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Aminoacyl-tRNA synthetases (ARSs) have evolved to acquire various additional domains. These domains allow ARSs to communicate with other cellular proteins in order to promote non-translational functions. Vertebrate cytoplasmic isoleucyl-tRNA synthetases (IARS1s) have an uncharacterized unique domain, UNE-I. Here, we present the crystal structure of the chicken IARS1 UNE-I complexed with glutamyl-tRNA synthetase 1 (EARS1). UNE-I consists of tandem ubiquitin regulatory X (UBX) domains that interact with a distinct hairpin loop on EARS1 and protect its neighboring proteins in the multi-synthetase complex from degradation. Phosphomimetic mutation of the two serine residues in the hairpin loop releases IARS1 from the complex. IARS1 interacts with BRCA1 in the nucleus, regulates its stability by inhibiting ubiquitylation via the UBX domains, and controls DNA repair function.

The CRISPR-Cas type III-E RNA-targeting effector complex gRAMP/Cas7-11 is associated with a caspase-like protein (TPR-CHAT/Csx29) to form Craspase (CRISPR-guided caspase). Here, we use cryo-electron microscopy snapshots of Craspase to explain its target RNA cleavage and protease activation mechanisms. Target-guide pairing extending into the 5' region of the guide RNA displaces a gating loop in gRAMP, which triggers an extensive conformational relay that allosterically aligns the protease catalytic dyad and opens an amino acid side-chain-binding pocket. We further define Csx30 as the endogenous protein substrate that is site-specifically proteolyzed by RNA-activated Craspase. This protease activity is switched off by target RNA cleavage by gRAMP and is not activated by RNA targets containing a matching protospacer flanking sequence. We thus conclude that Craspase is a target RNA-activated protease with self-regulatory capacity., Green Open Access added to TU Delft Institutional Repository 'You share, we take care!' - Taverne project https://www.openaccess.nl/en/you-share-we-take-care Otherwise as indicated in the copyright section: the publisher is the copyright holder of this work and the author uses the Dutch legislation to make this work public., BN/Stan Brouns Lab, BT/Environmental Biotechnology

Prokaryotes adapt to challenges from mobile genetic elements by integrating spacers derived from foreign DNA in the CRISPR array1. Spacer insertion is carried out by the Cas1–Cas2 integrase complex2–4. A substantial fraction of CRISPR–Cas systems use a Fe–S cluster containing Cas4 nuclease to ensure that spacers are acquired from DNA flanked by a protospacer adjacent motif (PAM)5,6 and inserted into the CRISPR array unidirectionally, so that the transcribed CRISPR RNA can guide target searching in a PAM-dependent manner. Here we provide a high-resolution mechanistic explanation for the Cas4-assisted PAM selection, spacer biogenesis and directional integration by type I-G CRISPR in Geobacter sulfurreducens, in which Cas4 is naturally fused with Cas1, forming Cas4/Cas1. During biogenesis, only DNA duplexes possessing a PAM-embedded 3′-overhang trigger Cas4/Cas1–Cas2 assembly. During this process, the PAM overhang is specifically recognized and sequestered, but is not cleaved by Cas4. This ‘molecular constipation’ prevents the PAM-side prespacer from participating in integration. Lacking such sequestration, the non-PAM overhang is trimmed by host nucleases and integrated to the leader-side CRISPR repeat. Half-integration subsequently triggers PAM cleavage and Cas4 dissociation, allowing spacer-side integration. Overall, the intricate molecular interaction between Cas4 and Cas1–Cas2 selects PAM-containing prespacers for integration and couples the timing of PAM processing with the stepwise integration to establish directionality., Accepted Author Manuscript, BN/Stan Brouns Lab, BT/Bioprocess Engineering, BN/Technici en Analisten

The isothermal compressibility and correlation length of supercooled water obtained from small-angle X-ray scattering (SAXS) were analyzed by fits based on an apparent power-law in the temperature range from 280 K down to the temperature of maximum compressibility at 229 K. Although the increase in thermodynamic response functions is not towards a critical point, it is still possible to obtain an apparent power law all the way to the maximum values with best-fit exponents of gamma = 0.40 +/- 0.01 for the isothermal compressibility and nu = 0.26 +/- 0.03 for the correlation length. The ratio between these exponents is close to a value of approximate to 0.5, as expected for a critical point, indicating the proximity of a potential second critical point. Comparison of gamma obtained from experiment with molecular dynamics simulations on the iAMOEBA water model shows that it would be located at pressures in the neighborhood of 1 kbar. The high value and sharpness of the compressibility maximum observed in the experiment are not reproduced by any of the existing classical water models, thus inviting further development of simulation models of water., QC 20190125

The isothermal compressibility and correlation length of supercooled water obtained from small-angle X-ray scattering (SAXS) were analyzed by fits based on an apparent power-law in the temperature range from 280 K down to the temperature of maximum compressibility at 229 K. Although the increase in thermodynamic response functions is not towards a critical point, it is still possible to obtain an apparent power law all the way to the maximum values with best-fit exponents of gamma = 0.40 +/- 0.01 for the isothermal compressibility and nu = 0.26 +/- 0.03 for the correlation length. The ratio between these exponents is close to a value of approximate to 0.5, as expected for a critical point, indicating the proximity of a potential second critical point. Comparison of gamma obtained from experiment with molecular dynamics simulations on the iAMOEBA water model shows that it would be located at pressures in the neighborhood of 1 kbar. The high value and sharpness of the compressibility maximum observed in the experiment are not reproduced by any of the existing classical water models, thus inviting further development of simulation models of water.

Caupin et al. have raised several issues regarding our recent paper on maxima in thermodynamic response and correlation functions in deeply supercooled water. We show that these issues can be addressed without affecting the conclusion of the paper., QC 201805131

Caupin et al have raised several issues regarding our recent paper on maxima in thermodynamic response and correlation functions in deeply supercooled water. We show that these issues can be addressed without affecting the conclusion of the paper.

Femtosecond x-ray laser pulses were used to probe micrometer-sized water droplets that were cooled down to 227 kelvin in vacuum. Isothermal compressibility and correlation length were extracted from x-ray scattering at the low-momentum transfer region. The temperature dependence of these thermodynamic response and correlation functions shows maxima at 229 kelvin for water and 233 kelvin for heavy water. In addition, we observed that the liquids undergo the fastest growth of tetrahedral structures at similar temperatures. These observations point to the existence of a Widom line, defined as the locus of maximum correlation length emanating from a critical point at positive pressures in the deeply supercooled regime. The difference in the maximum value of the isothermal compressibility between the two isotopes shows the importance of nuclear quantum effects., QC 20180116

Femtosecond x-ray laser pulses were used to probe micrometer-sized water droplets that were cooled down to 227 kelvin in vacuum. Isothermal compressibility and correlation length were extracted from x-ray scattering at the low-momentum transfer region. The temperature dependence of these thermodynamic response and correlation functions shows maxima at 229 kelvin for water and 233 kelvin for heavy water. In addition, we observed that the liquids undergo the fastest growth of tetrahedral structures at similar temperatures. These observations point to the existence of a Widom line, defined as the locus of maximum correlation length emanating from a critical point at positive pressures in the deeply supercooled regime. The difference in the maximum value of the isothermal compressibility between the two isotopes shows the importance of nuclear quantum effects.