Ion channels and G protein-coupled receptors (GPCRs) are regulated by lipids in their membrane environment. Structural studies combined with biophysical and molecular simulation investigations reveal ...interaction sites for specific lipids on membrane protein structures. For K channels, PIP
2
plays a key role in regulating Kv and Kir channels. Likewise, several recent cryo-EM structures of TRP channels have revealed bound lipids, including PIP
2
and cholesterol. Among the pentameric ligand-gated ion channel family, structural and biophysical studies suggest the M4 TM helix may act as a lipid sensor, e.g., forming part of the binding sites for neurosteroids on the GABA
A
receptor. Structures of GPCRs have revealed multiple cholesterol sites, which may modulate both receptor dynamics and receptor oligomerization. PIP
2
also interacts with GPCRs and may modulate their interactions with G proteins. Overall, it is evident that multiple lipid binding sites exist on channels and receptors that modulate their function allosterically and are potential druggable sites.
The extracellular domain (ECD) of class B1 G-protein-coupled receptors (GPCRs) plays a central role in signal transduction and is uniquely positioned to sense both the extracellular and membrane ...environments. Although recent studies suggest a role for membrane lipids in the modulation of class A and class F GPCR signaling properties, little is known about the effect of lipids on class B1 receptors. In this study, we employed multiscale molecular dynamics simulations to access the dynamics of the glucagon receptor (GCGR) ECD in the presence of native-like membrane bilayers. Simulations showed that the ECD could move about a hinge region formed by residues Q122–E126 to adopt both closed and open conformations relative to the transmembrane domain. ECD movements were modulated by binding of the glycosphingolipid GM3. These large-scale fluctuations in ECD conformation may affect the ligand binding and receptor activation properties. We also identify a unique phosphatidylinositol (4,5)-bisphosphate (PIP2) interaction profile near intracellular loop (ICL) 2/TM3 at the G-protein-coupling interface, suggesting a mechanism of engaging G-proteins that may have a distinct dependence on PIP2 compared with class A GPCRs. Given the structural conservation of class B1 GPCRs, the modulatory effects of GM3 and PIP2 on GCGR may be conserved across these receptors, offering new insights into potential therapeutic targeting.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Acetylcholinesterase, with a deep, narrow active-site gorge, attracts enormous interest due to its particularly high catalytic efficiency and its inhibitors used for treatment of Alzheimer's disease. ...To facilitate the massive pass-through of the substrate and inhibitors, "breathing" motions to modulate the size of the gorge are an important prerequisite. However, the molecular mechanism that governs such motions is not well explored. Here, to systematically investigate intrinsic motions of the enzyme, we performed microsecond molecular dynamics simulations on the monomer and dimer of Torpedo californica acetylcholinesterase (TcAChE) as well as the complex of TcAChE bound with the drug E2020. It has been revealed that protein-ligand interactions and dimerization both keep the gorge in bulk, and opening events of the gorge increase dramatically compared to the monomer. Dynamics of three subdomains, S3, S4 and the Ω-loop, are tightly associated with variations of the gorge size while the dynamics can be changed by ligand binding or protein dimerization. Moreover, high correlations among these subdomains provide a basis for remote residues allosterically modulating the gorge motions. These observations are propitious to expand our understanding of protein structure and function as well as providing clues for performing structure-based drug design.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The transcription factors of WRKY genes play essential roles in plant growth, stress responses, and metabolite biosynthesis. Erigeron breviscapus , a traditional Chinese herb, is abundant in ...flavonoids and has been used for centuries to treat cardiovascular and cerebrovascular diseases. However, the WRKY transcription factors that regulate flavonoid biosynthesis in E. breviscapus remain unknown. In this study, a total of 75 EbWRKY transcription factors were predicted through comprehensive genome-wide characterization of E. breviscapus and the chromosomal localization of each EbWRKY gene was investigated. RNA sequencing revealed transient responses of 74 predicted EbWRKY genes to exogenous abscisic acid (ABA), salicylic acid (SA), and gibberellin 3 (GA3) after 4 h of treatment. In contrast, the expression of key structural genes involved in flavonoid biosynthesis increased after 4 h in GA3 treatment. However, the content of flavonoid metabolites in leaves significantly increased at 12 h. The qRT-PCR results showed that the expression patterns of EbWRKY11 , EbWRKY30 , EbWRKY31 , EbWRKY36 , and EbWRKY44 transcription factors exhibited a high degree of similarity to the 11 structural genes involved in flavonoid biosynthesis. Protein-DNA interactions were performed between the key genes involved in scutellarin biosynthesis and candidate WRKYs . The result showed that F7GAT interacts with EbWRKY11, EbWRKY36, and EbWRKY44, while EbF6H has a self-activation function. This study provides comprehensive information on the regulatory control network of flavonoid accumulation mechanisms, offering valuable insights for breeding E. breviscapus varieties with enhanced scutellarin content.
Abstract
Cryo-electron microscopy (cryo-EM) enables the determination of membrane protein structures in native-like environments. Characterising how membrane proteins interact with the surrounding ...membrane lipid environment is assisted by resolution of lipid-like densities visible in cryo-EM maps. Nevertheless, establishing the molecular identity of putative lipid and/or detergent densities remains challenging. Here we present LipIDens, a pipeline for molecular dynamics (MD) simulation-assisted interpretation of lipid and lipid-like densities in cryo-EM structures. The pipeline integrates the implementation and analysis of multi-scale MD simulations for identification, ranking and refinement of lipid binding poses which superpose onto cryo-EM map densities. Thus, LipIDens enables direct integration of experimental and computational structural approaches to facilitate the interpretation of lipid-like cryo-EM densities and to reveal the molecular identities of protein-lipid interactions within a bilayer environment. We demonstrate this by application of our open-source LipIDens code to ten diverse membrane protein structures which exhibit lipid-like densities.
Erigeron breviscapus
is a Compositae plant, and its rich flavonoids have shown strong preventative and curative effects in the treatment of cardio- and cerebrovascular diseases.
bHLH
genes play a ...crucial role in plant growth and development. There are 116
EbbHLH
genes in
E. breviscapus
, and each gene has been named based on its chromosome location. Our phylogenetic analysis divided these genes into 18 subfamilies. To further investigate its function,
EbbHLH80
was isolated from
E. breviscapus
leaves. Next, transcriptomic and metabolomic analyses of tobacco leaves were performed. Among 421 differentially accumulated compounds, 98 flavonoids were identified. In addition, differentially expressed genes were identified using RNA-seq, and further analysis suggested that
EbbHLH80
-OE could not only regulate the expression of some structural genes in the flavonoid biosynthesis pathway to achieve flavonoid accumulation but also be involved in the regulation of a series of downstream pathways, such as stress response, ABA and ethylene signal transduction, to affect plant growth and development. The results of our analysis provide new insights into the function of
EbbHLH80
and lay the foundation for future functional studies on
E. breviscapus
.
Erigeron breviscapus
, a traditional Chinese medicinal plant, is enriched in flavonoids that are beneficial to human health. While we know that R2R3-MYB transcription factors (TFs) are crucial to ...flavonoid pathway, the transcriptional regulation of flavonoid biosynthesis in
E. breviscapus
has not been fully elucidated. Here,
EbMYBP1
, a R2R3-MYB transcription factor, was uncovered as a regulator involved in the regulation of flavonoid accumulation. Transcriptome and metabolome analysis revealed that a large group of genes related to flavonoid biosynthesis were significantly changed, accompanied by significantly increased concentrations of the flavonoid in
EbMYBP1-
OE transgenic tobacco compared with the wild-type (WT).
In vitro
and
in vivo
investigations showed that
EbMYBP1
participated in flavonoid biosynthesis, acting as a nucleus-localized transcriptional activator and activating the transcription of flavonoid-associated genes like
FLS
,
F3H
,
CHS
, and
CHI
by directly binding to their promoters. Collectively, these new findings are advancing our understanding of the transcriptional regulation that modulates the flavonoid biosynthesis.
contains an abundance of dammarane-type ginsenosides and gypenosides that exhibit extensive pharmacological activities. Increasing attention has been paid to the elucidation of cytochrome P450 ...monooxygenases (
s) and UDP-dependent glycosyltransferases (
s) that participate downstream of ginsenoside biosynthesis in the
genus. However, information on oxidosqualene cyclases (
), the upstream genes responsible for the biosynthesis of different skeletons of ginsenoside and gypenosides, is rarely reported. Here, an integrative study of the metabolome and the transcriptome in the leaf, stolon, and rattan was conducted and the function of
was demonstrated. In total, 46 triterpenes were detected and found to be highly abundant in the stolon, whereas gene expression analysis indicated that the upstream
genes responsible for saponin skeleton biosynthesis were highly expressed in the leaf. These findings indicated that the saponin skeletons were mainly biosynthesized in the leaf by
s, and subsequently transferred to the stolon via
s and
s biosynthesis to form various ginsenoside and gypenosides. Additionally, a new dammarane-II synthase (
),
, was identified by bioinformatics analysis, yeast expression assay, and enzyme assays. The results of the liquid chromatography-mass spectrometry (LC-MS) analysis proved that
could catalyze 2,3-oxidosqualene to form dammarenediol-II via cyclization. This work uncovered the biosynthetic mechanism of dammarenediol-II, an important starting substrate for ginsenoside and gypenosides biosynthesis, and may achieve the increased yield of valuable ginsenosides and gypenosides produced under excess substrate in a yeast cell factory through synthetic biology strategy.
Achlorophyllous plants are full mycoheterotrophic plants with no chlorophyll and they obtain their nutrients from soil fungi.
is a perennial, achlorophyllous orchid that displays distinctive ...evolutionary strategy of adaptation to the non-photosynthetic lifestyle. Here in this study, the genome of
was assembled to 1.12 Gb with a contig N50 size of 110 kb and a scaffold N50 size of 1.64 Mb so that it helped unveil the genetic basics of those adaptive changes. Based on the genomic data, key genes related to photosynthesis, leaf development, and plastid division pathways were found to be lost or under relaxed selection during the course of evolution. Thus, the genome sequence of
provides a good resource for future investigations of the evolution of orchids and other achlorophyllous plants.
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