CCP4i2: the new graphical user interface to theCCP4 program suite

Potterton, Liz; Agirre, Jon; Ballard, Charles; Cowtan, Kevin; Dodson, E.J.; Evans, Phil; Jenkins, Huw T.; Keegan, Ronan; Krissinel, Eugene; Stevenson, Kyle; Лебедев, А. А.; McNicholas, Stuart; Nicholls, Robert A.; Noble, M.E.M.; Pannu, Navraj S.; Roth, Christian; Sheldrick, George M.; Skubák, Pavol; Turkenburg, J.P.; Uski, Ville; Delft, Frank von; Waterman, David; Wilson, K.S.; Winn, Martyn; Wojdyr, Marcin

doi:10.1107/s2059798317016035

Cited by 443 publications

(389 citation statements)

References 41 publications

Supporting

Mentioning

389

Contrasting

Order By: Relevance

“…All other datasets were collected using a Pilatus 2 M detector on beamline X06DA (PXIII) at the Swiss Light Source, Paul Scherrer Institut, Villigen, Switzerland. The structures were solved by molecular replacement using the MOLREP program (51) in the CCP4 program suite (52). The structure was refined with REFMAC (53).…”

Section: Ccd-ibd Interaction Assay-in-mentioning

confidence: 99%

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Bonnard

Rouzic

Eiler

et al. 2018

Journal of Biological Chemistry

View full text Add to dashboard Cite

Recently, a new class of HIV-1 integrase (IN) inhibitors with a dual mode of action, called IN-LEDGF/p75 allosteric inhibitors (INLAIs), was described. Designed to interfere with the IN-LEDGF/p75 interaction during viral integration, unexpectedly, their major impact was on virus maturation. This activity has been linked to induction of aberrant IN multimerization, while inhibition of the IN-LEDGF/p75 interaction accounts for weaker antiretroviral effect at integration. Since these dual activities result from INLAI binding to IN at a single binding site, we expected that these activities co-evolved together, driven by the affinity for IN. Using an original INLAI, MUT-A, and its activity on an Ala-125 (A125) IN variant, we found that these two activities on A125 IN can be fully dissociated: MUT-A-induced IN multimerization and the formation of eccentric condensates in viral particles, that are responsible for inhibition of virus maturation, were lost, while inhibition of the IN-LEDGF/p75 interaction and consequently integration, was fully retained. Hence the mere binding of INLAI to A125 IN is insufficient to promote the conformational changes of IN required for aberrant multimerization. By analyzing the Xray structures of MUT-A bound to the IN catalytic core domain (CCD) with or without the A125 polymorphism, we discovered that the loss of IN multimerization is due to stabilization of the A125 IN variant CCD dimer, highlighting the importance of the CCD dimerization energy for IN multimerization. Our study reveals that affinity for the LEDGF/p75-binding pocket is not sufficient to induce INLAI-dependent IN multimerization and the associated inhibition of viral maturation.The integrase (IN) protein of Human Immunodeficiency Virus type 1 (HIV-1) catalyzes the stable insertion of the viral cDNA genome into the host cell chromatin, a step of the viral life cycle that is required for efficient viral gene expression. Integration occurs via a two-step reaction where IN initially cleaves after a conserved CA dinucleotide at the 3' end of the viral cDNA genome to free a 3'-OH group (3' processing), which is next used to carry out a nucleophilic attack on cellular chromosomal DNA (strand transfer).IN is one of the preferred targets for the development of antiretroviral (ARV) drugs. However, given the high genetic variability of HIV-1, IN mutations conferring cross-resistance to the first generation INSTIs, RAL and EVG, were described in patients receiving INSTI-containing regimens (2). The second generation INSTI DTG has a higher genetic barrier and conserves good ARV activity against a number of RAL-and EVGresistant strains. Recent reports showed that Bictegravir, a second generation INSTI still in development from Gilead Sciences, has a resistance profile similar to DTG (3). Nevertheless, DTG and Bictegravir are sensitive to the most detrimental INSTI resistant mutations albeit at lower levels than first generation INSTIs (4). Therefore, the development of small molecule inhibitors impairing IN functions with dis...

show abstract

Section: Ccd-ibd Interaction Assay-in-mentioning

confidence: 99%

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Bonnard

Rouzic

Eiler

et al. 2018

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…From this map, we were able to build our model using COOT (35). After model building, Refmac from the CCP4i suite was use for refinement (36)(37)(38). The final cross-validated R free after model refinement was 25.7% (Tables S1 and S2).…”

Section: Methodsmentioning

confidence: 99%

Crystal structure of Pistol, a class of self-cleaving ribozyme

Nguyen

Wang

Steitz

2017

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Small self-cleaving ribozymes have been discovered in all evolutionary domains of life. They can catalyze site-specific RNA cleavage, and as a result, they have relevance in gene regulation. Comparative genomic analysis has led to the discovery of a new class of small selfcleaving ribozymes named Pistol. We report the crystal structure of Pistol at 2.97-Å resolution. Our results suggest that the Pistol ribozyme self-cleavage mechanism likely uses a guanine base in the active site pocket to carry out the phosphoester transfer reaction. The guanine G40 is in close proximity to serve as the general base for activating the nucleophile by deprotonating the 2′-hydroxyl to initiate the reaction (phosphoester transfer). Furthermore, G40 can also establish hydrogen bonding interactions with the nonbridging oxygen of the scissile phosphate. The proximity of G32 to the O5′ leaving group suggests that G32 may putatively serve as the general acid. The RNA structure of Pistol also contains A-minor interactions, which seem to be important to maintain its tertiary structure and compact fold. Our findings expand the repertoire of ribozyme structures and highlight the conserved evolutionary mechanism used by ribozymes for catalysis.T he "RNA world" hypothesis speculates that RNA carried out the majority of biochemical reactions before the evolution of complex protein enzymes (1, 2). Ribozymes are noncoding RNA that carry out catalytic activities. Unlike protein enzymes, only a handful of ribozymes have known biological functions. Their biological functions range from regulating gene expression (e.g., riboswitches) and performing peptidyl-transfer reactions (e.g., ribosome) to removing intron sequences in genes (e.g., self-splicing Group I intron ribozymes) (2-9). The biological functions and mechanism of these ribozymes have been discovered through structural and biochemical studies.Currently, the classes of self-cleaving ribozymes consist of Hammerhead, Hairpin, Hepatitis Delta Virus ribozyme (HDV), Varkud Satellite (VS), glmS ribozyme, Twister, Twister sister, Hatchet, and Pistol (10). These classes differ based on their size, structure, and cleavage mechanism. Known for their site-specific cleavage, ribozymes with defined biological function include the Hammerhead, VS, and HDV, which all participate in rolling circle replication, whereas the glmS ribozyme functions in controlling gene expression (11,12). However, the biological function of a vast majority of the different self-cleaving ribozymes remains to be explored.Through comparative genome analysis, there have been three newly identified classes of self-cleaving ribozymes called Twister sister, Hatchet, and Pistol (3). Biochemical analysis reveals that Pistol can use a variety of divalent metal ions to carry out a complete site-specific, self-cleaving reaction, whereas utilization of monovalent cations results in modest cleavage rates (3, 10). The rate of Pistol self-cleavage has been estimated to be >10 min −1 under physiological conditions and >100 min −1 under optimal ...

show abstract

“…Initial phase estimates for the structure factors were obtained using the molecular replacement program Phaser [47] within the CCP4i GUI [48,49]. For molecular replacement, a tetramer of wt bradavidin [PDB:2Y32] (residues 2–119 of each chain A-D) was used as the search model.…”

Section: Methodsmentioning

confidence: 99%

Structural characterization of core-bradavidin in complex with biotin

et al. 2017

View full text Add to dashboard Cite

Bradavidin is a tetrameric biotin-binding protein similar to chicken avidin and bacterial streptavidin, and was originally cloned from the nitrogen-fixing bacteria Bradyrhizobium diazoefficiens. We have previously reported the crystal structure of the full-length, wild-type (wt) bradavidin with 138 amino acids, where the C-terminal residues Gly129-Lys138 (“Brad-tag”) act as an intrinsic ligand (i.e. Gly129-Lys138 bind into the biotin-binding site of an adjacent subunit within the same tetramer) and has potential as an affinity tag for biotechnological purposes. Here, the X-ray structure of core-bradavidin lacking the C-terminal residues Gly114-Lys138, and hence missing the Brad-tag, was crystallized in complex with biotin at 1.60 Å resolution [PDB:4BBO]. We also report a homology model of rhodavidin, an avidin-like protein from Rhodopseudomonas palustris, and of an avidin-like protein from Bradyrhizobium sp. Ai1a-2, both of which have the Brad-tag sequence at their C-terminus. Moreover, core-bradavidin V1, an engineered variant of the original core-bradavidin, was also expressed at high levels in E. coli, as well as a double mutant (Cys39Ala and Cys69Ala) of core-bradavidin (CC mutant). Our data help us to further engineer the core-bradavidin–Brad-tag pair for biotechnological assays and chemical biology applications, and provide deeper insight into the biotin-binding mode of bradavidin.

show abstract

CCP4i2: the new graphical user interface to theCCP4 program suite

Cited by 443 publications

References 41 publications

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Crystal structure of Pistol, a class of self-cleaving ribozyme

Structural characterization of core-bradavidin in complex with biotin

Contact Info

Product

Resources

About