PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-5 (5)
 

Clipboard (0)
None

Select a Filter Below

Journals
Year of Publication
Document Types
1.  Structure solution of DNA-binding proteins and complexes with ARCIMBOLDO libraries 
The structure solution of DNA-binding protein structures and complexes based on the combination of location of DNA-binding protein motif fragments with density modification in a multi-solution frame is described.
Protein–DNA interactions play a major role in all aspects of genetic activity within an organism, such as transcription, packaging, rearrangement, replication and repair. The molecular detail of protein–DNA interactions can be best visualized through crystallography, and structures emphasizing insight into the principles of binding and base-sequence recognition are essential to understanding the subtleties of the underlying mechanisms. An increasing number of high-quality DNA-binding protein structure determinations have been witnessed despite the fact that the crystallographic particularities of nucleic acids tend to pose specific challenges to methods primarily developed for proteins. Crystallographic structure solution of protein–DNA complexes therefore remains a challenging area that is in need of optimized experimental and computational methods. The potential of the structure-solution program ARCIMBOLDO for the solution of protein–DNA complexes has therefore been assessed. The method is based on the combination of locating small, very accurate fragments using the program Phaser and density modification with the program SHELXE. Whereas for typical proteins main-chain α-helices provide the ideal, almost ubiquitous, small fragments to start searches, in the case of DNA complexes the binding motifs and DNA double helix constitute suitable search fragments. The aim of this work is to provide an effective library of search fragments as well as to determine the optimal ARCIMBOLDO strategy for the solution of this class of structures.
doi:10.1107/S1399004714007603
PMCID: PMC4051508  PMID: 24914984
protein–DNA complexes and macromolecule structure solutions; structure-solution pipelines; molecular replacement; density modification
2.  The Structure of the Oligomerization Domain of Lsr2 from Mycobacterium tuberculosis Reveals a Mechanism for Chromosome Organization and Protection 
PLoS ONE  2012;7(6):e38542.
Lsr2 is a small DNA-binding protein present in mycobacteria and related actinobacteria that regulates gene expression and influences the organization of bacterial chromatin. Lsr2 is a dimer that binds to AT-rich regions of chromosomal DNA and physically protects DNA from damage by reactive oxygen intermediates (ROI). A recent structure of the C-terminal DNA-binding domain of Lsr2 provides a rationale for its interaction with the minor groove of DNA, its preference for AT-rich tracts, and its similarity to other bacterial nucleoid-associated DNA-binding domains. In contrast, the details of Lsr2 dimerization (and oligomerization) via its N-terminal domain, and the mechanism of Lsr2-mediated chromosomal cross-linking and protection is unknown. We have solved the structure of the N-terminal domain of Lsr2 (N-Lsr2) at 1.73 Å resolution using crystallographic ab initio approaches. The structure shows an intimate dimer of two ß–ß–a motifs with no close homologues in the structural databases. The organization of individual N-Lsr2 dimers in the crystal also reveals a mechanism for oligomerization. Proteolytic removal of three N-terminal residues from Lsr2 results in the formation of an anti-parallel β-sheet between neighboring molecules and the formation of linear chains of N-Lsr2. Oligomerization can be artificially induced using low concentrations of trypsin and the arrangement of N-Lsr2 into long chains is observed in both monoclinic and hexagonal crystallographic space groups. In solution, oligomerization of N-Lsr2 is also observed following treatment with trypsin. A change in chromosomal topology after the addition of trypsin to full-length Lsr2-DNA complexes and protection of DNA towards DNAse digestion can be observed using electron microscopy and electrophoresis. These results suggest a mechanism for oligomerization of Lsr2 via protease-activation leading to chromosome compaction and protection, and concomitant down-regulation of large numbers of genes. This mechanism is likely to be relevant under conditions of stress where cellular proteases are known to be upregulated.
doi:10.1371/journal.pone.0038542
PMCID: PMC3374832  PMID: 22719899
3.  High-Resolution Crystal Structure of an Outer Membrane-Anchored Endolytic Peptidoglycan Lytic Transglycosylase (MltE) from Escherichia coli 
Biochemistry  2011;50(13):2384-2386.
The crystal structure of the first endolytic peptidoglycan lytic transglycosylase MltE from Escherichia coli is reported herein. The degradative activity of this enzyme initiates the process of cell wall recycling, which is an integral event in the bacterial existence. The structure sheds light on how MltE recognizes its substrate, the cell wall peptidoglycan. It also explains the ability of this endolytic enzyme to cleave in the middle of the peptidoglycan chains. Furthermore, the structure reveals how the enzyme is sequestered on the inner leaf let of the outer membrane.
doi:10.1021/bi200085y
PMCID: PMC3068208  PMID: 21341761
4.  Practical structure solution with ARCIMBOLDO  
ARCIMBOLDO combines the location of small fragments with Phaser and density modification with SHELXE of all possible Phaser solutions. Its uses are explained and illustrated through practical test cases.
Since its release in September 2009, the structure-solution program ARCIMBOLDO, based on the combination of locating small model fragments such as polyalanine α-helices with density modification with the program SHELXE in a multisolution frame, has evolved to incorporate other sources of stereochemical or experimental information. Fragments that are more sophisticated than the ubiquitous main-chain α-­helix can be proposed by modelling side chains onto the main chain or extracted from low-homology models, as locally their structure may be similar enough to the unknown one even if the conventional molecular-replacement approach has been unsuccessful. In such cases, the program may test a set of alternative models in parallel against a specified figure of merit and proceed with the selected one(s). Experimental information can be incorporated in three ways: searching within ARCIMBOLDO for an anomalous fragment against anomalous differences or MAD data or finding model fragments when an anomalous substructure has been determined with another program such as SHELXD or is subsequently located in the anomalous Fourier map calculated from the partial fragment phases. Both sources of information may be combined in the expansion process. In all these cases the key is to control the workflow to maximize the chances of success whilst avoiding the creation of an intractable number of parallel processes. A GUI has been implemented to aid the setup of suitable strategies within the various typical scenarios. In the present work, the practical application of ARCIMBOLDO within each of these scenarios is described through the distributed test cases.
doi:10.1107/S0907444911056071
PMCID: PMC3322593  PMID: 22505254
ARCIMBOLDO; fragment search; Phaser; density modification; multi-solution phasing; SHELXE
5.  6-Benzyl-3,4-dimeth­oxy-10-methyl­pyrido[2′,1′:2,3]imidazo[4,5-c]isoquinolin-5(6H)-one 
Pyrido[2′,1′:2,3]imidazo[4,5-c]isoquinolin-5(6H)-ones such as the title compound, C24H21N3O3, can be obtained in a few minutes in a microwave-assisted three-component reaction from 2-amino­pyridines, isocyanides and 2-carboxy­benz­aldehydes. In the title compound, the pyrido[2′,1′:2,3]imidazo[4,5-c]isoquinolin-5(6H)-one ring system is almost planar (mean deviation 0.068 Å). The dihedral angle between the benzyl ring and the pyrido[2′,1′:2,3]imidazo[4,5-c]isoquinolin-5(6H)-one ring system is 78.2°. The crystal structure is stabilized by inter­molecular C—H⋯O and C—H⋯N hydrogen bonds.
doi:10.1107/S1600536809035806
PMCID: PMC2970274  PMID: 21577919

Results 1-5 (5)