Slapstick and Self-Reflexivity in George Herriman’s ‘Krazy Kat’

George Herriman’s newspaper comic strip ‘Krazy Kat’ has been cited by cartoonists and intellectuals over the years as a surrealistic masterpiece. It has been read as psychoanalytical, racial, socio-political and religious allegory. Yet attempts to define the strip do not do justice to its multifaceted nature, and its self-reflexivity. This essay examines how ‘Krazy Kat’ simultaneously invites and avoids interpretation, and how it fits into the slapstick tradition

Agon, conflict and dissent:Elfriede Jelinek's Ein Sportstück and its stagings by Einar Schleef and Just a Must Theatre

This article explores the directorial approaches of Einar Schleef’s radically choric staging of Elfriede Jelinek’s play Ein Sportstück at the Burgtheater (1998) and Vanda Butkovic’s English language premiere of Sports Play (Just a Must, 2012) in the context of the ancient Greek concept of the agon and the conflict between group and individual. A reworking of agon and conflict is not only central to Jelinek’s text but even more so to Schleef’s dramaturgy. This latter is read in terms of Schleef’s theory about the disappearance of the chorus from German theatre and woman from the centre of the conflict. While in terms of its content Jelinek’s play critiques society’s valorization of sportive agon and its relationship to collective violence and warfare, on the level of theatrical communication both Schleef’s and Butkovic’s stagings show that the play revives an agonistic tradition creating a public space for political dissent

Progress in rational methods of cryoprotection in macromolecular crystallography

Measurements of the average thermal contractions (294→72 K) of 26 different cryosolutions are presented and discussed in conjunction with other recent advances in the rational design of protocols for cryogenic cooling in macromolecular crystallography

Change of tRNA identity leads to a divergent orthogonal histidyl-tRNA synthetase/tRNAHis pair

Mature tRNAHis has at its 5′-terminus an extra guanylate, designated as G−1. This is the major recognition element for histidyl-tRNA synthetase (HisRS) to permit acylation of tRNAHis with histidine. However, it was reported that tRNAHis of a subgroup of α-proteobacteria, including Caulobacter crescentus, lacks the critical G−1 residue. Here we show that recombinant C. crescentus HisRS allowed complete histidylation of a C. crescentus tRNAHis transcript (lacking G−1). The addition of G−1 did not improve aminoacylation by C. crescentus HisRS. However, mutations in the tRNAHis anticodon caused a drastic loss of in vitro histidylation, and mutations of bases A73 and U72 also reduced charging. Thus, the major recognition elements in C. crescentus tRNAHis are the anticodon, the discriminator base and U72, which are recognized by the divergent (based on sequence similarity) C. crescentus HisRS. Transplantation of these recognition elements into an Escherichia coli tRNAHis template, together with addition of base U20a, created a competent substrate for C. crescentus HisRS. These results illustrate how a conserved tRNA recognition pattern changed during evolution. The data also uncovered a divergent orthogonal HisRS/tRNAHis pair

Dissecting conformational contributions to glycosidase catalysis and inhibition

Glycoside hydrolases (GHs) are classified into >100 sequence-based families. These enzymes process a wide variety of complex carbohydrates with varying stereochemistry at the anomeric and other ring positions. The shapes that these sugars adopt upon binding to their cognate GHs, and the conformational changes that occur along the catalysis reaction coordinate is termed the conformational itinerary. Efforts to define the conformational itineraries of GHs have focussed upon the critical points of the reaction: substrate-bound (Michaelis), transition state, intermediate (if relevant) and product-bound. Recent approaches to defining conformational itineraries that marry X-ray crystallography of enzymes bound to ligands that mimic the critical points, along with advanced computational methods and kinetic isotope effects are discussed

Radiation damage in room-temperature data acquisition with the PILATUS 6M pixel detector

Observations of the dose-rate effect in continuous X-ray diffraction data acquisition at room temperature are presented

Temperature-dependent macromolecular X-ray crystallography

The dynamical behaviour of crystalline macromolecules and their surrounding solvent as a function of cryo-temperature is reviewed

Sensitive Spectroscopic Detection of Large and Denatured Protein Aggregates in Solution by Use of the Fluorescent Dye Nile Red

The fluorescent dye Nile red was used as a probe for the sensitive detection of large, denatured aggregates of the model protein β-galactosidase (E. coli) in solution. Aggregates were formed by irreversible heat denaturation of β-galactosidase below and above the protein’s unfolding temperature of 57.4°C, and the presence of aggregates in heated solutions was confirmed by static light scattering. Interaction of Nile red with β-galactosidase aggregates led to a shift of the emission maximum (λmax) from 660 to 611 nm, and to an increase of fluorescence intensity. Time-resolved fluorescence and fluorescence correlation spectroscopy (FCS) measurements showed that Nile red detected large aggregates with hydrodynamic radii around 130 nm. By steady-state fluorescence measurements, it was possible to detect 1 nM of denatured and aggregated β-galactosidase in solution. The comparison with size exclusion chromatography (SEC) showed that native β-galactosidase and small aggregates thereof had no substantial effect on the fluorescence of Nile red. Large aggregates were not detected by SEC, because they were excluded from the column. The results with β-galactosidase demonstrate the potential of Nile red for developing complementary analytical methods that overcome the size limitations of SEC, and can detect the formation of large protein aggregates at early stages