HK1-12 (69.8%) and the positive control tebuconazole·trifloxystrobin (71.9%). This study indicated that sorbicillinoids are key antibiosis agents responsible for T. reesei HK1-12 as a potential biocontrol agent of R. solani.

Phytoene synthase (PSY) catalyzes the committed and rate-limiting step in carotenoid biosynthesis by condensing two geranylgeranyl diphosphate molecules into 15-cis-phytoene. As the gateway enzyme, PSY directs metabolic flux toward diverse carotenoids, which functions as essential pigments, antioxidants, and provitamin A precursors in nutrition and agriculture. PSY enzymes, widespread across biological kingdoms, have evolved lineage-specific regulatory mechanism while retaining conserved catalytic features, including hydrophobic loops and Asp-rich motifs, indicating a universal synthesis mechanism. This review summarizes evolutionary phylogeny and structure-function relationship of PSYs from prokaryotes to eukaryotes, highlighting their roles as key biotechnological targets for metabolic engineering to improve crop nutritional quality, stress resilience, and biofortification. Future research should elucidate PSY's regulation, catalytic machinery, and rate-limiting control over carotenoid profiles.

Lycopene β-cyclase (LCYb) serves as a pivotal gatekeeper enzyme in carotenoid biosynthesis, catalyzing an enzymatic conversion of lycopene to β-carotene. While phylogenetically widespread across life domains, structural and functional characterization of archaeal LCYbs remains conspicuously understudied. Here, we report biochemical validation and mechanistic dissection of a putative LCYb from the hypersaline-adapted archaeon Haloarcula marismortui (Hma-LCYb). Heterologous expression of Hma-LCYb in the lycopene-accumulating Escherichia coli strain resulted in the efficient β-carotene biosynthesis (0.91 ± 0.01 mg/g DCW, dried cell weight), demonstrating cross-domain functionality. Remarkably, introducing the Hma-LCYb gene into the LCYb-deficient haloarchaeon Haloferax volcanii not only enabled β-carotene production but achieved the titers (0.21 ± 0.002 mg/g DCW) surpassing the endogenous bacterioruberin levels (0.06 ± 0.003 mg/g DCW), establishing the first archaeal platform for halo-adapted carotenoid engineering. Mutational analyses revealed that a combination of residues D55, W64, E82, Y140, R168, and E214 of Hma-LCYb is critical for catalyzing lycopene-to-β-carotene conversion. These findings resolve long-standing questions regarding archaeal carotenoid cyclization mechanisms and highlight biotechnological potential of extremophilic enzymes in industrial carotenoid production.

A novel azaphilone dimer, humilone A (1), along with its related monomer, humilone B (2), was isolated from cultures of the marine-derived fungus Humicola sp. HK1-8 using a molecular networking-guided discovery approach. Further investigation of the molecular family of azaphilone dimers led to the putative identification of four analogues, humilones C-F (3-6), based on systematic analysis of their characteristic MS/MS fragmentation patterns. Detailed fragmentation studies of the dimers revealed that the predominant cleavage fragments originated from C-C bond scission at the dimeric methylene bridge. Compound 1 displayed antifungal activity against Rhizoctonia solani.

Tandem mass spectrometry (MS/MS)-based molecular networking has emerged as a powerful tool for rapid dereplication of known compounds and discovery of novel structural analogues within the same metabolite class. In this study, the chemical diversity of indolo-sesquiterpene hybrids from the mangrove rhizosphere soil-derived fungus, Aspergillus terreus N4-9, was investigated by using molecular networking strategies. The known indolo-sesquiterpene hybrid terreuside B (1) along with three new analogues, terreusides C-E (2-4), were targeted isolation from the fungal cultures. Additionally, three putative new congeners, terreusides F-H (8-10), were tentatively identified through systematic analysis of their characteristic MS/MS fragmentation patterns. Detailed fragmentation studies revealed two predominant cleavage pathways for these hybrids related to fracture of the methylene bridge connecting Rings A and B (Type I) and furan opening in Ring C (Type II). Compound 2 demonstrated significant growth inhibitory activity against human gastric cancer SGC-7901 cells with an IC₅₀ value of 6.25 μM.

Phytoene synthase (PSY) is one of key enzymes in carotenogenesis that catalyze two molecules of geranylgeranyl diphosphate to produce phytoene. PSY is widespread in bacteria, archaea, and eukaryotes. Currently, functional role and catalytic mechanism of archaeal PSY homologues have not been fully clarified due to the limited reports. Herein, we identified a rate-limiting role of the PSY from the halophilic euryarchaeon Haloferax volcanii (HVO-PSY) in bacterioruberin biosynthesis and dissected the roles of its seven conserved residues. Compared with the parental H. volcanii strain, the recombinant HVO-PSY strain had the increased bacterioruberin yields, confirming that HVO-PSY is a rate-limiting enzyme in bacterioruberin biosynthesis in H. volcanii. Additionally, we optimized the culture condition for bacterioruberin biosynthesis in the recombinant strain: 150 g/L NaCl, 45 °C, and pH 6.5-7.5. Mutational data demonstrate that residues D47, D51, D110, D168, D172, and R178 of HVO-PSY are essential for catalysis since these recombinant mutant strains harboring these mutations in the enzyme had the reduced bacterioruberin yields relative to the recombinant wild-type strain. Overall, our findings have verified the rate-limiting role of HVO-PSY in bacterioruberin biosynthesis in H. volcanii and clarified the essential roles of its conserved residues D47, D51, D110, D168, D172, and R178 in catalysis.