Characterisation of chalcone isomerase gene isolated from Pueraria montana var.lobata plant

Chalcone isomerase (CHI) is well-known as an important enzyme in the biosynthetic pathways such as flavonoid, isoflavonoid, and anthocyanin biosynthesis. The enzyme was investigated in some kinds of plants in Fabaceae but no research was conducted about the CHI gene of Pueraria montana var. lobata (P. lobata) in Vietnam. In order to provide more information and characterisation of the gene, our study isolated the CHI gene by RT-PCR and Sangersequencing. The sequence of the CHIgene was analysed with nucleotide and deduced amino acid sequences to find the main domains. A full-length CDS of CHI gene from P. lobata is 672 bp encoded 224 amino acids. By using bioinformatic tools to compare, the isolated gene shared 99.7% homology with the same species reference (code D63577.1). Two different nucleotides in the gene were altered the amino acids in the protein, but the differences have not happened in active sites. Additionally, the conserved amino acids related to active catalysis of a hydrogen bond network also appeared in the P. lobataCHI gene. SWISS-MODEL was used to build the complete protein modeling showing that P. lobataCHI protein was the most similar with CHI of Medicago sativa - was defined structure in which all alpha-helix and beta-helix were completelyhomologies.

The Sorghum (Sorghum bicolor) Brown Midrib 30 Gene Encodes a Chalcone Isomerase Required for Cell Wall Lignification

In sorghum (Sorghum bicolor) and other C4 grasses, brown midrib (bmr) mutants have long been associated with plants impaired in their ability to synthesize lignin. The brown midrib 30 (Bmr30) gene, identified using a bulk segregant analysis and next-generation sequencing, was determined to encode a chalcone isomerase (CHI). Two independent mutations within this gene confirmed that loss of its function was responsible for the brown leaf midrib phenotype and reduced lignin concentration. Loss of the Bmr30 gene function, as shown by histochemical staining of leaf midrib and stalk sections, resulted in altered cell wall composition. In the bmr30 mutants, CHI activity was drastically reduced, and the accumulation of total flavonoids and total anthocyanins was impaired, which is consistent with its function in flavonoid biosynthesis. The level of the flavone lignin monomer tricin was reduced 20-fold in the stem relative to wild type, and to undetectable levels in the leaf tissue of the mutants. The bmr30 mutant, therefore, harbors a mutation in a phenylpropanoid biosynthetic gene that is key to the interconnection between flavonoids and monolignols, both of which are utilized for lignin synthesis in the grasses.

Overexpression of chalcone isomerase A gene in Astragalus trigonus for stimulating apigenin

Apigenin is one of the most studied flavonoids and is widely distributed in the plant kingdom. Apigenin exerts important antioxidant, antibacterial, antifungal, antitumor activities, and anti-inflammatory effects in neurological or cardiovascular disease. Chalcone isomerase A (chiA) is an important enzyme of the flavonoid biosynthesis pathway. In order to enhance the apigenin production, the petunia chi A gene was transformed for Astragalus trigonus. Bialaphos survived plants were screened by PCR, dot blot hybridization and RT-PCR analysis. Also, jasmonic acid, salicylic acid, chitosan and yeast extract were tested to evaluate their capacity to work as elicitors for apigenin. Results showed that yeast extract was the best elicitor for induction of apigenin with an increase of 3.458 and 3.9 fold of the control for calli and cell suspension culture, respectively. Transformed cell suspension showed high apigenin content with a 20.17 fold increase compared to the control and 6.88 fold more than the yeast extract treatment. While, transformed T1 calli derived expressing chiA gene produced apigenin 4.2 fold more than the yeast extract treatment. It can be concluded that the highest accumulation of apigenin was obtained with chiA transgenic cell suspension system and it can be utilized to enhancement apigenin production in Astragalus trigonus.

Biochemical and Molecular Characterization of the Rice Chalcone Isomerase Family

Chalcone isomerase (CHI) is a key enzyme in flavonoid biosynthesis. In plants, CHIs occur in multigene families, and they are divided into four types, types I–IV. Type I and II CHIs are bona fide CHIs with CHI activity, and type III and IV CHIs are non-catalytic members with different functions. Rice contains seven CHI family genes (OsCHIs). Molecular analysis suggested that OsCHI3 is a type I CHI, and the other OsCHIs were classified into types III and IV. To elucidate their biochemical functions, OsCHI1, OsCHI3, OsCHI6, and OsCHI7 were expressed in Escherichia coli, and the recombinant OsCHI proteins were purified. An activity assay of recombinant OsCHIs showed that OsCHI3 catalyzed the isomerization of naringenin chalcone and isoliquiritigenin, whereas the other recombinant OsCHIs had no CHI activity. OsCHI3 also exhibited a strong preference to naringenin chalcone compared to isoliquiritigenin, which agrees well with the catalytic properties of type I CHIs. These results ascertain OsCHI3 to be a bona fide CHI in rice. OsCHI3 and the other OsCHIs were expressed constitutively throughout the rice growth period and different tissues. OsCHI3 expression was induced immediately in response to ultra-violet (UV) stress, suggesting its involvement in the biosynthesis of sakuranetin, a flavonoid phytoalexin in rice.

Identification and Characterization of Chalcone Isomerase Genes Involved in Flavonoid Production in Dracaena cambodiana

Dragon’s blood is a traditional medicine in which flavonoids are the main bioactive compounds; however, the underlying formation mechanism of dragon’s blood remains largely poorly understood. Chalcone isomerase (CHI) is the key enzyme in the flavonoid biosynthesis pathway. However, CHI family genes are not well understood in Dracaena cambodiana Pierre ex Gagnep, an important source plant of dragon’s blood. In this study, 11 CHI family genes were identified from D. cambodiana, and they were classified into three types. Evolutionary and transcriptional profiling analysis revealed that DcCHI1 and DcCHI4 might be involved in flavonoid production. Both DcCHI1 and DcCHI4 displayed low expression levels in stem under normal growth conditions and were induced by methyl jasmonate (MeJA), 6-benzyl aminopurine (6-BA, synthetic cytokinin), ultraviolet-B (UV-B), and wounding. The recombinant proteins DcCHI1 and DcCHI4 were expressed in Escherichia coli and purified by His-Bind resin chromatography. Enzyme activity assay indicated that DcCHI1 catalyzed the formation of naringenin from naringenin chalcone, while DcCHI4 lacked this catalytic activity. Overexpression of DcCHI1 or DcCHI4 enhanced the flavonoid production in D. cambodiana and tobacco. These findings implied that DcCHI1 and DcCHI4 play important roles in flavonoid production. Thus, our study will not only contribute to better understand the function and expression regulation of CHI family genes involved in flavonoid production in D. cambodiana but also lay the foundation for developing the effective inducer of dragon’s blood.