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Udo Heinemann - One of the best experts on this subject based on the ideXlab platform.
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rna single strands bind to a conserved surface of the major Cold Shock protein in crystals and solution
RNA, 2012Co-Authors: Rolf Sachs, Udo Heinemann, Klaas E A Max, Jochen BalbachAbstract:Bacterial Cold Shock Proteins (CSPs) regulate the cellular response to temperature downshift. Their general principle of function involves RNA chaperoning and transcriptional antitermination. Here we present two crystal structures of Cold Shock protein B from Bacillus subtilis (Bs-CspB) in complex with either a hexanucleotide (5′-UUUUUU-3′) or heptanucleotide (5′-GUCUUUA-3′) single-stranded RNA (ssRNA). Hydrogen bonds and stacking interactions between RNA bases and aromatic sidechains characterize individual binding subsites. Additional binding subsites which are not occupied by the ligand in the crystal structure were revealed by NMR spectroscopy in solution on Bs-CspB·RNA complexes. Binding studies demonstrate that Bs-CspB associates with ssDNA as well as ssRNA with moderate sequence specificity. Varying affinities of oligonucleotides are reflected mainly in changes of the dissociation rates. The generally lower binding affinity of ssRNA compared to its ssDNA analog is attributed solely to the substitution of thymine by uracil bases in RNA.
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common mode of dna binding to Cold Shock domains crystal structure of hexathymidine bound to the domain swapped form of a major Cold Shock protein from bacillus caldolyticus
FEBS Journal, 2007Co-Authors: Klaas E A Max, Jochen Balbach, Markus Zeeb, Ralf Bienert, Udo HeinemannAbstract:Bacterial Cold Shock Proteins (CSPs) regulate cellular adaptation to Cold stress. Functions ascribed to CSP include roles as RNA chaperones and in transcription antitermination. We present the crystal structure of the Bacillus caldolyticus CSP (Bc-Csp) in complex with hexathymidine (dT6) at a resolution of 1.29 A. Bound to dT6, crystalline Bc-Csp forms a domain-swapped dimer in which β strands 1–3 associate with strands 4 and 5 from the other subunit to form a closed β barrel and vice versa. The globular units of dimeric Bc-Csp closely resemble the well-known structure of monomeric CSP. Structural reorganization from the monomer to the domain-swapped dimer involves a strictly localized change in the peptide bond linking Glu36 and Gly37 of Bc-Csp. Similar structural reorganizations have not been found in any other CSP or oligonucleotide/oligosaccharide-binding fold structures. Each dT6 ligand is bound to one globular unit of Bc-Csp via an amphipathic protein surface. Individual binding subsites interact with the DNA bases through stacking and hydrogen bonding. The sugar–phosphate backbone remains solvent exposed. Based on crystallographic and biochemical studies of deoxyoligonucleotide binding to CSP, we suggest a common mode of binding of single-stranded heptanucleotide motifs to Proteins containing Cold Shock domains, including the eukaryotic Y-box factors.
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t rich dna single strands bind to a preformed site on the bacterial Cold Shock protein bs cspb
Journal of Molecular Biology, 2006Co-Authors: Klaas E A Max, Jochen Balbach, Markus Zeeb, Ralf Bienert, Udo HeinemannAbstract:Bacterial Cold Shock Proteins (CSPs) are involved in cellular adaptation to Cold stress. They bind to single-stranded nucleic acids with a KD value in the micro- to nanomolar range. Here we present the structure of the Bacillus subtilis CspB (Bs-CspB) in complex with hexathymidine (dT6) at a resolution of 1.78 A. Bs-CspB binds to dT6 with nanomolar affinity via an amphipathic interface on the protein surface. Individual binding subsites interact with single nucleobases through stacking interactions and hydrogen bonding. The sugar-phosphate backbone and the methyl groups of the thymine nucleobases remain solvent exposed and are not contacted by protein groups. Fluorescence titration experiments monitoring the binding of oligopyrimidines to Bs-CspB reveal binding preferences at individual subsites and allow the design of an optimised heptapyrimidine ligand, which is bound with sub-nanomolar affinity. This study reveals the stoichiometry and sequence determinants of the binding of single-stranded nucleic acids to a preformed site on Bs-CspB and thus provides the structural basis of the RNA chaperone and transcription antitermination activities of the CSP.
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two exposed amino acid residues confer thermostability on a Cold Shock protein
Nature Structural & Molecular Biology, 2000Co-Authors: Dieter Perl, Udo Heinemann, Uwe Mueller, Franz X SchmidAbstract:Thermophilic organisms produce Proteins of exceptional stability. To understand protein thermostability at the molecular level we studied a pair of Cold Shock Proteins, one of mesophilic and one of thermophilic origin, by systematic mutagenesis. Although the two Proteins differ in sequence at 12 positions, two surface-exposed residues are responsible for the increase in stability of the thermophilic protein (by 15.8 kJ mol−1 at 70 °C). 11.5 kJ mol−1 originate from a predominantly electrostatic contribution of Arg 3 and 5.2 kJ mol−1 from hydrophobic interactions of Leu 66 at the carboxy terminus. The mesophilic protein could be converted to a highly thermostable form by changing the Glu residues at positions 3 and 66 to Arg and Leu, respectively. The variation of surface residues may thus provide a simple and powerful approach for increasing the thermostability of a protein.
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thermal stability and atomic resolution crystal structure of the bacillus caldolyticus Cold Shock protein
Journal of Molecular Biology, 2000Co-Authors: U Mueller, Franz X Schmid, Dieter Perl, Udo HeinemannAbstract:The bacterial Cold Shock Proteins are small compact beta-barrel Proteins without disulfide bonds, cis-proline residues or tightly bound cofactors. Bc-Csp, the Cold Shock protein from the thermophile Bacillus caldolyticus shows a twofold increase in the free energy of stabilization relative to its homolog Bs-CspB from the mesophile Bacillus subtilis, although the two Proteins differ by only 12 out of 67 amino acid residues. This pair of Cold Shock Proteins thus represents a good system to study the atomic determinants of protein thermostability. Bs-CspB and Bc-Csp both unfold reversibly in cooperative transitions with T(M) values of 49.0 degrees C and 77.3 degrees C, respectively, at pH 7.0. Addition of 0.5 M salt stabilizes Bs-CspB but destabilizes Bc-Csp. To understand these differences at the structural level, the crystal structure of Bc-Csp was determined at 1.17 A resolution and refined to R=12.5% (R(free)=17.9%). The molecular structures of Bc-Csp and Bs-CspB are virtually identical in the central beta-sheet and in the binding region for nucleic acids. Significant differences are found in the distribution of surface charges including a sodium ion binding site present in Bc-Csp, which was not observed in the crystal structure of the Bs-CspB. Electrostatic interactions are overall favorable for Bc-Csp, but unfavorable for Bs-CspB. They provide the major source for the increased thermostability of Bc-Csp. This can be explained based on the atomic-resolution crystal structure of Bc-Csp. It identifies a number of potentially stabilizing ionic interactions including a cation-binding site and reveals significant changes in the electrostatic surface potential.
Masayori Inouye - One of the best experts on this subject based on the ideXlab platform.
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Escherichia coli RNase R has dual activities, helicase and RNase
2016Co-Authors: Naoki Awano, Masayori Inouye, Vaishnavi Rajagopal, Mark Arbing, Smita Patel, John Hunt, Sangita PhadtareAbstract:In Escherichia coli, the Cold Shock response occurs when there is a temperature downshift from 37°C to 15°C, and this response is characterized by induction of several Cold Shock Proteins, including the DEAD-box helicase CsdA, during the acclimation phase. CsdA is involved in a variety of cellular processes. Our previous studies showed that the helicase activity of CsdA is critical for its function in Cold Shock acclimation of cells and that the only Proteins that were able to complement its function were another helicase, RhlE, an RNA chaperone, CspA, and a Cold-inducible exoribonuclease, RNase R. Interestingly, other major 3-to-5 process-ing exoribonucleases of E. coli, such as polynucleotide phosphorylase and RNase II, cannot complement the Cold Shock function of CsdA. Here we carried out a domain analysis of RNase R and showed that this protein has two distinct activities, RNase and helicase, which are independent of each other and are due to different domains. Mutant RNase R Proteins that lack the RNase activity but exhibit the helicase activity were able to complement the Cold Shock function of CsdA, suggesting that only the helicase activity of RNase R is essential for complementation of the Cold Shock function of CsdA. We also observed that in vivo deletion of the two Cold Shock domains resulted in a loss of the ability of RNase R to complement the Cold Shock function of CsdA. W
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The Cold Shock Response.
EcoSal Plus, 2008Co-Authors: Sangita Phadtare, Masayori InouyeAbstract:This review focuses on the Cold Shock response of Escherichia coli. Change in temperature is one of the most common stresses that an organism encounters in nature. Temperature downshift affects the cell on various levels: (i) decrease in the membrane fluidity; (ii) stabilization of the secondary structures of RNA and DNA; (iii) slow or inefficient protein folding; (iv) reduced ribosome function, affecting translation of non-Cold Shock Proteins; (v) increased negative supercoiling of DNA; and (vi) accumulation of various sugars. Cold Shock Proteins and certain sugars play a key role in dealing with the initial detrimental effect of Cold Shock and maintaining the continued growth of the organism at low temperature. CspA is the major Cold Shock protein of E. coli, and its homologues are found to be widespread among bacteria, including psychrophilic, psychrotrophic, mesophilic, and thermophilic bacteria, but are not found in archaea or cyanobacteria. Significant, albeit transient, stabilization of the cspA mRNA immediately following temperature downshift is mainly responsible for its Cold Shock induction. Various approaches were used in studies to detect Cold Shock induction of cspA mRNA. Sugars are shown to confer protection to cells undergoing Cold Shock. The study of the Cold Shock response has implications in basic and health-related research as well as in commercial applications. The Cold Shock response is elicited by all types of bacteria and affects these bacteria at various levels, such as cell membrane, transcription, translation, and metabolism.
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three amino acids in escherichia coli cspe surface exposed aromatic patch are critical for nucleic acid melting activity leading to transcription antitermination and Cold acclimation of cells
Journal of Biological Chemistry, 2002Co-Authors: Sangita Phadtare, Sanjay Tyagi, Masayori Inouye, Konstantin SeverinovAbstract:Abstract Cold-Shock Proteins of the CspA family ofEscherichia coli help the cells to acclimate to low temperature conditions through an unknown mechanism. In vitro, these Proteins bind to single-stranded nucleic acids and destabilize nucleic acid secondary structures. An unusual surface-exposed patch of 6 evolutionarily conserved aromatic amino acids is thought to be involved in RNA binding by the Cold-Shock Proteins. Here we investigated the functional role of the aromatic patch in E. coli CspE by substituting individual aromatic residues with positively charged Arg residues. These substitutions do not affect the RNA binding activity of the CspE mutants. We show that substitutions of three centrally located aromatic patch amino acid residues, Phe17, Phe30, and His32, abolish the ability of the mutant CspE to acclimatize cells to Cold, antiterminate transcription and melt nucleic acids but have no effect on RNA binding. On the other hand, peripherally located Trp10, Phe19, and Phe33 can be substituted with Arg without loss of any of the in vivo and in vitro CspE functions tested. The results thus indicate that these aromatic patch residues have clearly distinct functional roles and further extend the correlation between the essential function of CspA homologues in Cold acclimation and their ability to antiterminate transcription.
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selective mrna degradation by polynucleotide phosphorylase in Cold Shock adaptation in escherichia coli
Journal of Bacteriology, 2001Co-Authors: Kunitoshi Yamanaka, Masayori InouyeAbstract:Upon Cold Shock, Escherichia coli cell growth transiently stops. During this acclimation phase, specific Cold Shock Proteins (CSPs) are highly induced. At the end of the acclimation phase, their synthesis is reduced to new basal levels, while the non-Cold Shock protein synthesis is resumed, resulting in cell growth reinitiation. Here, we report that polynucleotide phosphorylase (PNPase) is required to repress CSP production at the end of the acclimation phase. A pnp mutant, upon Cold Shock, maintained a high level of CSPs even after 24 h. PNPase was found to be essential for selective degradation of CSP mRNAs at 15 degrees C. In a poly(A) polymerase mutant and a CsdA RNA helicase mutant, CSP expression upon Cold Shock was significantly prolonged, indicating that PNPase in concert with poly(A) polymerase and CsdA RNA helicase plays a critical role in Cold Shock adaptation.
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acquirement of Cold sensitivity by quadruple deletion of the cspa family and its suppression by pnpase s1 domain in escherichia coli
Molecular Microbiology, 2001Co-Authors: Bing Xia, Masayori InouyeAbstract:Escherichia coli contains a large CspA family, CspA to CspI. Here, we demonstrate that E. coli is highly protected against Cold-Shock stress, as these CspA homologues existed at approximately a total of two million molecules per cell at low temperature and growth defect was not observed until four csp genes (cspA, cspB, cspE and cspG) were deleted. The quadruple-deletion strain acquired Cold sensitivity and formed filamentous cells at 15°C although chromosomes were normally segregated. The Cold-sensitivity and filamentation phenotypes were suppressed by all members of the CspA family except for CspD, which causes lethality upon overexpression. Interestingly, the Cold sensitivity of the mutant was also suppressed by the S1 domain of polynucleotide phosphorylase (PNPase), which also folds into a β-barrel structure similar to that of CspA. The present results show that Cold-Shock Proteins and S1 domains share not only the tertiary structural similarity but also common functional properties, suggesting that these seemingly distinct protein categories may have evolved from a common primordial RNA-binding protein.
Ryozo Imai - One of the best experts on this subject based on the ideXlab platform.
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arabidopsis Cold Shock domain protein 2 is a negative regulator of Cold acclimation
New Phytologist, 2013Co-Authors: Kentaro Sasaki, Ryozo Imai, Myunghee KimAbstract:Bacterial Cold Shock Proteins (CSPs) act as RNA chaperones that destabilize mRNA secondary structures at low temperatures. Bacterial CSPs are composed solely of a nucleic acid-binding domain termed the Cold Shock domain (CSD). Plant CSD Proteins contain an auxiliary domain in addition to the CSD but also show RNA chaperone activity. However, their biological functions are poorly understood. We examined Arabidopsis Cold Shock DOMAIN PROTEIN 2 (AtCSP2) using overexpressing and mutant lines. A double mutant, with reduced AtCSP2 and no AtCSP4, showed higher freezing tolerance than the wild-type when Cold-acclimated. The increase in freezing tolerance was associated with up-regulation of CBF transcription factors and their downstream genes. By contrast, overexpression of AtCSP2 resulted in decreased freezing tolerance when Cold-acclimated. In addition, late flowering and shorter siliques were observed in the overexpressing lines. AtCSP2 negatively regulates freezing tolerance and is partially redundant with its closest paralog, AtCSP4.
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erratum to interactome analysis reveals versatile functions of arabidopsis Cold Shock domain protein 3 in rna processing within the nucleus and cytoplasm
Cell Stress & Chaperones, 2013Co-Authors: Myunghee Kim, Kentaro Sasaki, Yutaka Sonoda, Hironori Kaminaka, Ryozo ImaiAbstract:Arabidopsis Cold Shock DOMAIN PROTEIN 3 (AtCSP3) shares an RNA chaperone function with E. coli Cold Shock Proteins and regulates freezing tolerance during Cold acclimation. Here, we screened for AtCSP3-interacting Proteins using a yeast two-hybrid system and 38 candidate interactors were identified. Sixteen of these were further confirmed in planta interaction between AtCSP3 by a bi-molecular fluorescence complementation assay. We found that AtCSP3 interacts with CONSTANS-LIKE protein 15 and nuclear poly(A)-binding Proteins in nuclear speckles. Three 60S ribosomal Proteins (RPL26A, RPL40A/UBQ2, and RPL36aB) and the Gar1 RNA-binding protein interacted with AtCSP3 in the nucleolus and nucleoplasm, suggesting that AtCSP3 functions in ribosome biogenesis. Interactions with LOS2/enolase and glycine-rich RNA-binding protein 7 that are Cold inducible, and an mRNA decapping protein 5 (DCP5) were observed in the cytoplasm. These data suggest that AtCSP3 participates in multiple complexes that reside in nuclear and cytoplasmic compartments and possibly regulates RNA processing and functioning.
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Cold Shock domain protein 3 is involved in salt and drought stress tolerance in arabidopsis
FEBS Open Bio, 2013Co-Authors: Kentaro Sasaki, Ryozo Imai, Myunghee Kim, Shunya Sato, Wataru Saburi, Hirokazu MatsuiAbstract:Cold Shock Proteins (CSPs) of bacteria are produced in response to Cold and function as RNA chaperones that are essential for Cold adaptation. Arabidopsis thaliana Cold Shock DOMAIN PROTEIN 3 (AtCSP3) shares a domain with bacterial CSPs and is involved in acquisition of freezing tolerance. Our previous study revealed that many of the genes that are down regulated in an AtCSP3 knockout mutant (atcsp3–2) are functionally associated with responses to salt and drought as well as Cold. Here, we examined the involvement of AtCSP3 in salt and drought stress tolerance. We found that AtCSP3 is induced during salt and drought stresses, and is regulated by ABA. A knockout mutant of AtCSP3 (atcsp3–2) showed lower survival rates after salt and drought stress treatments. Conversely, the AtCSP3-overexpressing plants displayed higher survival rates after treatment with these stresses. Most of the genes that were down regulated in the atcsp3–2 mutant were found to be inducible upon salt and drought stresses, and upregulated in the AtCSP3-overexpressors. Together, our data demonstrates that AtCSP3 is involved in the regulation of salt and drought stress tolerance in Arabidopsis.
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Cold Shock domain protein 3 regulates freezing tolerance in arabidopsis thaliana
Journal of Biological Chemistry, 2009Co-Authors: Kentaro Sasaki, Ryozo ImaiAbstract:In response to Cold, Escherichia coli produces Cold Shock Proteins (CSPs) that have essential roles in Cold adaptation as RNA chaperones. Here, we demonstrate that Arabidopsis Cold Shock domain protein 3 (AtCSP3), which shares a Cold Shock domain with bacterial CSPs, is involved in the acquisition of freezing tolerance in plants. AtCSP3 complemented a Cold-sensitive phenotype of the E. coli CSP quadruple mutant and displayed nucleic acid duplex melting activity, suggesting that AtCSP3 also functions as an RNA chaperone. Promoter-GUS transgenic plants revealed tissue-specific expression of AtCSP3 in shoot and root apical regions. When exposed to low temperature, GUS activity was extensively induced in a broader region of the roots. In transgenic plants expressing an AtCSP3-GFP fusion, GFP signals were detected in both the nucleus and cytoplasm. An AtCSP3 knock-out mutant (atcsp3-2) was sensitive to freezing compared with wild-type plants under non-acclimated and Cold-acclimated conditions, whereas expression of C-repeat-binding factors and their downstream genes during Cold acclimation was not altered in the atcsp3-2 mutant. Overexpression of AtCSP3 in transgenic plants conferred enhanced freezing tolerance over wild-type plants. Together, the data demonstrated an essential role of RNA chaperones for Cold adaptation in higher plants.
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conservation of the Cold Shock domain protein family in plants
Plant Physiology, 2003Co-Authors: Dale Karlson, Ryozo ImaiAbstract:In this paper, we report the widespread occurrence of the nucleic acid-binding Cold Shock domain (CSD) in plants and identify the first eukaryotic homologs that are nearly identical to bacterial Cold Shock Proteins (CSP). Using Arabidopsis as a model system, we determined that its four unique CSD
Jochen Balbach - One of the best experts on this subject based on the ideXlab platform.
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rna single strands bind to a conserved surface of the major Cold Shock protein in crystals and solution
RNA, 2012Co-Authors: Rolf Sachs, Udo Heinemann, Klaas E A Max, Jochen BalbachAbstract:Bacterial Cold Shock Proteins (CSPs) regulate the cellular response to temperature downshift. Their general principle of function involves RNA chaperoning and transcriptional antitermination. Here we present two crystal structures of Cold Shock protein B from Bacillus subtilis (Bs-CspB) in complex with either a hexanucleotide (5′-UUUUUU-3′) or heptanucleotide (5′-GUCUUUA-3′) single-stranded RNA (ssRNA). Hydrogen bonds and stacking interactions between RNA bases and aromatic sidechains characterize individual binding subsites. Additional binding subsites which are not occupied by the ligand in the crystal structure were revealed by NMR spectroscopy in solution on Bs-CspB·RNA complexes. Binding studies demonstrate that Bs-CspB associates with ssDNA as well as ssRNA with moderate sequence specificity. Varying affinities of oligonucleotides are reflected mainly in changes of the dissociation rates. The generally lower binding affinity of ssRNA compared to its ssDNA analog is attributed solely to the substitution of thymine by uracil bases in RNA.
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common mode of dna binding to Cold Shock domains crystal structure of hexathymidine bound to the domain swapped form of a major Cold Shock protein from bacillus caldolyticus
FEBS Journal, 2007Co-Authors: Klaas E A Max, Jochen Balbach, Markus Zeeb, Ralf Bienert, Udo HeinemannAbstract:Bacterial Cold Shock Proteins (CSPs) regulate cellular adaptation to Cold stress. Functions ascribed to CSP include roles as RNA chaperones and in transcription antitermination. We present the crystal structure of the Bacillus caldolyticus CSP (Bc-Csp) in complex with hexathymidine (dT6) at a resolution of 1.29 A. Bound to dT6, crystalline Bc-Csp forms a domain-swapped dimer in which β strands 1–3 associate with strands 4 and 5 from the other subunit to form a closed β barrel and vice versa. The globular units of dimeric Bc-Csp closely resemble the well-known structure of monomeric CSP. Structural reorganization from the monomer to the domain-swapped dimer involves a strictly localized change in the peptide bond linking Glu36 and Gly37 of Bc-Csp. Similar structural reorganizations have not been found in any other CSP or oligonucleotide/oligosaccharide-binding fold structures. Each dT6 ligand is bound to one globular unit of Bc-Csp via an amphipathic protein surface. Individual binding subsites interact with the DNA bases through stacking and hydrogen bonding. The sugar–phosphate backbone remains solvent exposed. Based on crystallographic and biochemical studies of deoxyoligonucleotide binding to CSP, we suggest a common mode of binding of single-stranded heptanucleotide motifs to Proteins containing Cold Shock domains, including the eukaryotic Y-box factors.
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recognition of t rich single stranded dna by the Cold Shock protein bs cspb in solution
Nucleic Acids Research, 2006Co-Authors: Markus Zeeb, Klaas E A Max, Ulrich Weininger, Christian Low, Heinrich Sticht, Jochen BalbachAbstract:Cold Shock Proteins (CSP) belong to the family of single-stranded nucleic acid binding Proteins with OB-fold. CSP are believed to function as ‘RNA chaperones’ and during anti-termination. We determined the solution structure of Bs-CspB bound to the single-stranded DNA (ssDNA) fragment heptathymidine (dT7) by NMR spectroscopy. BsCspB reveals an almost invariant conformation when bound to dT7 with only minor reorientations in loop b1–b2 and b3–b4 and of few aromatic side chains involved in base stacking. Binding studies of protein variants and mutated ssDNA demonstrated that Bs-CspB associates with ssDNA at almost diffusion controlled rates and low sequence specificity consistent with its biological function. A variation of the ssDNA affinity is accomplished solely by changes of the dissociation rate. 15 N NMR relaxation and H/D exchange experiments revealed that binding of dT7 increases the stability of Bs-CspB and reduces the sub-nanosecond dynamics of the entire protein and especially of loop b3–b4.
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t rich dna single strands bind to a preformed site on the bacterial Cold Shock protein bs cspb
Journal of Molecular Biology, 2006Co-Authors: Klaas E A Max, Jochen Balbach, Markus Zeeb, Ralf Bienert, Udo HeinemannAbstract:Bacterial Cold Shock Proteins (CSPs) are involved in cellular adaptation to Cold stress. They bind to single-stranded nucleic acids with a KD value in the micro- to nanomolar range. Here we present the structure of the Bacillus subtilis CspB (Bs-CspB) in complex with hexathymidine (dT6) at a resolution of 1.78 A. Bs-CspB binds to dT6 with nanomolar affinity via an amphipathic interface on the protein surface. Individual binding subsites interact with single nucleobases through stacking interactions and hydrogen bonding. The sugar-phosphate backbone and the methyl groups of the thymine nucleobases remain solvent exposed and are not contacted by protein groups. Fluorescence titration experiments monitoring the binding of oligopyrimidines to Bs-CspB reveal binding preferences at individual subsites and allow the design of an optimised heptapyrimidine ligand, which is bound with sub-nanomolar affinity. This study reveals the stoichiometry and sequence determinants of the binding of single-stranded nucleic acids to a preformed site on Bs-CspB and thus provides the structural basis of the RNA chaperone and transcription antitermination activities of the CSP.
Hermann Schindelin - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of cspa the major Cold Shock protein of escherichia coli
Proceedings of the National Academy of Sciences of the United States of America, 1994Co-Authors: Hermann Schindelin, Masayori Inouye, Weining Jiang, Udo HeinemannAbstract:The major Cold Shock protein of Escherichia coli, CspA, produced upon a rapid downshift in growth temperature, is involved in the transcriptional regulation of at least two genes. The protein shares high homology with the nucleic acid-binding domain of the Y-box factors, a family of eukaryotic Proteins involved in transcriptional and translational regulation. The crystal structure of CspA has been determined at 2-A resolution and refined to R = 0.187. CspA is composed of five antiparallel beta-strands forming a closed five-stranded beta-barrel. The three-dimensional structure of CspA is similar to that of the major Cold Shock protein of Bacillus subtilis, CspB, which has recently been determined at 2.45-A resolution. However, in contrast to CspB, no dimer is formed in the crystal. The surface of CspA is characteristic for a protein interacting with single-stranded nucleic acids. Due to the high homology of the bacterial Cold Shock Proteins with the Y-box factors, E. coli CspA and B. subtilis CspB define a structural framework for the common Cold Shock domain.
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universal nucleic acid binding domain revealed by crystal structure of the b subtilis major Cold Shock protein
Nature, 1993Co-Authors: Hermann Schindelin, Moharned A. Marahiel, Udo HeinemannAbstract:THE Cold-Shock response in both Escherichia coli and Bacillus subtilis is induced by an abrupt downshift in growth temperature. It leads to the increased production of the major Cold-Shock Proteins, CS7.4 and CspB, respectively1–3. CS7.4 is a transcriptional activator of two genes4,5. CS7.4 and CspB share 43 per cent sequence identity with the nucleic acid-binding domain of the eukaryotic gene-regulatory Y-box factors6. This Cold-Shock domain is conserved from bacteria to man7 and contains the RNA-binding RNP1 sequence motif8. As a prototype of the Cold-Shock domain, the structure of CspB has been determined here from two crystal forms. In both, CspB is present as an antiparallel five-stranded β-barrel. Three consecutive β-strands, the central one containing the RNP1 motif, create a surface rich in aromatic and basic residues that are presumably involved in nucleic acid binding. Preferential binding of CspB to single-stranded DNA is observed in gel retardation experiments.
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universal nucleic acid binding domain revealed by crystal structure of the b subtilis major Cold Shock protein
Nature, 1993Co-Authors: Hermann Schindelin, Udo HeinemannAbstract:The Cold-Shock response in both Escherichia coli and Bacillus subtilis is induced by an abrupt downshift in growth temperature. It leads to the increased production of the major Cold-Shock Proteins, CS7.4 and CspB, respectively. CS7.4 is a transcriptional activator of two genes. CS7.4 and CspB share 43 per cent sequence identity with the nucleic acid-binding domain of the eukaryotic gene-regulatory Y-box factors. This Cold-Shock domain is conserved from bacteria to man and contains the RNA-binding RNP1 sequence motif. As a prototype of the Cold-Shock domain, the structure of CspB has been determined here from two crystal forms. In both, CspB is present as an antiparallel five-stranded beta-barrel. Three consecutive beta-strands, the central one containing the RNP1 motif, create a surface rich in aromatic and basic residues that are presumably involved in nucleic acid binding. Preferential binding of CspB to single-stranded DNA is observed in gel retardation experiments.