scholarly journals Thermodynamics of Ligand Binding to a Heterogeneous RNA Population in the Malachite Green Aptamer

Biochemistry ◽  
2011 ◽  
Vol 51 (1) ◽  
pp. 565-572 ◽  
Author(s):  
Joshua E. Sokoloski ◽  
Sarah E. Dombrowski ◽  
Philip C. Bevilacqua
Keyword(s):  
Ligand Binding ◽  
Malachite Green ◽  
Malachite Green Aptamer

2.8 Å crystal structure of the malachite green aptamer 1 1Edited by J. A. Doudna

2000 ◽  
Vol 301 (1) ◽  
pp. 117-128 ◽  
Author(s):  
Christopher Baugh ◽  
Dilârâ Grate ◽  
Charles Wilson
Keyword(s):  
Crystal Structure ◽  
Malachite Green ◽  
Malachite Green Aptamer

scholarly journals Labeling RNAs in live cells using malachite green aptamer scaffolds as fluorescent probes

2017 ◽  
Author(s):  
V. Siddartha Yerramilli ◽  
Kyung Hyuk Kim
Keyword(s):  
Malachite Green ◽  
Tandem Repeats ◽  
Fluorescent Proteins ◽  
Rna Aptamers ◽  
Live Cells ◽  
High Background ◽  
Rna Molecules ◽  
High Background Noise ◽  
Fluorescent Tags ◽  
Malachite Green Aptamer

AbstractRNAs mediate many different processes that are central to cellular function. The ability to quantify or image RNAs in live cells is very useful in elucidating such functions of RNA. RNA aptamerfluorogen systems have been increasingly used in labeling RNAs in live cells. Here, we use the malachite green aptamer (MGA), an RNA aptamer that can specifically bind to malachite green (MG) dye and induces it to emit far-red fluorescence signals. Previous studies on MGA showed a potential for the use of MGA for genetically tagging other RNA molecules in live cells. However, these studies also exhibited low fluorescence signals and high background noise. Here we constructed and tested RNA scaffolds containing multiple tandem repeats of MGA as a strategy to increase the brightness of the MGA aptamer-fluorogen system as well as to make the system fluoresce when tagging various RNA molecules, in live cells. We demonstrate that our MGA scaffolds can induce fluorescence signals by up to ~20 fold compared to the basal level as a genetic tag for other RNA molecules. We also show that our scaffolds function reliably as genetically-encoded fluorescent tags for mRNAs of fluorescent proteins and other RNA aptamers.

CarR, a MarR-family regulator from Corynebacterium glutamicum, modulated antibiotic and aromatic compound resistance

2019 ◽  
Vol 476 (21) ◽  
pp. 3141-3159 ◽  
Author(s):  
Meiru Si ◽  
Can Chen ◽  
Zengfan Wei ◽  
Zhijin Gong ◽  
GuiZhi Li ◽  
...  
Keyword(s):  
Stress Response ◽  
Ligand Binding ◽  
Corynebacterium Glutamicum ◽  
Conformational Changes ◽  
Cysteine Oxidation ◽  
Transcriptional Regulatory ◽  
Ligand Free ◽  
Redox Sensing

Abstract MarR (multiple antibiotic resistance regulator) proteins are a family of transcriptional regulators that is prevalent in Corynebacterium glutamicum. Understanding the physiological and biochemical function of MarR homologs in C. glutamicum has focused on cysteine oxidation-based redox-sensing and substrate metabolism-involving regulators. In this study, we characterized the stress-related ligand-binding functions of the C. glutamicum MarR-type regulator CarR (C. glutamicum antibiotic-responding regulator). We demonstrate that CarR negatively regulates the expression of the carR (ncgl2886)–uspA (ncgl2887) operon and the adjacent, oppositely oriented gene ncgl2885, encoding the hypothetical deacylase DecE. We also show that CarR directly activates transcription of the ncgl2882–ncgl2884 operon, encoding the peptidoglycan synthesis operon (PSO) located upstream of carR in the opposite orientation. The addition of stress-associated ligands such as penicillin and streptomycin induced carR, uspA, decE, and PSO expression in vivo, as well as attenuated binding of CarR to operator DNA in vitro. Importantly, stress response-induced up-regulation of carR, uspA, and PSO gene expression correlated with cell resistance to β-lactam antibiotics and aromatic compounds. Six highly conserved residues in CarR were found to strongly influence its ligand binding and transcriptional regulatory properties. Collectively, the results indicate that the ligand binding of CarR induces its dissociation from the carR–uspA promoter to derepress carR and uspA transcription. Ligand-free CarR also activates PSO expression, which in turn contributes to C. glutamicum stress resistance. The outcomes indicate that the stress response mechanism of CarR in C. glutamicum occurs via ligand-induced conformational changes to the protein, not via cysteine oxidation-based thiol modifications.

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