A vast class of weak, millimolar-affinity molecular interactions governs cellular function, yet their quantitative characterization has remained largely beyond conventional methods. For over a century, biochemistry has worked within a concentration-based framework where molarity scales with molecular number per volume (N/V), and experiments have usually, often implicitly, changed concentration by moving N while holding V fixed. The weak-interaction measurement bottleneck arises from this paradigm: reading weak binding through bulk concentration requires concentrations beyond practical limits, a framework constraint rather than one of instrumental sensitivity. Here we show that shifting experimental control from N to accessible volume V overcomes this bottleneck and opens previously intractable affinity ranges through nanoscale spatial confinement. Controlling V means controlling what biochemists have called "local concentration" and "proximity effects," recasting these long-ambiguous notions as quantitative variables grounded in first principles. Implemented in DNA nanocavities, the approach showed that geometric arrangement alone can override solution-phase binding hierarchies. The same spatial control quantified a protein-peptide interaction of order 10 mM from femtomoles per well, totalling under a picomole per titration. Even so, a standard plate reader gave a signal-to-noise ratio near 10^3, leaving headroom for still weaker interactions. The affinity-and-geometry readout also enabled rational screening for protein-protein-interaction modulators, identifying compounds that enhance weak associations by reweighting local encounters rather than binding tightly on their own or forming a stable ternary complex. Together, this volume-based paradigm and its implementation provide a general strategy for probing and modulating previously inaccessible biochemical phenomena.

Enzymes that process two substrates through sequential steps recognize structural motifs on the substrates that define selectivity at each reaction step. However, whether and how selectivity emerges in sequential, multi-substrate reactions remains incompletely understood. Here, we hypothesized that first-substrate binding reshapes the enzyme-substrate intermediate to program second-substrate selectivity. Using a transpeptidase, ancestral sortase variant (AcSE5), as a model to test the hypothesis, we developed a Contact Reporter assay that enables rapid and quantitative profiling of ligation across systematically diversified first-second substrate combinations. This comprehensive analysis revealed that single-residue changes in the first substrate generate distinct selectivity profiles for subsequent nucleophiles. To extend this analysis beyond peptide-based substrates, we further established a fluorescence readout and visualized selectivity across a chemically diverse set of 79 primary amines. Together, our findings reveal how selectivity emerges in sequential, multi-substrate reactions: first-substrate binding biases second-substrate preference via the enzyme–substrate intermediate, offering a generalizable framework for directing ligation outcomes in transpeptidases and related bioconjugation platforms

Abstract Strigolactones (SLs) are structurally diverse plant hormones and rhizosphere signals whose receptor-mediated functions are increasingly accessible through molecular tools. This Highlight Review focuses on receptor-centric assay platforms, including fluorogenic probes and TR-FRET systems, that enable quantitative evaluation of SL perception. These tools have accelerated the discovery of antagonists and agonists, clarified receptor selectivity and structure-activity relationships, and, together with advances in SL synthesis, support the development of selective agrochemical and breeding strategies.

Striga hermonthica, a root parasitic weed that infests a diverse range of crops, germinates through the perception of host-derived strigolactones (SLs) with receptor protein ShHTL. The development of effective strategies to suppress Striga represents an urgent challenge in combating poverty and starvation in Africa. We conducted high throughput screening to identify inhibitors for SL receptor in Striga. Through the screening of 800 compounds against eight receptor proteins (ShHTL4-11), 22 compounds were discovered as the ShHTL antagonist. Striga germination assay revealed that one out of 22 compounds has inhibitory activity to SL-induced Striga germination. Structure-activity relationship study of the primary hit compound afforded the Striga germination modulator (SGM-3) with higher potency. In vitro binding assay demonstrated that SGM-3 selectively binds to ShHTL7 which is known as the dominant receptor for Striga seed germination. SGM-3 will contribute to the suppression of Striga infection and elucidation of the parasitization mechanism.

Chemical kinetics has long inferred local molecular behaviour through the flask-and-molarity pairing, where well-mixed concentrations serve as the experimental readout. Yet many biological reactions occur in structured environments. Researchers have long recognized that concentration may not carry the same operational meaning in such environments, but even local concepts such as effective molarity usually translate local effects back into a single value with units of concentration. What has been missing is the complementary path: a bench-compatible way to make local structure an experimental variable, rather than only a correction to molarity. Here we show a chemistry-geometry crossover that the flask-and-molarity interface could not make visible. In the micromolar-or-weaker affinity regime, inhibition can switch sharply out of the familiar concentration-and-affinity mode: chemical binding strength no longer determines the response, and the shape of the target's local space does. A bench-compatible interface made this switch measurable by separating bulk dose from local geometry. This blind spot arose from the hidden premise that macroscopic pooling makes a structured local state readable as a single local concentration. The chemistry-geometry crossover breaks that premise: in a structured target environment, a macroscopic assay can remain sensitive to the probability distribution of local states, so collapsing that distribution to one concentration-valued number removes the geometric control axis from the readout. By preserving that axis in the experiment, the interface bypasses molarity's hidden bottleneck and provides a routine experimental route to remeasure and reinterpret molecular interactions in structured space.

Photocleavable molecules are valuable tools for biological studies, enabling spatiotemporal activation of molecular functions within cellular environments. In particular, coumarin-based photolytic molecules are useful because of their ability to flexibly tune the wavelength of photostimulation through their structural modifications. Ideal photocleavable molecular tools require hydrolytic stability and selective susceptibility to photo stimuli. However conventional coumarin-based molecules have not simultaneously achieved both highly efficient photocleavage and hydrolysis resistance. Herein, we proposed a novel molecular design concept that introduces a silyl group into coumarin-based molecules at a position remote from the photolabile bond, creating an ideal photocleavable molecule for chemical biology tools. The established orbital effect of the remotely introduced silyl group improves the photolysis efficiency of coumarin-based molecules, while its bulkiness substantially enhances their hydrolytic stability in aqueous environments and under enzymatic conditions. Furthermore, this improvement in molecular functionality contributes to the development of high-performance protein-release biomaterials.

Photocleavable molecules are widely used in fields such as materials science and chemical biology. In particular, coumarin-based photocleavable molecules are indispensable for photomanipulation techniques in chemical biology because of their ability to flexibly adjust the wavelength of photostimulation through straightforward structural modifications. However, traditional coumarin-based molecules have several limitations, including hydrolysis in aqueous environments and susceptibility to intracellular carboxylesterases. Additionally, substituting bonds in these molecules to enhance hydrolytic stability often decreases their photolysis efficiency. Herein, we proposed a novel molecular design concept that introduces a silyl group into coumarin-based molecules at a position remote from the photolabile bond, creating an ideal photocleavable molecule for chemical biology tools. The established orbital effect of the remotely introduced silyl group improves the photolysis efficiency of coumarin-based molecules, while its bulkiness substantially enhances their hydrolytic stability in aqueous environments and under enzymatic conditions. Furthermore, this improvement in molecular functionality contributes to the development of high-performance protein-release biomaterials.

Strigolactones are one of the phytohormones, which have multiple activities on plant growth and development. Since these strigolactone activities are highly associated with crop yield, use of strigolactone could be a promising technology in modern sustainable agriculture. The major strigolactones in corn root exudates have been identified as zealactone 1a/b and zeapyranolactone. We recently disclosed the first total synthesis of zealactone 1a/b together with its biological activity in corn. Herein, we describe the design and synthesis of simplified analogues of both corn-derived strigolactones with their bioavailability in soil and their biological activities. These compounds would be potential leads for the development of synthetic strigolactones for the agronomical use in a more sustainable crop production. image

Polycyclic aromatic hydrocarbons (PAHs) are an important family of molecules in science and technology. Amino‐substituted PAHs are promising building blocks to create attractive molecules for materials science. However, the synthetic limitations have hampered to produce diverse structures of amino‐substituted PAHs. Here we describe a novel efficient synthetic method to access amino‐substituted PAHs through the electrocyclization of highly reactive keteniminium species. We demonstrated the synthesis of various amino‐substituted PAHs and disclosed some of their photophysical properties. Furthermore, combination with theoretical calculation revealed that the multiple electrocyclization reactions of keteniminium species is a stepwise process and the fusion of additional aromatic ring to the substrate accelerates the electrocyclization.

Maize (Zea mays) is a major staple crop in Africa, where its yield and the livelihood of millions are compromised by the parasitic witchweed Striga. Germination of Striga is induced by strigolactones exuded from maize roots into the rhizosphere. In a maize germplasm collection, we identified two strigolactones, zealactol and zealactonoic acid, which stimulate less Striga germination than the major maize strigolactone, zealactone. We then showed that a single cytochrome P450, ZmCYP706C37, catalyzes a series of oxidative steps in the maize-strigolactone biosynthetic pathway. Reduction in activity of this enzyme and two others involved in the pathway, ZmMAX1b and ZmCLAMT1, can change strigolactone composition and reduce Striga germination and infection. These results offer prospects for breeding Striga-resistant maize.

We describe herein a high yielding and metal-free methodology to access 9-aminophenanthrene derivatives and heterocyclic analogues starting from biaryl acetamides. The conversion of the substrates to their corresponding keteniminium salts triggers a spontaneous electrocyclization occurring at room temperature within a few minutes. DFT Calculations and competition reactions were carried out to rationalize the observed reactivity and understand the relative kinetics compared to other known reactions involving keteniminium salts.

Cage-like molecules, assembled by the coordination of multiple metal ions and organic links, are pushing new frontiers in science due to their design flexibility and the resulting diverse and unique chemical properties. This field has been advanced by two close but distinct chemistry communities. Consequently, the family of molecules referred to as coordination cages (CCs) constituted of metal-pyridine coordination bonds or metal–organic cages (MOCs) based on dinuclear tetracarboxylates paddlewheel complexes in each community had not been reviewed cross-sectionally, even though they are conceptually similar. This review article extracted and compared experimental information on a total of 197 CCs and 78 MOCs from 182 reports to identify their synthetic and structural signatures. We did not merely enumerate the reports we collected; we meta-analyzed the data extracted from the reports and highlighted both the similarities and dissimilarities between CCs and MOCs. As a result, we clarified the key parameters governing the synthetic conditions. Furthermore, we identified a new research direction by visualizing unexplored features and properties of CCs and MOCs. This review article provides a good tutorial both for researchers attempting to cross the boundary between CCs and MOCs and those who are new to the field.

Strigolactones (SLs) are plant hormones that suppress shoot branching through perception by their receptor protein DWARF 14 (D14). The artificial regulation of SL signaling has been considered a potent agricultural technique because plant architecture is strongly related to crop yield. In this communication, we describe the development of a small-molecule D14 inhibitor that functions at sub-micromolar levels. This potent inhibitor may be a lead compound for a first-in-class plant growth regulator.

Two new methods for selective deprotection of diphenylmethylsilyl (DPMS) ethers are described. Unmasking can be achieved with either catalytic amounts of perfluoro‐1‐butanesulfonyl fluoride (a SuFEx reagent) under mild, aqueous micellar conditions, or using stoichiometric amounts of 18‐crown‐6 ether in aqueous ethanol.

Strigolactones are plant hormones, which play pivotal roles in plant growth and development with potential application in sustainable agriculture. Recently, zealactone 1a/b has been identified as the major strigolactone from the root exudates of corn. Although zealactone is a promising molecule affecting signaling in the rhizosphere as well as in planta, evaluating its biological activities has been hampered by its low natural abundance and its relative chemical instability. Herein, we present the total synthesis of zealactone 1a/b based on our studies employing a [2+2]-cycloaddition strategy and a chemoselective Baeyer-Villiger oxidation to forge the gamma-butyrolactone fragment. Furthermore, we disclose the biological activities of zealactone 1a/b on corn and in soil in comparison with related synthetic analogues.

DWARF14 (D14) is a strigolactone receptor that plays a central role in suppression of shoot branching, and hence is a potential target to increase crop productions and biomass. Recently, we reported a fluorescence turn-on probe, Yoshimulactone Green (YLG), which generates a strong fluorescence upon the hydrolysis by D14-type strigolactone receptors. Herein, we applied a YLG-based in vitro assay to a high-throughput chemical screening and identified a novel small molecule DL1 as a potent inhibitor of D14. DL1 competes with endogenous strigolactones, thereby increasing the number of shoot branching in a model plant Arabidopsis as well as in rice. Thus, DL1 is expected to be useful not only as a tool to understand the biological roles of D14 receptors in plant growth and development, but also as a potent agrochemical to improve the crop yield.

Plant-derived strigolactones have diverse functions at ecological scale, including effects upon the growth of plants themselves. The parasitic plants from the family Orobanchaceae interfere with the ecological and hormonal functions of strigolactones to generate unique germination abilities based on the sensing of host-derived strigolactones. Although the recent discovery of strigolactone receptors has enabled us to begin elucidating the mechanism of strigolactone perception, how perception relates to plant parasitism is still a mystery. In this review, we explore emerging questions by introducing recent advances in strigolactone research in parasitic plants. We also attempt to construct a conceptual framework for the unique in planta dynamics of strigolactone perception uncovered through the use of fluorescent probes for strigolactone receptors. Understanding the mechanisms of strigolactone-related processes is essential for controlling the parasitic plant Striga hermonthica, which has caused devastating damage to crop production in Africa.

The phytohormone auxin indole-3-acetic acid (IAA) regulates nearly all aspects of plant growth and development. Despite substantial progress in our understanding of auxin biology, delineating specific auxin response remains a major challenge. Auxin regulates transcriptional response via its receptors, TIR1 and AFB F-box proteins. Here we report an engineered, orthogonal auxin-TIR1 receptor pair, developed through a bump-and-hole strategy, that triggers auxin signaling without interfering with endogenous auxin or TIR1/AFBs. A synthetic, convex IAA (cvxIAA) hijacked the downstream auxin signaling in vivo both at the transcriptomic level and in specific developmental contexts, only in the presence of a complementary, concave TIR1 (ccvTIR1) receptor. Harnessing the cvxIAA-ccvTIR1 system, we provide conclusive evidence for the role of the TIR1-mediated pathway in auxin-induced seedling acid growth. The cvxIAA-ccvTIR1 system serves as a powerful tool for solving outstanding questions in auxin biology and for precise manipulation of auxin-mediated processes as a controllable switch.

Aromaticity of photoexcited molecules is an important concept in organic chemistry. Its theory, Baird's rule for triplet aromaticity since 1972 gives the rationale of photoinduced conformational changes and photochemical reactivities of cyclic π-conjugated systems. However, it is still challenging to monitor the dynamic structural change induced by the excited-state aromaticity, particularly in condensed materials. Here we report direct structural observation of a molecular motion and a subsequent packing deformation accompanied by the excited-state aromaticity. Photoactive liquid crystal (LC) molecules featuring a π-expanded cyclooctatetraene core unit are orientationally ordered but loosely packed in a columnar LC phase, and therefore a photoinduced conformational planarization by the excited-state aromaticity has been successfully observed by time-resolved electron diffractometry and vibrational spectroscopy. The structural change took place in the vicinity of excited molecules, producing a twisted stacking structure. A nanoscale torque driven by the excited-state aromaticity can be used as the working mechanism of new photoresponsive materials.