Tolyporphins A–R are unusual tetrapyrrole macrocycles produced by the non-axenic filamentous cyanobacterium HT-58-2. A putative biosynthetic gene cluster for biosynthesis of tolyporphins (here termed BGC-1) was previously identified in the genome of HT-58-2. Here, homology searching of BGC-1 in HT-58-2 led to identification of similar BGCs in seven other filamentous cyanobacteria, including strains Nostoc sp. 106C, Nostoc sp. RF31YmG, Nostoc sp. FACHB-892, Brasilonema octagenarum UFV-OR1, Brasilonema octagenarum UFV-E1, Brasilonema sennae CENA114 and Oculatella sp. LEGE 06141, suggesting their potential for tolyporphins production. A similar gene cluster (BGC-2) also was identified unexpectedly in HT-58-2. Tolyporphins BGCs were not identified in unicellular cyanobacteria. Phylogenetic analysis based on 16S rRNA and a common component of the BGCs, TolD, points to a close evolutionary history between each strain and their respective tolyporphins BGC. Though identified with putative tolyporphins BGCs, examination of pigments extracted from three cyanobacteria has not revealed the presence of tolyporphins. Overall, the identification of BGCs and potential producers of tolyporphins presents a collection of candidate cyanobacteria for genetic and biochemical analysis pertaining to these unusual tetrapyrrole macrocycles.

Tolyporphins are distinctive tetrapyrrole natural products found singularly in a filamentous cyanobacterial‐microbial holobiont (termed HT‐58‐2) from Micronesia. The absorption and fluorescence features of tolyporphins resemble those of chlorophyll a, complicating direct analysis of culture samples. Treatment of the crude (unfractionated) organic extract (CH2Cl2/2‐propanol, 1:1) of HT‐58‐2 cultures with NaBH4 in methanol causes reduction of the peripheral ketone auxochromes, whereupon tolyporphins (predominantly 7,17‐dioxobacteriochlorins) exhibit a bathochromic shift (λabs ˜ 676 → ˜ 700 nm) and chlorophyll a (a 131‐oxochlorin) exhibits a hypsochromic shift (λabs 665 → 634 nm). Fluorescence excitation spectroscopy (at 368 and 491 nm with λem 710 nm) enabled detection of reduced tolyporphins amidst abundant reduced chlorophyll a (1:19 ratio), a detection sensitivity >5 times that without reduction. The resulting assay combines simple sample preparation from non‐axenic cultures at microscale quantities (2 mL, 2 μm), absence of any fractionation procedures, and fluorescence detection. Tolyporphins were readily detected in cultures of HT‐58‐2 at reasonable growth periods in the absence of environmental stressors, which was not possible previously.

Aggregation-induced emission (AIE) probes that can be triggered by enzymatic activity are valuable for applications across the life sciences.

A targeted strategy for treating cancer is antibody-directed enzyme prodrug therapy, where the enzyme attached to the antibody causes conversion of an inactive small-molecule prodrug into an active drug. A limitation may be the diffusion of the active drug away from the antibody target site. A related strategy with radiotherapeutics entails enzymatically promoted conversion of a soluble to insoluble radiotherapeutic agent, thereby immobilizing the latter at the target site. Such a molecular brachytherapy has been scarcely investigated. In distinct research, the advent of molecular designs for aggregation-induced emission (AIE) suggests translational use in molecular brachytherapy. Here, several 2-(2-hydroxyphenyl)benzothiazole substrates that readily aggregate in aqueous solution (and afford AIE) were elaborated in this regard. In particular, (1) the 2-(2-hydroxyphenyl) unit was derivatized to bear a pegylated phosphodiester that imparts water solubility yet undergoes enzymatic cleavage, and (2) a p-phenol unit was attached to the benzo moiety to provide a reactive site for final-step iodination (here examined with natural abundance iodide). The pegylated phosphodiester-iodinated benzothiazole undergoes conversion from aqueous-soluble to aqueous-insoluble upon treatment with a phosphatase or phosphodiesterase. The aggregation is essential to molecular brachytherapy, whereas the induced emission of AIE is not essential but provides a convenient basis for research development. Altogether, 21 compounds were synthesized (18 new, 3 known via new routes). Taken together, blending biomedical strategies of enzyme prodrug therapy with materials chemistry concerning substances that undergo AIE may comprise a step forward on the long road toward molecular brachytherapy.