NMR chemical shift assignment of a constitutively active fragment of the antitermination protein LicT

International audienceLicT belongs to an essential family of bacterial antitermination proteins which bind to nascent mRNAs in order to stimulate transcription of sugar-metabolizing operons. As most of other antitermination proteins involved in carbohydrate metabolism, LicT is composed of a N-terminal RNA-binding module (CAT) and two homologous regulatory modules (PRD1 and PRD2). The activity of the CAT effector module is controlled by antagonist phosphorylations by the phosphotransferase system on conserved histidines of the two C-terminal PRDs in response to available carbon sources. Previous studies on truncated and mutant constructs have provided partial structural insight into the mechanism of signal transduction between the N-terminal RNA-binding domain and the two regulation modules. However, no structure at atomic resolution has been ever solved that contain the RNA-binding domain and a regulation module. We report the NMR assignment of a constitutively active fragment of LicT, named D99A-CAT-PRD1 or CAT-PRD1*. This fragment is composed of the RNA-binding module and the first N-terminal regulation module which bears the mutation of Asp99 to an asparagine. It is dimeric as the native protein, with a 40 kD molecular weight. The D99N mutation is sufficient to endow this fragment with a high RNA-binding constitutive activity, in a phosphorylation-free context. The assignment reported here should set the base of future NMR investigation of signal transduction between the regulatory module and the effector module in the active state of the protein, and in the long term enable the structural study of the full length protein structure in interaction with its target RNA

NMR chemical shift backbone assignment of the viral protein P1 encoded by the African Rice Yellow Mottle Virus

International audienceRNA silencing describes a pan-eukaryotic pathway of gene regulation where doubled stranded RNA are processed by the RNAse III enzyme Dicer or homologs. In particular, plants use it as a way to defend themselves against pathogen invasions. In turn, to evade the plant immune response, viruses have developed anti-RNA silencing mechanisms. They may indeed code for proteins called "viral suppressor of RNA silencing" which block the degrading of viral genomic or messenger RNA by the plant. The Rice Mottle Virus is an African virus of the sobemovirus family, which attacks the most productive rice varieties cultivated on this continent. It encodes P1, a cysteine-rich protein described as a potential RNA silencing suppressor. P1 is a 157 amino-acid long protein, characterized by a high propensity to aggregate concomitant with a limited stability with time in the conditions used in structural studies. To overcome this problem, shorter fragments were also studied. This strategy enabled the assignment of more than 90% backbone resonances of P1. This assignment should set the base of future NMR investigation of the protein structure and of its interactions with rice cellular partners

QTL identification and candidate gene identification for monoterpene content in grape (Vitis vinifera L.) berries

Great progress has been made during the last decade in clarifying the molecular details of aroma accumulation in grape berries. However, the multigene complex controlling monoterpene accumulation in grape is not well understood. To shed light on this issue, the grapes of 149 F1 progenies from the cross 87-1 (Vitis vinifera L.) × 9-22 (Vitis vinifera L.) were characterized at the mature stage for three representative free monoterpenes during five growing seasons. A total of 202, 184 and 255 polymorphic SSR (simple sequence repeat) markers were contracted on the maternal 87-1, paternal 9-22 and consensus genetic maps, respectively. On the consensus map, we confirmed a major QTL (quantitative trait locus) for free linalool, nerol and α-terpineol content on linkage group (LG) 5, and a stable QTL for free linalool and α-terpineol was detected on LG 10. In addition, two new stable QTLs for free monoterpene (linalool, nerol and α-terpineol) contents were identified on LG 11 and LG 18 that explained up to 42.5 % of the total variance. Eleven promising candidate genes related to pentatricopeptide repeat (PPR)-containing proteins, seed maturation protein, RING finger protein, and AP2/ERF transcription factors might be potentially involved in monoterpene accumulation. The stable QTLs and candidate genes identified in this study provide new insights into free monoterpene accumulation in grape

Gut microbiota and calcium balance

Microorganisms living on the surface and inside the human body play an important role in the physiological activities of the human body. The largest microecosystem in the human body is the gut microbiome. Calcium disorders are found in many diseases. For example, patients with chronic renal insufficiency present with secondary hyperparathyroidism, which is caused by a calcium imbalance in the body. In addition, calcium dysregulation may affect lipid metabolism in the liver through the calmodulator pathway, leading to cirrhosis, etc. Currently, a considerable number of probiotics have been proven to enhance the body’s absorption of calcium. This paper reviews the effects of intestinal flora and related factors such as short-chain fatty acids, estrogen, immune factors and vitamin D on calcium balance

Non-Stationarity of Aerosol Extinction Coefficient per Unit of Mass in Autumn and Winter in Chengdu, China

Based on hourly observation data from the aethalometer and GRIMM180 environment particle monitor as well as the simultaneous data of visibility (V), relative humidity (RH) and nitrogen dioxide (NO2) from October to December in 2017 in Chengdu, the corresponding time series of aerosol extinction coefficient per unit of mass is retrieved. The generalized additive models (GAMs) are adopted to analyze the non-stationarity of the time series of aerosol extinction coefficient per unit of mass and to explore the responses of the aerosol extinction coefficient per unit of mass to the aerosol component structure factors (ρBC/ρPM10, ρPM1/ρPM2.5, ρPM1~2.5/ρPM2.5 and ρPM2.5/ρPM10; ρ represents particle mass concentration) and RH. The results show that through the comparative analysis of stationary and non-stationary models, the time series of aerosol extinction coefficient per unit of mass in autumn and winter in Chengdu is non-stationary. In addition, the RH and aerosol component structure factors are all significant nonlinear covariates that affect the non-stationarity of the aerosol extinction coefficient per unit of mass. According to the influence of covariates, the sequence is as follows: RH > ρBC/ρPM10 > ρPM2.5/ρPM10 > ρPM1/ρPM2.5. At PM2.5 pollution concentration (ρPM2.5 > 75 μg m−3), according to the influence of covariates, the sequence is as follows: RH > ρPM1~2.5/ρPM2.5 > ρBC/ρPM10 > ρPM2.5/ρPM10. Moreover, the interaction between RH and aerosol component structure factors significantly affects the aerosol extinction coefficient per unit of mass. The condition of high RH, high ρPM2.5/ρPM10, high ρPM1/ρPM2.5 and low ρBC/ρPM10 has a synergistic amplification effect on the increase of the aerosol extinction coefficient per unit of mass. At PM2.5 pollution concentration, the synergistic effect of high RH, high ρPM2.5/ρPM10, high ρPM1~2.5/ρPM2.5 and low ρBC/ρPM10 is beneficial to the increase of the aerosol extinction coefficient per unit of mass

Resolving the activation mechanism of the D99N antiterminator LicT protein

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1H, 13C and 15N backbone chemical shift assignments of camelid single-domain antibodies against active state µ-opioid receptor

International audienceNanobodies are single chain antibodies that have become a highly valuable and versatile tool for biomolecular and therapeutic research. One application field is the stabilization of active states of flexible proteins, among which G-protein coupled receptors (GPCRs) represent a very important class of membrane proteins. Here we present the backbone and side-chain assignment of the 1H, 13C and 15N resonances of Nb33 and Nb39, two nanobodies that recognize and stabilize the μ-opioid receptor (μOR) to opioids in its active agonist-bound conformation. In addition, we present a comparison of their secondary structures as derived from NMR chemical shifts

Structural characterization of the mammalian deoxynucleotide N-hydrolase Rcl and its stabilizing interactions with two inhibitors.

The gene Rcl encodes a deoxynucleoside 5'-monophosphate N-glycosidase that catalyzes the hydrolysis of the N-glycosidic bond of the nucleotide to give deoxyribose 5-phosphate and a nucleobase, preferentially a purine. This enzyme is over-expressed in several cancers, and its rate of expression is correlated to the degree of aggressiveness of tumors, which makes it a new and attractive therapeutic target. We describe here its structural characterization in the presence of two inhibitory substrate mimics. One of these ligands corresponds to the monophosphorylated form of acyclovir, which is used in the clinic. This study reveals an important ligand-induced stabilization of the dimer structure of the enzyme. The original structural features of Rcl will be helpful for designing new inhibitors

Epithelial Splicing Regulatory Protein 1 (ESRP1) is a new regulator of stomach smooth muscle development and plasticity

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