1,721,319 research outputs found
Novel Structural Insights into Class I and II Histone Deacetylases
No full textThe deacetylation of modified lysine residues of histones and other proteins is catalyzed by histone deacetlyases (HDACs). HDACs play an important role in the regulation of many biological processes such as cell-cycle, cell differentiation and survival. Since the inhibition of HDACs leads to growth arrest, differentiation or apoptosis of tumor cell lines, HDACs are promising targets for cancer therapy. Knowledge of the three-dimensional structures of HDACs with bound substrate or inhibitor molecules is a prerequisite for rational structure-based drug design. Here recent developments in the crystal structure analysis of human HDAC4, HDAC7, and HDAC8, which all belong to the family of zinc ion-dependent HDACs, are described. Crystallographic and biochemical studies of the catalytic domains of HDAC4 and HDAC7 revealed the molecular basis for their low enzymatic activity. Furthermore, the role of a second, structural zinc ion has been elucidated. The structures of HDAC8 with bound substrate-like peptide molecule demonstrate the functional role of a conserved aspartate residue located at the rim of the active site in substrate recognition. Structures of these three HDACs with various bound inhibitor molecules will provide the structural basis for further development of HDAC inhibitors with improved isoform-specific selectivity
Refined crystal structure of phycoerythrin from Porphyridium cruentum at 0.23-nm resolution and localization of the gamma subunit
No full textSequence analysis and overexpression of the Zymomonas mobilis tgt gene encoding tRNA-guanine transglycosylase: purification and biochemical characterization of the enzyme
No full texttRNA-guanine transglycosylase (Tgt) is involved in the biosynthesis of the hypermodified tRNA nucleoside queuosine (Q). It catalyzes the posttranscriptional base exchange of the Q precursor 7-aminomethyl-7-deazaguanine (preQ1) with the genetically encoded guanine in the anticodon of tRNA(Asp), tRNA(Asn), tRNA(His), and tRNA(Tyr). A partially sequenced gene upstream of the DNA ligase (lig) gene of the Zymomonas mobilis chromosome shows strong homology to the tgt gene of Escherichia coli (K.B. Shark and T. Conway, FEMS Microbiol. Lett. 96:19-26, 1992). We showed that this gene is able to complement the tgt mutation in E. coli SJ1505, and we determined its complete sequence. Four start codons were possible for this gene, resulting in proteins of 386 to 399 amino acids (M(r), 42,800 to 44,300) showing 60.4% sequence identity with Tgt from E. coli. The smallest of the four possible reading frames, which was still extended at its 5' end compared with the E. coli tgt gene, was overexpressed in E. coli. The gene product was purified to homogeneity and was biochemically characterized. The kinetical parameters were virtually identical to those published for the E. coli enzyme. In contrast to E. coli Tgt, which is reported to be a homotrimer, Z. mobilis Tgt was found to be a monomer according to gel filtration. In this study, it was shown that the formation of homotrimers by the E. coli enzyme is readily reversible and is dependent on protein concentration.tRNA-guanine transglycosylase (Tgt) is involved in the biosynthesis of the hypermodified tRNA nucleoside queuosine (Q). It catalyzes the posttranscriptional base exchange of the Q precursor 7-aminomethyl-7-deazaguanine (preQ1) with the genetically encoded guanine in the anticodon of tRNA(Asp), tRNA(Asn), tRNA(His), and tRNA(Tyr). A partially sequenced gene upstream of the DNA ligase (lig) gene of the Zymomonas mobilis chromosome shows strong homology to the tgt gene of Escherichia coli (K.B. Shark and T. Conway, FEMS Microbiol. Lett. 96:19-26, 1992). We showed that this gene is able to complement the tgt mutation in E. coli SJ1505, and we determined its complete sequence. Four start codons were possible for this gene, resulting in proteins of 386 to 399 amino acids (M(r), 42,800 to 44,300) showing 60.4% sequence identity with Tgt from E. coli. The smallest of the four possible reading frames, which was still extended at its 5' end compared with the E. coli tgt gene, was overexpressed in E. coli. The gene product was purified to homogeneity and was biochemically characterized. The kinetical parameters were virtually identical to those published for the E. coli enzyme. In contrast to E. coli Tgt, which is reported to be a homotrimer, Z. mobilis Tgt was found to be a monomer according to gel filtration. In this study, it was shown that the formation of homotrimers by the E. coli enzyme is readily reversible and is dependent on protein concentration
Assessment of model bias in crystallographic maps and its implications for validation of crystal structures
No full textRNA-modification by Base Exchange: Structure, Function and Application of tRNA-guanine Transglycosylases
No full texthttp://dx.doi.org/10.13039/501100001659 German Research Foundatio
RNA synthesis and purification for structural studies
No full textRNAs play pivotal roles in the cell, ranging from catalysis (e.g., RNase P), acting as adaptor molecule (tRNA) to regulation (e.g., riboswitches). Precise understanding of its three-dimensional structures has given unprecedented insight into the molecular basis for all of these processes. Nevertheless, structural studies on RNA are still limited by the very special nature of this polymer. The most common methods for the determination of 3D RNA structures are NMR and X-ray crystallography. Both methods have their own set of requirements and give different amounts of information about the target RNA. For structural studies, the major bottleneck is usually obtaining large amounts of highly pure and homogeneously folded RNA. Especially for X-ray crystallography it can be necessary to screen a large number of variants to obtain well-ordered single crystals. In this mini-review we give an overview about strategies for the design, in vitro production, and purification of RNA for structural studies
Structural analysis of the spliceosomal RNA helicase Prp28 from the thermophilic eukaryote Chaetomium thermophilum
No full textPrp28 (pre-mRNA-splicing ATP-dependent RNA helicase 28) is a spliceosomal DEAD-box helicase which is involved in two steps of spliceosome assembly. It is required for the formation of commitment complex 2 in an ATP-independent manner as well as for the formation of the pre-catalytic spliceosome, which in contrast is ATP-dependent. During the latter step, Prp28 is crucial for the integration of the U4/U6.U5 tri-snRNP since it displaces the U1 snRNP and allows the U6 snRNP to base-pair with the 5'-splice site. Here, the crystal structure of Prp28 from the thermophilic fungus Chaetomium thermophilum is reported at 3.2 angstrom resolution and is compared with the available structures of homologues
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