Cloning and Characterization of Oxidosqualene Cyclases Involved in Taraxasterol, Taraxerol and Bauerenol Triterpene Biosynthesis in Taraxacum coreanum

2019 ◽  
Vol 60 (7) ◽  
pp. 1595-1603 ◽  
Author(s):  
Jung Yeon Han ◽  
Hye-Jeong Jo ◽  
Eun Kyung Kwon ◽  
Yong Eui Choi
Keyword(s):  
Functional Characterization ◽  
Taraxacum Officinale ◽  
Oxidosqualene Cyclases ◽  
Oxidosqualene Cyclase ◽  
The Family ◽  
Triterpene Synthases ◽  
Taraxacum Coreanum ◽  
Heterologous Expression In Yeast ◽  
Triterpene Biosynthesis

Abstract Triterpenes, consisting of six isoprene units, are one of the largest classes of natural compounds in plants. The genus Taraxacum is in the family Asteraceae and is widely distributed in the Northern Hemisphere. Various triterpenes, especially taraxerol and taraxasterol, are present in Taraxacum plants. Triterpene biosynthesis occurs through the action of oxidosqualene cyclase (OSC), which generates various types of triterpenes from 2,3-oxidosqualene after the rearrangement of the triterpene skeleton. However, no functional characterization of the OSC genes involved in triterpene biosynthesis, except for a lupeol synthase in Taraxacum officinale, has been performed. Taraxacum coreanum, or Korean dandelion, grows in Korea and China. Putative OSC genes in T. coreanum plants were isolated by transcriptome analysis, and four of these (TcOSC1, TcOSC2, TcOSC3 and TcOSC4) were functionally characterized by heterologous expression in yeast. Both TcOSC1 and TcOSC2 were closely related to dammarenediol-II synthases. TcOSC3 and TcOSC4 were strongly grouped with β-amyrin synthases. Functional analysis revealed that TcOSC1 produced several triterpenes, including taraxasterol; Ψ-taraxasterol; α-, β- and δ-amyrin; and dammarenediol-II. TcOSC2 catalyzed the production of bauerenol and another unknown triterpene, TcOSC3 catalyzed the production of β-amyrin. TcOSC4 catalyzed the production of taraxerol. Moreover, we identified taraxasterol, ψ-taraxasterol, taraxerol, lupeol, δ-amyrin, α-amyrin, β-amyrin and bauerenol in the roots and leaves of T. coreanum. Our results suggest that TcOSC1, TcOSC2, TcOSC3 and TcOSC4 are key triterpene biosynthetic enzymes in T. coreanum. These enzymes are novel triterpene synthases involved in the production of taraxasterol, bauerenol and taraxerol.

Functional characterization of high-affinity Na+/dicarboxylate cotransporter found in Xenopus laevis kidney and heart

2005 ◽  
Vol 289 (5) ◽  
pp. C1159-C1168 ◽  
Author(s):  
Naomi Oshiro ◽  
Ana M. Pajor
Keyword(s):  
Xenopus Laevis ◽  
Cdna Sequence ◽  
Citric Acid Cycle ◽  
Functional Characterization ◽  
Activation Kinetics ◽  
Electrophysiological Properties ◽  
High Affinity ◽  
The Family ◽  
Kidney Liver

The SLC13 gene family includes sodium-coupled transporters for citric acid cycle intermediates and sulfate. The present study describes the sequence and functional characterization of a SLC13 family member from Xenopus laevis, the high-affinity Na+/dicarboxylate cotransporter xNaDC-3. The cDNA sequence of xNaDC-3 codes for a protein of 602 amino acids that is ∼70% identical to the sequences of mammalian NaDC-3 orthologs. The message for xNaDC-3 is found in the kidney, liver, intestine, and heart. The xNaDC-3 has a high affinity for substrate, including a Km for succinate of 4 μM, and it is inhibited by the NaDC-3 test substrates 2,3-dimethylsuccinate and adipate. The transport of succinate by xNaDC-3 is dependent on sodium, with sigmoidal activation kinetics, and lithium can partially substitute for sodium. As with other members of the family, xNaDC-3 is electrogenic and exhibits inward substrate-dependent currents in the presence of sodium. However, other electrophysiological properties of xNaDC-3 are unique and involve large leak currents, possibly mediated by anions, that are activated by binding of sodium or lithium to a single site.

Cloning and characterization of a 2,3-oxidosqualene cyclase from Eleutherococcus trifoliatus

Holzforschung ◽  
2013 ◽  
Vol 67 (4) ◽  
pp. 463-471 ◽  
Author(s):  
Li-Ting Ma ◽  
Sheng-Yang Wang ◽  
Yen-Hsueh Tseng ◽  
Yi-Ru Lee ◽  
Fang-Hua Chu
Keyword(s):  
Physiological Role ◽  
Active Site Residues ◽  
Reading Frame ◽  
Oxidosqualene Cyclases ◽  
Oxidosqualene Cyclase ◽  
Acid Protein ◽  
Leaf Tissues ◽  
New Perspective ◽  
Amino Acid Protein

Abstract The 2,3-oxidosqualene cyclases (OSCs) are a family of enzymes that have an important role in plant triterpene biosynthesis. In this study, an OSC gene designed EtLUS from Eleutherococcus trifoliatus has been cloned. EtLUS includes a 2292-bp open reading frame and encodes a 763-amino acid protein. EtLUS has an MLCYCR motif, which is conserved in lupeol synthases. Comparison of active-site residues and gene expression in yeast showed that EtLUS synthesizes lupeol. However, EtLUS has the highest sequence identity with β-amyrin synthases from Araliaceae rather than lupeol synthases, adding new perspective to the evolution of the OSCs of Araliaceae. Furthermore, EtLUS is upregulated in leaf tissues under methyl jasmonate treatment, which can be interpreted that lupeol and its derivatives play an ecological and physiological role in plant defense against pathogens and insect herbivores.

Identification and functional characterization of a novel locust peptide belonging to the family of insect growth blocking peptides

Peptides ◽  
2015 ◽  
Vol 74 ◽  
pp. 23-32 ◽  
Author(s):  
Tewodros Firdissa Duressa ◽  
Kurt Boonen ◽  
Yoichi Hayakawa ◽  
Roger Huybrechts
Keyword(s):  
Functional Characterization ◽  
Insect Growth ◽  
The Family

Cloning, Expression And Functional Characterization Of Two Oxidosqualene Cyclases From Hoodia Gordonii

Planta Medica ◽  
2016 ◽  
Vol 82 (05) ◽  
Author(s):  
I Parveen ◽  
M Wang ◽  
J Zhao ◽  
AG Chittiboyina ◽  
N Techen ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Oxidosqualene Cyclases ◽  
Hoodia Gordonii

Mechanism and Inhibition of Matrix Metalloproteinases

2019 ◽  
Vol 26 (15) ◽  
pp. 2609-2633 ◽  
Author(s):  
Linda Cerofolini ◽  
Marco Fragai ◽  
Claudio Luchinat
Keyword(s):  
Matrix Metalloproteinases ◽  
Functional Characterization ◽  
Structural Data ◽  
Therapeutic Strategies ◽  
Outer Cell ◽  
Drug Candidates ◽  
The Family ◽  
Outer Cell Membrane ◽  
Membrane Bound

Matrix metalloproteinases hydrolyze proteins and glycoproteins forming the extracellular matrix, cytokines and growth factors released in the extracellular space, and membrane-bound receptors on the outer cell membrane. The pathological relevance of MMPs has prompted the structural and functional characterization of these enzymes and the development of synthetic inhibitors as possible drug candidates. Recent studies have provided a better understanding of the substrate preference of the different members of the family, and structural data on the mechanism by which these enzymes hydrolyze the substrates. Here, we report the recent advancements in the understanding of the mechanism of collagenolysis and elastolysis, and we discuss the perspectives of new therapeutic strategies for targeting MMPs.

scholarly journals Polyphosphate-dependent synthesis of ATP and ADP by the family-2 polyphosphate kinases in bacteria

2008 ◽  
Vol 105 (46) ◽  
pp. 17730-17735 ◽  
Author(s):  
Boguslaw Nocek ◽  
Samvel Kochinyan ◽  
Michael Proudfoot ◽  
Greg Brown ◽  
Elena Evdokimova ◽  
...  
Keyword(s):  
Sinorhizobium Meliloti ◽  
Molecular Mechanisms ◽  
Catalytic Mechanism ◽  
Functional Characterization ◽  
High Energy ◽  
Inorganic Polyphosphate ◽  
The Family ◽  
Adp Crystal ◽  
Intracellular Energy

Inorganic polyphosphate (polyP) is a linear polymer of tens or hundreds of phosphate residues linked by high-energy bonds. It is found in all organisms and has been proposed to serve as an energy source in a pre-ATP world. This ubiquitous and abundant biopolymer plays numerous and vital roles in metabolism and regulation in prokaryotes and eukaryotes, but the underlying molecular mechanisms for most activities of polyP remain unknown. In prokaryotes, the synthesis and utilization of polyP are catalyzed by 2 families of polyP kinases, PPK1 and PPK2, and polyphosphatases. Here, we present structural and functional characterization of the PPK2 family. Proteins with a single PPK2 domain catalyze polyP-dependent phosphorylation of ADP to ATP, whereas proteins containing 2 fused PPK2 domains phosphorylate AMP to ADP. Crystal structures of 2 representative proteins, SMc02148 from Sinorhizobium meliloti and PA3455 from Pseudomonas aeruginosa, revealed a 3-layer α/β/α sandwich fold with an α-helical lid similar to the structures of microbial thymidylate kinases, suggesting that these proteins share a common evolutionary origin and catalytic mechanism. Alanine replacement mutagenesis identified 9 conserved residues, which are required for activity and include the residues from both Walker A and B motifs and the lid. Thus, the PPK2s represent a molecular mechanism, which potentially allow bacteria to use polyP as an intracellular energy reserve for the generation of ATP and survival.

scholarly journals Ferric Enterochelin Transport in Yersinia enterocolitica: Molecular and Evolutionary Aspects

1999 ◽  
Vol 181 (20) ◽  
pp. 6387-6395 ◽  
Author(s):  
S. Schubert ◽  
D. Fischer ◽  
J. Heesemann
Keyword(s):  
Gene Cluster ◽  
Yersinia Enterocolitica ◽  
Insertional Mutagenesis ◽  
Yersinia Pseudotuberculosis ◽  
Functional Characterization ◽  
In Vitro Transcription ◽  
E Coli ◽  
The Family

ABSTRACT Yersinia enterocolitica is well equipped for siderophore piracy, encompassing the utilization of siderophores such as ferrioxamine, ferrichrome, and ferrienterochelin. In this study, we report on the molecular and functional characterization of theYersinia fep-fes gene cluster orthologous to theEscherichia coli ferrienterochelin transport genes (fepA, fepDGC, and fepB) and the esterase gene fes. In vitro transcription-translation analysis identified polypeptides of 30 and 35 kDa encoded byfepC and fes, respectively. A frameshift mutation within the fepA gene led to expression of a truncated polypeptide of 40 kDa. The fepD,fepG, and fes genes of Y. enterocolitica were shown to complement corresponding E. coli mutants. Insertional mutagenesis of fepD orfes genes abrogates enterochelin-supported growth ofY. enterocolitica on iron-chelated media. In contrast toE. coli, the fep-fes gene cluster inY. enterocolitica consists solely of genes required for uptake and utilization of enterochelin (fep) and not of enterochelin synthesis genes such as entF. By Southern hybridization, fepDGC and fes sequences could be detected in Y. enterocolitica biotypes IB, IA, and II but not in biotype IV strains, Yersinia pestis, andYersinia pseudotuberculosis strains. According to sequence alignment data and the coherent structure of the Yersinia fep-fes gene cluster, we suggest early genetic divergence of ferrienterochelin uptake determinants among species of the familyEnterobacteriaceae.

scholarly journals Functional characterization of the transcription regulator WhiB1 from Gordonia sp. IITR100

Microbiology ◽  
2020 ◽  
Vol 166 (12) ◽  
pp. 1181-1190
Author(s):  
Pooja Murarka ◽  
Aditi Keshav ◽  
Bintu Kumar Meena ◽  
Preeti Srivastava
Keyword(s):  
Type Species ◽  
Functional Characterization ◽  
Rhodococcus Erythropolis ◽  
Multiple Sequence ◽  
Content Type ◽  
Transcription Regulator ◽  
Link Type ◽  
Family Protein ◽  
The Family

WhiB is a transcription regulator which has been reported to be involved in the regulation of cell morphogenesis, cell division, antibiotic resistance, stress, etc., in several members of the family Actinomycetes . The present study describes functional characterization of a WhiB family protein, WhiB1 (protein ID: WP_065632651.1), from Gordonia sp. IITR100. We demonstrate that WhiB1 affects chromosome segregation and cell morphology in recombinant Escherichia coli , Gordonia sp. IITR100 as well as in Rhodococcus erythropolis . Multiple sequence alignment suggests that WhiB1 is a conserved protein among members of the family Actinomycetes . It has been reported that overexpression of WhiB1 leads to repression of the biodesulfurization operon in recombinant E. coli , Gordonia sp. IITR100 and R. erythropolis . A WhiB1-mut containing a point mutation Q116A in the DNA binding domain of WhiB1 led to partial alleviation of repression of the biodesulfurization operon. We show for the first time that the WhiB family protein WhiB1 is also involved in repression of the biodesulfurization operon by directly binding to the dsz promoter DNA.

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