Structural characterization of amorphous calcium carbonate-binding protein: an insight into the mechanism of amorphous calcium carbonate formation

2013 ◽  
Vol 453 (2) ◽  
pp. 179-186 ◽  
Author(s):  
Jingtan Su ◽  
Xiao Liang ◽  
Qiang Zhou ◽  
Guiyou Zhang ◽  
Hongzhong Wang ◽  
...  
Keyword(s):  
Calcium Carbonate ◽  
Binding Sites ◽  
Binding Protein ◽  
Homology Modelling ◽  
Size Exclusion ◽  
Structural Basis ◽  
Amorphous Calcium Carbonate ◽  
Carbonate Formation

ACC (amorphous calcium carbonate) plays an important role in biomineralization process for its function as a precursor for calcium carbonate biominerals. However, it is unclear how biomacromolecules regulate the formation of ACC precursor in vivo. In the present study, we used biochemical experiments coupled with bioinformatics approaches to explore the mechanisms of ACC formation controlled by ACCBP (ACC-binding protein). Size-exclusion chromatography, chemical cross-linking experiments and negative staining electron microscopy reveal that ACCBP is a decamer composed of two adjacent pentamers. Sequence analyses and fluorescence quenching results indicate that ACCBP contains two Ca2+-binding sites. The results of in vitro crystallization experiments suggest that one Ca2+-binding site is critical for ACC formation and the other site affects the ACC induction efficiency. Homology modelling demonstrates that the Ca2+-binding sites of pentameric ACCBP are arranged in a 5-fold symmetry, which is the structural basis for ACC formation. To the best of our knowledge, this is the first report on the structural basis for protein-induced ACC formation and it will significantly improve our understanding of the amorphous precursor pathway.

scholarly journals A yeast ARS-binding protein activates transcription synergistically in combination with other weak activating factors.

1990 ◽  
Vol 10 (3) ◽  
pp. 887-897 ◽  
Author(s):  
A R Buchman ◽  
R D Kornberg
Keyword(s):  
Dna Sequences ◽  
Transcriptional Activation ◽  
Binding Sites ◽  
Essential Gene ◽  
Specific Binding ◽  
Binding Protein ◽  
Point Mutations ◽  
Yeast Genes

ABFI (ARS-binding protein I) is a yeast protein that binds specific DNA sequences associated with several autonomously replicating sequences (ARSs). ABFI also binds sequences located in promoter regions of some yeast genes, including DED1, an essential gene of unknown function that is transcribed constitutively at a high level. ABFI was purified by specific binding to the DED1 upstream activating sequence (UAS) and was found to recognize related sequences at several other promoters, at an ARS (ARS1), and at a transcriptional silencer (HMR E). All ABFI-binding sites, regardless of origin, provided weak UAS function in vivo when examined in test plasmids. UAS function was abolished by point mutations that reduced ABFI binding in vitro. Analysis of the DED1 promoter showed that two ABFI-binding sites combine synergistically with an adjacent T-rich sequence to form a strong constitutive activator. The DED1 T-rich element acted synergistically with all other ABFI-binding sites and with binding sites for other multifunctional yeast activators. An examination of the properties of sequences surrounding ARS1 left open the possibility that ABFI enhances the initiation of DNA replication at ARS1 by transcriptional activation.

scholarly journals The role of phosphorylation and dephosphorylation of shell matrix proteins in shell formation: an in vivo and in vitro study

CrystEngComm ◽  
2018 ◽  
Vol 20 (27) ◽  
pp. 3905-3916 ◽  
Author(s):  
Jinzhe Du ◽  
Guangrui Xu ◽  
Chuang Liu ◽  
Rongqing Zhang
Keyword(s):  
Calcium Carbonate ◽  
In Vitro Study ◽  
Matrix Proteins ◽  
Shell Formation ◽  
Carbonate Formation ◽  
Shell Matrix ◽  
Shell Matrix Proteins

Phosphorylation of shell matrix proteins is critical for shell formation in vivo and can modulate calcium carbonate formation in vitro.

Potential Inhibition of COVID-19 RNA-dependent RNA Polymerase by Hepatitis C Virus Non-nucleoside Inhibitors: An In-silico Perspective

2020 ◽  
Vol 17 ◽  
Author(s):  
Yee Siew Choong ◽  
Theam Soon Lim ◽  
Hanyun Liu ◽  
Rubin Jiang ◽  
Zimu Cai ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Rna Polymerase ◽  
Binding Sites ◽  
Homology Modelling ◽  
Clinical Importance ◽  
Rna Dependent Rna Polymerase ◽  
Nucleoside Inhibitors

Background: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a novel member of the genus betacoronavirus in the Coronaviridae family. It has been identified as the causative agent of coronavirus disease 2019 (COVID-19) spreading rapidly in Asia, America and Europe. Like some other RNA viruses, RNA replication and transcription of SARS-CoV-2 relies on its RNA-dependent RNA polymerase (RdRP), which is a therapeutic target of clinical importance. Crystal structure of SARS-CoV-2 that was solved recently (PDB ID 6M71) with some missing residues. Objective: We used SARS-CoV-2 RdRP as a target protein to screen for possible chemical molecules with potential antiviral effects. Method: Here we modelled the missing residues 896-905 via homology modelling and then analysed the interactions of Hepatitis C virus allosteric non-nucleoside inhibitors (NNIs) in the reported NNIs binding sites in SARS-CoV-2 RdRP. Results and Discussion: We found that MK-3281, filibuvir, setrobuvir and dasabuvir might be able to inhibit SARS-CoV-2 RdRP based on their binding affinities in the respective binding sites. Conclusion: Further in vitro and in vivo experimental research will be carried out to evaluate their effectiveness in COVID19 treatment in the near future.

scholarly journals Structural basis and mechanism for metallochaperone-assisted assembly of the CuA center in cytochrome oxidase

2019 ◽  
Vol 5 (7) ◽  
pp. eaaw8478 ◽  
Author(s):  
Fabia Canonica ◽  
Daniel Klose ◽  
Raphael Ledermann ◽  
Maximilian M. Sauer ◽  
Helge K. Abicht ◽  
...  
Keyword(s):  
Binding Sites ◽  
Structural Data ◽  
Structural Basis ◽  
Copper Binding ◽  
Redox Center ◽  
Cytochrome Oxidases ◽  
Cu Ions ◽  
Copper Transfer

The mechanisms underlying the biogenesis of the structurally unique, binuclear Cu1.5+•Cu1.5+ redox center (CuA) on subunit II (CoxB) of cytochrome oxidases have been a long-standing mystery. Here, we reconstituted the CoxB•CuA center in vitro from apo-CoxB and the holo-forms of the copper transfer chaperones ScoI and PcuC. A previously unknown, highly stable ScoI•Cu2+•CoxB complex was shown to be rapidly formed as the first intermediate in the pathway. Moreover, our structural data revealed that PcuC has two copper-binding sites, one each for Cu1+ and Cu2+, and that only PcuC•Cu1+•Cu2+ can release CoxB•Cu2+ from the ScoI•Cu2+•CoxB complex. The CoxB•CuA center was then formed quantitatively by transfer of Cu1+ from a second equivalent of PcuC•Cu1+•Cu2+ to CoxB•Cu2+. This metalation pathway is consistent with all available in vivo data and identifies the sources of the Cu ions required for CuA center formation and the order of their delivery to CoxB.

A yeast ARS-binding protein activates transcription synergistically in combination with other weak activating factors

1990 ◽  
Vol 10 (3) ◽  
pp. 887-897
Author(s):  
A R Buchman ◽  
R D Kornberg
Keyword(s):  
Dna Sequences ◽  
Transcriptional Activation ◽  
Binding Sites ◽  
Essential Gene ◽  
Specific Binding ◽  
Binding Protein ◽  
Point Mutations ◽  
Yeast Genes

ABFI (ARS-binding protein I) is a yeast protein that binds specific DNA sequences associated with several autonomously replicating sequences (ARSs). ABFI also binds sequences located in promoter regions of some yeast genes, including DED1, an essential gene of unknown function that is transcribed constitutively at a high level. ABFI was purified by specific binding to the DED1 upstream activating sequence (UAS) and was found to recognize related sequences at several other promoters, at an ARS (ARS1), and at a transcriptional silencer (HMR E). All ABFI-binding sites, regardless of origin, provided weak UAS function in vivo when examined in test plasmids. UAS function was abolished by point mutations that reduced ABFI binding in vitro. Analysis of the DED1 promoter showed that two ABFI-binding sites combine synergistically with an adjacent T-rich sequence to form a strong constitutive activator. The DED1 T-rich element acted synergistically with all other ABFI-binding sites and with binding sites for other multifunctional yeast activators. An examination of the properties of sequences surrounding ARS1 left open the possibility that ABFI enhances the initiation of DNA replication at ARS1 by transcriptional activation.

scholarly journals Highly Hydrated Paramagnetic Amorphous Calcium Carbonate Nanoclusters as a Superior MRI Contrast Agent

2021 ◽  
Author(s):  
Liang Dong ◽  
Yun-Jun Xu ◽  
Cong Sui ◽  
Yang Zhao ◽  
Li-Bo Mao ◽  
...  
Keyword(s):  
Calcium Carbonate ◽  
Contrast Agents ◽  
Water Content ◽  
Mutual Effect ◽  
Mri Contrast Agent ◽  
High Water Content ◽  
Amorphous Calcium Carbonate ◽  
Mri Contrast ◽  
Carbonate Formation

Abstract Amorphous calcium carbonate (ACC) plays a key role as transient precursor in the early stages of biogenic calcium carbonate formation in nature. However, due to its instability in aqueous solution, there is still rare success to utilize ACC in biomedicine. Here, we report the mutual effect between paramagnetic gadolinium ions and ACC, resulting in ultrafine paramagnetic amorphous carbonate nanoclusters (ACNC) in the presence of both gadolinium occluded highly hydrated ACC-like environment and poly(acrylic acid). Gadolinium is confirmed to enhance the water content in ACC, and the high water content of ACNC (23 molecules H2O per 1 Gd) contributes to the much enhanced magnetic resonance imaging (MRI) contrast efficiency compared with commercially available gadolinium-based contrast agents. Furthermore, the enhanced T1 weighted MRI performance and biocompatibility of ACNC are further evaluated in various animals including rat, rabbit and beagle dog, in combination with promising safety in vivo. Overall, exceptionally facile mass-productive ACNC exhibits superb imaging performance and impressive stability, which provides a promising strategy to design MR contrast agents.

scholarly journals Docosahexaenoic Acid Suppresses Expression of Adipogenic Tetranectin through Sterol Regulatory Element-Binding Protein and Forkhead Box O Protein in Pigs

Nutrients ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 2315
Author(s):  
Jui-Ting Yang ◽  
Yu-Jen Chen ◽  
Chao-Wei Huang ◽  
Ya-Chin Wang ◽  
Harry J. Mersmann ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Binding Sites ◽  
Binding Protein ◽  
Regulatory Element ◽  
Forkhead Box ◽  
Transcriptional Suppression ◽  
Element Binding Protein ◽  
Sterol Regulatory Element

Tetranectin (TN), a plasminogen-binding protein originally involved in fibrinolysis and bone formation, was later identified as a secreted adipokine from human and rat adipocytes and positively correlated with adipogenesis and lipid metabolism in adipocytes. To elucidate the nutritional regulation of adipogenic TN from diets containing different sources of fatty acids (saturated, n-6, n-3) in adipocytes, we cloned the coding region of porcine TN from a cDNA library and analyzed tissue expressions in weaned piglets fed with 2% soybean oil (SB, enriched in n-6 fatty acids), docosahexaenoic acid oil (DHA, an n-3 fatty acid) or beef tallow (BT, enriched in saturated and n-9 fatty acids) for 30 d. Compared with tissues in the BT- or SB-fed group, expression of TN was reduced in the adipose, liver and lung tissues from the DHA-fed group, accompanied with lowered plasma levels of triglycerides and cholesterols. This in vivo reduction was also confirmed in porcine primary differentiated adipocytes supplemented with DHA in vitro. Then, promoter analysis was performed. A 1956-bp putative porcine TN promoter was cloned and transcription binding sites for sterol regulatory-element binding protein (SREBP)-1c or forkhead box O proteins (FoxO) were predicted on the TN promoter. Mutating binding sites on porcine TN promoters showed that transcriptional suppression of TN by DHA on promoter activity was dependent on specific response elements for SREBP-1c or FoxO. The inhibited luciferase promoter activity by DHA on the TN promoter coincides with reduced gene expression of TN, SREBP-1c, and FoxO1 in human embryonic kidney HEK293T cells supplemented with DHA. To conclude, our current study demonstrated that the adipogenic TN was negatively regulated by nutritional modulation of DHA both in pigs in vivo and in humans/pigs in vitro. The transcriptional suppression by DHA on TN expression was partly through SREBP-1c or FoxO. Therefore, down-regulation of adipogenic tetranectin associated with fibrinolysis and adipogenesis may contribute to the beneficial effects of DHA on ameliorating obesity-induced metabolic syndromes such as atherosclerosis and adipose dysfunctions.

CHARACTERIZATION OF A NEW LCAT MUTATION CAUSING FAMILIAL LCAT DEFICIENCY (FLD) AND THE ROLE OF APOE AS A MODIFIER GENE OF THE FLD PHENOTYPE

2009 ◽  
Vol 32 (6S) ◽  
pp. 3
Author(s):  
A Baass ◽  
H Wassef ◽  
M Tremblay ◽  
L Bernier ◽  
R Dufour ◽  
...  
Keyword(s):  
Modifier Gene ◽  
Size Exclusion ◽  
Plasma Lipoproteins ◽  
Lipoprotein Profile ◽  
Exclusion Chromatography ◽  
Low Hdl ◽  
Lcat Deficiency ◽  
Apoe Genotypes

Introduction: LCAT (lecithin:cholesterol acyltransferase ) is an enzyme which plays an essential role in cholesterol esterification and reverse cholesterol transport. Familial LCAT deficiency (FLD) is a disease characterized by a defect in LCAT resulting in extremely low HDL-C, premature corneal opacities, anemia as well as proteinuria and renal failure. Method: We have identified two brothers presenting characteristics of familial LCAT deficiency. We sequenced the LCAT gene, measured the lipid profile as well as the LCAT activity in 15 members of this kindred. We also characterized the plasma lipoproteins by agarose gel electrophoresis and size exclusion chromatography and sequenced several candidate genes related to dysbetalipoproteinemia in this family. Results: We have identified the first French Canadian kindred with familial LCAT deficiency. Two brothers affected by FLD, were homozygous for a novel LCAT mutation. This c.102delG mutation occurs at the codon for His35 causing a frameshift that stops transcription at codon 61 abolishing LCAT enzymatic activity both in vivo and in vitro. It has a dramatic effect on the lipoprotein profile, with an important reduction of HDL-C in both heterozygotes (22%) and homozygotes (88%) and a significant decrease in LDL-C in heterozygotes (35%) as well as homozygotes (58%). Furthermore, the lipoprotein profile differed markedly between the two affected brothers who had different APOE genotypes. We propose that APOE could be an important modifier gene explaining heterogeneity in lipoprotein profiles observed among FLD patients. Our results suggest that a LCAT-/- genotype associated with an APOE ?2 allele could be a novel mechanism leading to dysbetalipoproteinemia.

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