tRNASer acceptor stem: conformation and hydration of a microhelix in a crystal structure at 1.8 A resolution.

The crystal structure of a serine-specific tRNA acceptor-stem microhelix, the binding site for the seryl-tRNA synthetase, was solved by X-ray analysis. This seven-base-pair tRNA(Ser) microhelix forms endless rows of helices in the crystal lattice, with two helices stacking 'head-to-head' onto each other, resulting in an intermolecular guanosine stacking of the first purine nucleotides at the 5'-strands of the tRNA(Ser) microhelices. A network of 75 water loci could be associated with each RNA duplex.

Human tRNA(Gly) acceptor-stem microhelix: crystallization and preliminary X-ray diffraction analysis at 1.2 A resolution.

The major dissimilarities between the eukaryotic/archaebacterial-type and eubacterial-type glycyl-tRNA synthetase systems (GlyRS; class II aminoacyl-tRNA synthetases) represent an intriguing example of evolutionarily divergent solutions to similar biological functions. The differences in the identity elements of the respective tRNA(Gly) systems are located within the acceptor stem and include the discriminator base U73. In the present work, the human tRNA(Gly) acceptor-stem microhelix was crystallized in an attempt to analyze the structural features that govern the correct recognition of tRNA(Gly) by the eukaryotic/archaebacterial-type glycyl-tRNA synthetase.

Cocrystallizing natural RNA with its unnatural mirror image: biochemical and preliminary X-ray diffraction analysis of a 5S rRNA A-helix racemate.

Chemically synthesized RNAs with the unnatural L-configuration possess enhanced in vivo stability and nuclease resistance, which is a highly desirable property for pharmacological applications. For a structural comparison, both L- and D-RNA oligonucleotides of a shortened Thermus flavus 5S rRNA A-helix were chemically synthesized. The enantiomeric RNA duplexes were stochiometrically cocrystallized as a racemate, which enabled analysis of the D- and L-RNA enantiomers in the same crystals.

Resisting degradation by human elastase: commonality of design features shared by 'canonical' plant and bacterial macrocyclic protease inhibitor scaffolds.

A previously unexplained difference in the resistance to enzymatic hydrolysis of 11-mer Bowman-Birk-type inhibitors of human leukocyte elastase that differ in P1 is found to correlate with the strength of a particular intramolecular hydrogen bond within the inhibitor. This transannular hydrogen bond stabilizes the side chain of the conserved P2 Thr in a 'canonical' +60 degrees -rotamer chi(1) conformation and thereby directs it for a close interaction with the enzyme's catalytic His. As the implications of this NMR analysis are neither limited to this macrocyclic scaffold derived from plant proteins nor to a particular serine protease, we present a unified analysis with inhibitory bacterial depsipeptides of 7-12 residues in length that share key design features for which we propose communal functional explanations.

Crystallization and preliminary X-ray diffraction analysis of an Escherichia coli tRNA(Gly) acceptor-stem microhelix.

The tRNA(Gly) and glycyl-tRNA synthetase (GlyRS) system is an evolutionary special case within the class II aminoacyl-tRNA synthetases because two divergent types of GlyRS exist: an archaebacterial/human type and an eubacterial type. The tRNA identity elements which determine the correct aminoacylation process are located in the aminoacyl domain of tRNA(Gly). To obtain further insight concerning structural investigation of the identity elements, the Escherichia coli seven-base-pair tRNA(Gly) acceptor-stem helix was crystallized.

Comparative crystallization and preliminary X-ray diffraction studies of locked nucleic acid and RNA stems of a tenascin C-binding aptamer.

The pharmacokinetic properties of an aptamer against the tumour-marker protein tenascin-C have recently been successfully improved by the introduction of locked nucleic acids (LNAs) into the terminal stem of the aptamer. Since it is believed that this post-SELEX optimization is likely to provide a more general route to enhance the in vitro and in vivo stability of aptamers, elucidation of the structural basis of this improvement was embarked upon. Here, the crystallographic and X-ray diffraction data of the isolated aptamer stem encompassed in a six-base-pair duplex both with and without the LNA modification are presented.

Crystallization and preliminary X-ray diffraction analysis of a tRNASer acceptor-stem microhelix.

In order to understand elongator tRNA(Ser) and suppressor tRNA(Sec) identity elements, the respective acceptor-stem helices have been synthesized and crystallized in order to analyse and compare their structures in detail at high resolution. The synthesis, crystallization and preliminary X-ray diffraction results for a seven-base-pair tRNA(Ser) acceptor-stem helix are presented here. Diffraction data were collected to 1.

Solution structure of a novel C2-symmetrical bifunctional bicyclic inhibitor based on SFTI-1.

A novel bifunctional bicyclic inhibitor has been created that combines features both from the Bowman-Birk inhibitor (BBI) proteins, which have two distinct inhibitory sites, and from sunflower trypsin inhibitor-1 (SFTI-1), which has a compact bicyclic structure. The inhibitor was designed by fusing together a pair of reactive loops based on a sequence derived from SFTI-1 to create a backbone-cyclized disulfide-bridged 16-mer peptide. This peptide has two symmetrically spaced trypsin binding sites.

The conserved P1' Ser of Bowman-Birk-type proteinase inhibitors is not essential for the integrity of the reactive site loop.

The isolated reactive site beta-hairpin loop of Bowman-Birk-type proteinase inhibitors has become a widely studied proteinomimetic because it retains the three-dimensional structure and much of the inhibitory potency of the corresponding region of the complete protein. Here we analyse the role of the P1' Ser residue which is highly conserved and intramolecularly hydrogen bonded in the complete proteins. A combined kinetic and structural analysis of variant proteinomimetic peptides demonstrates that the hydrogen-bond potential of the side-chain oxygen atom of the P1' Ser is not essential for the integrity of the reactive site loop and that it provides only a small contribution to the trypsin affinity and no apparent contribution to the stability against tryptic turnover.

The structural basis of a conserved P2 threonine in canonical serine proteinase inhibitors.

Bowman-Birk inhibitors (BBIs) are a well-studied family of canonical inhibitor proteins of serine proteinases. In nature, the active region of BBIs possesses a highly conserved Thr at the P2 position. The importance of this residue has been reemphasized by synthetic BBI reactive site loop proteinomimetics.

Peptide mimics of the Bowman-Birk inhibitor reactive site loop.

Bowman-Birk Inhibitors (BBIs) are small highly cross-linked proteins that typically display an almost symmetrical "double-headed" structure. Each "head" contains an independent proteinase binding domain. The realization that one BBI molecule could form a 1:1:1 complex with two enzymes led early workers to dissect this activity.

A conserved cis peptide bond is necessary for the activity of Bowman-Birk inhibitor protein.

The Bowman-Birk inhibitor (BBI) family of protease inhibitors has an inhibitory region comprising a disulfide-linked nine-residue loop that adopts the characteristic canonical motif found in many serine protease inhibitors. A unique feature of the BBI loop is the presence of a cis peptide bond at the edge of the inhibitory loop. BBI-related protein fragments that encapsulate this loop retain the structure and inhibitory activity of the parent protein.

The (1)H-NMR solution structure of the antitryptic core peptide of Bowman-Birk inhibitor proteins: a minimal canonical loop.

Bowman-Birk inhibitor (BBI) proteins contain an inhibitory motif comprising a disulfide-bonded sequence that interacts with serine proteinases. Recently, a small 14-residue peptide from sunflowers (SFTI-1), which has potent anti-trypsin activity, has been found to have the same motif. However, this peptide also has an unusual head-to-tail cyclisation.