Exploring the Diversity and Regulation of Apocarotenoid Metabolic Pathways in Plants

In plants, carotenoids are subjected to enzyme-catalyzed oxidative cleavage reactions as well as to non-enzymatic degradation processes, which produce various carbonyl products called apocarotenoids. These conversions control carotenoid content in different tissues and give rise to apocarotenoid hormones and signaling molecules, which play important roles in plant growth and development, response to environmental stimuli, and in interactions with surrounding organisms. In addition, carotenoid cleavage gives rise to apocarotenoid pigments and volatiles that contribute to the color and flavor of many flowers and several fruits. Some apocarotenoid pigments, such as crocins and bixin, are widely utilized as colorants and additives in food and cosmetic industry and also have health-promoting properties. Considering the importance of this class of metabolites, investigation of apocarotenoid diversity and regulation has increasingly attracted the attention of plant biologists. Here, we provide an update on the plant apocarotenoid biosynthetic pathway, especially highlighting the diversity of the enzyme carotenoid cleavage dioxygenase 4 (CCD4) from different plant species with respect to substrate specificity and regioselectivity, which contribute to the formation of diverse apocarotenoid volatiles and pigments. In addition, we summarize the regulation of apocarotenoid metabolic pathway at transcriptional, post-translational, and epigenetic levels. Finally, we describe inter- and intraspecies variation in apocarotenoid production observed in many important horticulture crops and depict recent progress in elucidating the genetic basis of the natural variation in the composition and amount of apocarotenoids. We propose that the illustration of biochemical, genetic, and evolutionary background of apocarotenoid diversity would not only accelerate the discovery of unknown biosynthetic and regulatory genes of bioactive apocarotenoids but also enable the identification of genetic variation of causal genes for marker-assisted improvement of aroma and color of fruits and vegetables and CRISPR-based next-generation metabolic engineering of high-value apocarotenoids.

Toehold mediated strand displacement to measure released product from self-cleaving ribozymes

This paper presents a probe comprising a fluorophore and a quencher, enabling measurement of released product from self-cleaving hammerhead ribozyme, without labeled RNA molecules, regular sampling or use of polyacrylamide gels. The probe is made of two DNA strands; one strand is labelled with a fluorophore at its 5′-end, while the other strand is labelled with a quencher at its 3′-end. These two DNA strands are perfectly complementary, but with a 3′-overhang of the fluorophore strand. These unpaired nucleotides act as a toehold, which is utilized by a detached cleaved fragment (coming from a self-cleaving hammerhead ribozyme) as the starting point for a strand displacement reaction. This reaction causes the separation of the fluorophore strand from the quencher strand, culminating in fluorescence, detectable in a plate reader. Notably, the emitted fluorescence is proportional to the amount of detached cleaved-off RNAs, displacing the DNA quencher strand. This method can replace or complement radio-hazardous unstable 32P as a method of measurement of the product release from ribozyme cleavage reactions; it also eliminates the need for polyacrylamide gels, for the same purpose. Critically, this method allows to distinguish between the total amount of cleaved ribozymes and the amount of detached fragments, resulting from that cleavage reaction.

C-Glycoside metabolism in the gut and in nature: Identification, characterization, structural analyses and distribution of C-C bond-cleaving enzymes

C-Glycosides, in which a sugar moiety is linked via a carbon-carbon (C-C) bond to a non-sugar moiety (aglycone), are found in our food and medicine. The C-C bond is cleaved by intestinal microbes and the resulting aglycones exert various bioactivities. Although the enzymes responsible for the reactions have been identified, their catalytic mechanisms and the generality of the reactions in nature remain to be explored. Here, we present the identification and structural basis for the activation of xenobiotic C-glycosides by heterocomplex C-deglycosylation enzymes from intestinal and soil bacteria. They are found to be metal-dependent enzymes exhibiting broad substrate specificity toward C-glycosides. X-ray crystallographic and cryo-electron microscopic analyses, as well as structure-based mutagenesis, reveal the structural details of these enzymes and the detailed catalytic mechanisms of their remarkable C-C bond cleavage reactions. Furthermore, bioinformatic and biochemical analyses suggest that the C-deglycosylation enzymes are widely distributed in the gut, soil, and marine bacteria.

Nickel-catalyzed Homo-coupling of Aryl Ethers with Magnesium Anthracene Reductant

Nickel-catalyzed reductive homo-coupling of aryl ethers has been achieved with Mg(anthracene)(thf)3 as a readily available low-cost reductant. DFT calculations provided a rationale for the specific efficiency of the diorganomagnesium-type two-electron reducing agent. The calculations showed that the dianionic anthracene-9,10-diyl ligand reduces the two aryl ether substrates resulting in the homo-coupling reaction through supplying the electrons to the Ni-Mg bimetallic system to form organomagnesium nickel(0)-ate complexes, which cause two sequential C–O bond cleavage reactions. The calculations also showed cooperative actions of Lewis-acidic magnesium atoms and electron-rich nickel atoms in the C–O cleavage reactions.

[38]Octaphyrin bis-Sn(IV) complexes with unique coordination geometries

Metal complexation of octaphyrin(1.1.1.1.1.1.1.1) triggers unique ring-fixation aptitudes or unexpected rearrangement (cleavage) reactions. In this paper, a unique complexation behavior of [38]octaphyrin upon tin(IV) metalation is showcased. Two new [38]octaphyrin bis-Sn(IV) complexes 2Sn and 3Sn were isolated and characterized as weakly aromatic molecules. While 2Sn with the [Formula: see text] molecular symmetry displayed a similar characteristic to octaphyrin bis-Si(IV) and bis-Ge(IV) complexes reported previously, 3Sn showed a different coordination mode that is fixed by intramolecular hydrogen bondings between pyrrolic NH and axially ligated OH on the tin ion as revealed by X-ray diffraction analysis. An unexpected dimeric structure was also observed during an attempt to grow crystals of 2Sn. These characteristic behaviors indicate that the ring-fixation aptitude of octaphyrin is quite sensitive to the nature of metal ions even for the same group 14 elements.

Transamidation of Amides and Amidation of Esters by Selective N–C(O)/O–C(O) Cleavage Mediated by Air- and Moisture-Stable Half-Sandwich Nickel(II)–NHC Complexes

The formation of amide bonds represents one of the most fundamental processes in organic synthesis. Transition-metal-catalyzed activation of acyclic twisted amides has emerged as an increasingly powerful platform in synthesis. Herein, we report the transamidation of N-activated twisted amides by selective N–C(O) cleavage mediated by air- and moisture-stable half-sandwich Ni(II)–NHC (NHC = N-heterocyclic carbenes) complexes. We demonstrate that the readily available cyclopentadienyl complex, [CpNi(IPr)Cl] (IPr = 1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene), promotes highly selective transamidation of the N–C(O) bond in twisted N-Boc amides with non-nucleophilic anilines. The reaction provides access to secondary anilides via the non-conventional amide bond-forming pathway. Furthermore, the amidation of activated phenolic and unactivated methyl esters mediated by [CpNi(IPr)Cl] is reported. This study sets the stage for the broad utilization of well-defined, air- and moisture-stable Ni(II)–NHC complexes in catalytic amide bond-forming protocols by unconventional C(acyl)–N and C(acyl)–O bond cleavage reactions.