Identification of the C2-1H histidine NMR resonances in chloramphenicol acetyltransferase by a 13C-1H heteronuclear multiple quantum coherence method

AbstractChloramphenicol acetyltransferase (CAT) was used to assess the feasibility of study of specific proton resonances in an enzyme of overall molecular mass 75000. [ring2-13C]Histidine was selectively incorporated into the type III chloramphenicol acetyltransferase (CATIII) using a histidine auxotroph of E. coli. Heteronuclear multiple and single quantum experiments were used to select the C2 protons in the histidyl imidazole ring. One- and two-dimensional spectra revealed six signals out of a total of seven histidine residues in CATIII. pH titration, chemical modification and ligand binding were used to demonstrate that the signal from H195, the histidine at the active site, is not among those observed. Nevertheless, this work demonstrates that selective isotopic enrichment and multiple quantum coherence techniques can be used to distinguish proton resonances in a protein of high molecular mass

Biophysical and functional characterization of hippocalcin mutants responsible for human dystonia

Dystonia is a neurological movement disorder that forces the body into twisting, repetitive movements or sometimes painful abnormal postures. With the advent of next-generation sequencing technologies, the homozygous mutations T71N and A190T in the neuronal calcium sensor (NCS) hippocalcin were identified as the genetic cause of primary isolated dystonia (DYT2 dystonia). However, the effect of these mutations on the physiological role of hippocalcin has not yet been elucidated. Using a multidisciplinary approach, we demonstrated that hippocalcin oligomerises in a calcium-dependent manner and binds to voltage-gated calcium channels. Mutations T71N and A190T in hippocalcin did not affect stability, calcium-binding affinity or translocation to cellular membranes (Ca2+/myristoyl switch). We obtained the first crystal structure of hippocalcin and alignment with other NCS proteins showed significant variability in the orientation of the C-terminal part of the molecule, the region expected to be important for target binding. We demonstrated that the disease-causing mutations did not affect the structure of the protein, however both mutants showed a defect in oligomerisation. In addition, we observed an increased calcium influx in KCl-depolarised cells expressing mutated hippocalcin, mostly driven by N-type voltage-gated calcium channels. Our data demonstrate that the dystonia-causing mutations strongly affect hippocalcin cellular functions which suggest a central role for perturbed calcium signalling in DYT2 dystonia

Properties of collagen/sodium alginate hydrogels for bioprinting of skin models

3D printing technology has great potential for the reconstruction of human skin. However, the reconstructed skin has some differences from natural skin, largely because the hydrogel used does not have the appropriate biological and physical properties to allow healing and regeneration. This study examines the swelling, degradability, microstructure and biological properties of Collagen/Sodium Alginate (Col/SA) hydrogels of differing compositions for the purposes of skin printing. Increasing the content of sodium alginate causes the hydrogel to exhibit stronger mechanical and swelling properties, a faster degradation ratio, smaller pore size, and less favorable biological properties. An optimal 1% collagen hydrogel was used to print bi-layer skin in which fibroblasts and keratinocytes showed improved spreading and proliferation as compared to other developed formulations. The Col/SA hydrogels presented suitable tunability and properties to be used as a bioink for bioprinting of skin aiming at finding applications as 3D models for wound healing research.This research was funded by the National Key R&D Program of China (2018YFE0207900), and People's Liberation Army (BWS17J036, 18-163-13-ZT-003-011-01) and the National Natural Science Foundation of China (51835010 and 51375371), and Xi’an Science and Technology Plan Project (21ZCZZHXJS-QCY6-0012). Shaanxi Science and Technology Project (2022KXJ-147). Thanks to Shi Changquan and Yang Chuncheng of Shaanxi Ketao-AM Technology Co., Ltd. for their technical support for printing equipment

An ECG-on-Chip with 535-nW/Channel Integrated Lossless Data Compressor for Wireless Sensors

This paper presents a low-power ECG recording system-on-chip (SoC) with on-chip low-complexity lossless ECG compression for data reduction in wireless/ambulatory ECG sensor devices. The chip uses a linear slope predictor for data compression, and incorporates a novel low-complexity dynamic coding-packaging scheme to frame the prediction error into fixed-length 16-bit format. The proposed technique achieves an average compression ratio of 2.25x on MIT/BIH ECG database. Implemented in a standard 0.35 um process, the compressor uses 0.565K gates/channel occupying 0.4 mm2 for four channels, and consumes 535 nW/channel at 2.4 V for ECG sampled at 512 Hz. Small size and ultra-low power consumption makes the proposed technique suitable for wearable ECG sensor applications

Theoretical investigation of structural, energetic and electronic properties of titanate pyrochlores

Ab initio total energy calculations using the plane-wave pseudopotential method based on density functional theory were carried out to investigate the structural, energetic and electronic properties of A2Ti2O7 (A =  La, Gd and Yb) pyrochlores. It turned out that the formation energies of antisite defects are not linearly dependent on the ratio of the cation radii, and, for the three compositions, the cation antisite formation energy is largest for Gd2Ti2O7 pyrochlore. It was indicated that Gd2Ti2O7 compound is the least likely to form defect fluorite structure, which gives rise to the least resistance to radiation-induced amorphization. DOS analysis showed that stronger interaction exists in the Gd2Ti2O7 compound, and its electronic structure is very different from that of La2Ti2O7 and Yb2Ti2O7. Our calculations suggested that the electronic structure of the A cation and bond type should be taken into account when explaining the response behavior of A2Ti2O7 (A =  La, Gd, Yb) pyrochlores to ion irradiation-induced amorphization.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/58124/2/cm7_34_346203.pd