The structure of SSO2064, the first representative of Pfam family PF01796, reveals a novel two-domain zinc-ribbon OB-fold architecture with a potential acyl-CoA-binding role

AbstractConserved telomere maintenance component 1 (CTC1) is an important component of the CST (CTC1-STN1-TEN1) complex, involved in maintaining the stability of telomeric DNA. Several non-synonymous single-nucleotide polymorphisms (nsSNPs) in CTC1 have been reported to cause Coats plus syndrome and Dyskeratosis congenital diseases. Here, we have performed sequence and structure analyses of nsSNPs of CTC1 using state-of-the-art computational methods. The structure-based study focuses on the C-terminal OB-fold region of CTC1. There are 11 pathogenic mutations identified, and detailed structural analyses were performed. These mutations cause a significant disruption of noncovalent interactions, which may be a possible reason for CTC1 instability and consequent diseases. To see the impact of such mutations on the protein conformation, all-atom molecular dynamics (MD) simulations of CTC1-wild-type (WT) and two of the selected mutations, R806C and R806L for 200 ns, were carried out. A significant conformational change in the structure of the R806C mutant was observed. This study provides a valuable direction to understand the molecular basis of CTC1 dysfunction in disease progression, including Coats plus syndrome.

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Engineering of Functional Replication Protein A Homologs Based on Insights into the Evolution of Oligonucleotide/ Oligosaccharide-Binding Folds

Journal of Bacteriology ◽

10.1128/jb.01930-07 ◽

2008 ◽

Vol 190 (17) ◽

pp. 5766-5780 ◽

Cited By ~ 10

Author(s):

Yuyen Lin ◽

Li-Jung Lin ◽

Palita Sriratana ◽

Kelli Coleman ◽

Taekjip Ha ◽

...

Keyword(s):

Homologous Recombination ◽

Protein A ◽

Replication Protein A ◽

Replication Protein ◽

Methanosarcina Acetivorans ◽

Functional Studies ◽

Functional Homolog ◽

Ob Fold ◽

Methanothermobacter Thermautotrophicus ◽

Related Proteins

ABSTRACT The bacterial single-stranded DNA-binding protein (SSB) and the archaeal/eukaryotic functional homolog, replication protein A (RPA), are essential for most aspects of DNA metabolism. Structural analyses of the architecture of SSB and RPA suggest that they are composed of different combinations of a module called the oligonucleotide/oligosaccharide-binding (OB) fold. Members of the domains Bacteria and Eukarya, in general, contain one type of SSB or RPA. In contrast, organisms in the archaeal domain have different RPAs made up of different organizations of OB folds. Interestingly, the euryarchaeon Methanosarcina acetivorans harbors multiple functional RPAs named MacRPA1 (for M. acetivorans RPA 1), MacRPA2, and MacRPA3. Comparison of MacRPA1 with related proteins in the publicly available databases suggested that intramolecular homologous recombination might play an important role in generating some of the diversity of OB folds in archaeal cells. On the basis of this information, from a four-OB-fold-containing RPA, we engineered chimeric modules to create three-OB-fold-containing RPAs to mimic a novel form of RPA found in Methanococcoides burtonii and Methanosaeta thermophila. We further created two RPAs that mimicked the RPAs in Methanocaldococcus jannaschii and Methanothermobacter thermautotrophicus through fusions of modules from MacRPA1 and M. thermautotrophicus RPA. Functional studies of these engineered proteins suggested that fusion and shuffling of OB folds can lead to well-folded polypeptides with most of the known properties of SSB and RPAs. On the basis of these results, different models that attempt to explain how intramolecular and intermolecular homologous recombination can generate novel forms of SSB or RPAs are proposed.

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Redefining the PF06864 Pfam Family Based on Burkholderia pseudomallei PilO2Bp S-SAD Crystal Structure

PLoS ONE ◽

10.1371/journal.pone.0094981 ◽

2014 ◽

Vol 9 (4) ◽

pp. e94981 ◽

Cited By ~ 4

Author(s):

Patricia Lassaux ◽

Oscar Conchillo-Solé ◽

Babu A. Manjasetty ◽

Daniel Yero ◽

Lucia Perletti ◽

...

Keyword(s):

Crystal Structure ◽

Burkholderia Pseudomallei ◽

Pfam Family ◽

Family Based

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Overexpression of the Bacteriophage T4 motB Gene Alters H-NS Dependent Repression of Specific Host DNA

Viruses ◽

10.3390/v13010084 ◽

2021 ◽

Vol 13 (1) ◽

pp. 84

Author(s):

Jennifer Patterson-West ◽

Chin-Hsien Tai ◽

Bokyung Son ◽

Meng-Lun Hsieh ◽

James R. Iben ◽

...

Keyword(s):

Bacteriophage T4 ◽

Early Gene ◽

Dna Binding Domain ◽

Host Gene Expression ◽

Rna Seq ◽

Specific Host ◽

Dnase I Footprinting ◽

Ob Fold ◽

Host Genes

The bacteriophage T4 early gene product MotB binds tightly but nonspecifically to DNA, copurifies with the host Nucleoid Associated Protein (NAP) H-NS in the presence of DNA and improves T4 fitness. However, the T4 transcriptome is not significantly affected by a motB knockdown. Here we have investigated the phylogeny of MotB and its predicted domains, how MotB and H-NS together interact with DNA, and how heterologous overexpression of motB impacts host gene expression. We find that motB is highly conserved among Tevenvirinae. Although the MotB sequence has no homology to proteins of known function, predicted structure homology searches suggest that MotB is composed of an N-terminal Kyprides-Onzonis-Woese (KOW) motif and a C-terminal DNA-binding domain of oligonucleotide/oligosaccharide (OB)-fold; either of which could provide MotB’s ability to bind DNA. DNase I footprinting demonstrates that MotB dramatically alters the interaction of H-NS with DNA in vitro. RNA-seq analyses indicate that expression of plasmid-borne motB up-regulates 75 host genes; no host genes are down-regulated. Approximately 1/3 of the up-regulated genes have previously been shown to be part of the H-NS regulon. Our results indicate that MotB provides a conserved function for Tevenvirinae and suggest a model in which MotB functions to alter the host transcriptome, possibly by changing the association of H-NS with the host DNA, which then leads to conditions that are more favorable for infection.

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The YebC Family Protein PA0964 Negatively Regulates the Pseudomonas aeruginosa Quinolone Signal System and Pyocyanin Production

Journal of Bacteriology ◽

10.1128/jb.00428-08 ◽

2008 ◽

Vol 190 (18) ◽

pp. 6217-6227 ◽

Cited By ~ 72

Author(s):

Haihua Liang ◽

Lingling Li ◽

Zhaolin Dong ◽

Michael G. Surette ◽

Kangmin Duan

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Virulence Factors ◽

Response Regulator ◽

Initial Study ◽

Pfam Family ◽

Swarming Motility ◽

Sensing Systems ◽

Family Protein ◽

Pyocyanin Production

ABSTRACT Bacterial pathogenicity is often manifested by the expression of various cell-associated and secreted virulence factors, such as exoenzymes, protease, and toxins. In Pseudomonas aeruginosa, the expression of virulence genes is coordinately controlled by the global regulatory quorum-sensing systems, which includes the las and rhl systems as well as the Pseudomonas quinolone signal (PQS) system. Phenazine compounds are among the virulence factors under the control of both the rhl and PQS systems. In this study, regulation of the phzA1B1C1D1E1 (phzA1) operon, which is involved in phenazine synthesis, was investigated. In an initial study of inducing conditions, we observed that phzA1 was induced by subinhibitory concentrations of tetracycline. Screening of 13,000 mutants revealed 32 genes that altered phzA1 expression in the presence of subinhibitory tetracycline concentrations. Among them, the gene PA0964, designated pmpR ( p qsR-mediated P QS r egulator), has been identified as a novel regulator of the PQS system. It belongs to a large group of widespread conserved hypothetical proteins with unknown function, the YebC protein family (Pfam family DUF28). It negatively regulates the quorum-sensing response regulator pqsR of the PQS system by binding at its promoter region. Alongside phzA1 expression and phenazine and pyocyanin production, a set of virulence factors genes controlled by both rhl and the PQS were shown to be modulated by PmpR. Swarming motility and biofilm formation were also significantly affected. The results added another layer of regulation in the rather complex quorum-sensing systems in P. aeruginosa and demonstrated a clear functional clue for the YebC family proteins.

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Cell Cycle Localization, Dimerization, and Binding Domain Architecture of the Telomere Protein cPot1

Molecular and Cellular Biology ◽

10.1128/mcb.24.5.2091-2102.2004 ◽

2004 ◽

Vol 24 (5) ◽

pp. 2091-2102 ◽

Cited By ~ 28

Author(s):

Chao Wei ◽

Carolyn M. Price

Keyword(s):

Cell Cycle ◽

Dna Binding ◽

Binding Site ◽

Domain Architecture ◽

Binding Domain ◽

Binding Motif ◽

Dna Binding Domain ◽

Telomere Protein ◽

Ob Fold

ABSTRACT Pot1 is a single-stranded-DNA-binding protein that recognizes telomeric G-strand DNA. It is essential for telomere capping in Saccharomyces pombe and regulates telomere length in humans. Human Pot1 also interacts with proteins that bind the duplex region of the telomeric tract. Thus, like Cdc13 from S. cerevisiae, Pot 1 may have multiple roles at the telomere. We show here that endogenous chicken Pot1 (cPot1) is present at telomeres during periods of the cell cycle when t loops are thought to be present. Since cPot1 can bind internal loops and directly adjacent DNA-binding sites, it is likely to fully coat and protect both G-strand overhangs and the displaced G strand of a t loop. The minimum binding site of cPot1 is double that of the S. pombe DNA-binding domain. Although cPot can self associate, dimerization is not required for DNA binding and hence does not explain the binding-site duplication. Instead, the DNA-binding domain appears to be extended to contain a second binding motif in addition to the conserved oligonucleotide-oligosaccharide (OB) fold present in other G-strand-binding proteins. This second motif could be another OB fold. Although dimerization is inefficient in vitro, it may be regulated in vivo and could promote association with other telomere proteins and/or telomere compaction.

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The C-terminal domain of Clostridioides difficile TcdC is exposed on the bacterial cell surface

10.1101/872283 ◽

2019 ◽

Author(s):

Ana M. Oliveira Paiva ◽

Leen de Jong ◽

Annemieke H. Friggen ◽

Wiep Klaas Smits ◽

Jeroen Corver

Keyword(s):

Sigma Factor ◽

Negative Regulator ◽

Toxin Gene ◽

Cytoplasmic Localization ◽

Terminal Domain ◽

Clostridioides Difficile ◽

C Terminus ◽

N Terminus ◽

Ob Fold ◽

Toxin Gene Expression

AbstractClostridioides difficile is an anaerobic gram-positive bacterium that can can produce the large clostridial toxins, Toxin A and Toxin B, encoded within the pathogenicity locus (PaLoc). The PaLoc also encodes the sigma factor TcdR, that positively regulates toxin gene expression, and TcdC, a putative negative regulator of toxin expression. TcdC is proposed to be an anti-sigma factor, however, several studies failed to show an association between tcdC genotype and toxin production. Consequently, TcdC function is not yet fully understood. Previous studies have characterized TcdC as a membrane-associated protein with the ability to bind G-quadruplex structures. The binding to the DNA secondary structures is mediated through the OB-fold domain present at the C-terminus of the protein. This domain was previously also proposed to be responsible for the inhibitory effect on toxin gene expression, implicating a cytoplasmic localization of the C-terminal OB-fold.In this study we aimed to obtain topological information on the C-terminus of TcdC. Using Scanning Cysteine Accessibility Mutagenesis and a HiBiT-based system, we demonstrate that the C-terminus of TcdC is located extracellularly. The extracellular location of TcdC is not compatible with direct binding of the OB-fold domain to intracellular nucleic acid or protein targets, and suggests a mechanism of action that is different from characterized anti-sigma factors.ImportanceTranscription of the C. difficile large clostrididial toxins (TcdA and TcdB) is directed by the sigma factor TcdR. TcdC has been implicated as a negative regulator, possible acting as an anti-sigma factor.Activity of TcdC has been mapped to its C-terminal OB fold domain. TcdC is anchored in the bacterial membrane, through its hydrophobic N-terminus and acting as an anti-sigma factor would require cytoplasmic localization of the C-terminal domain.Remarkably, topology predictions for TcdC suggest the N-terminus to be membrane localized and the C-terminal domain to be located extracellularly. Using independent assays, we show that the C-terminus of TcdC indeed is located in the extracellular environment, which is incompatible with its proposed role as anti-sigma factor in toxin regulation.

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