Methods for expression of recombinant proteins using a Pichia pastoris cell-free system

Cell‐free protein synthesis is a powerful tool for engineering biology and has been utilized in many diverse applications, from biosensing and protein prototyping to biomanufacturing and the design of metabolic pathways. By exploiting host cellular machinery decoupled from cellular growth, proteins can be produced in vitro both on demand and rapidly. Eukaryotic cell‐free platforms are often neglected due to perceived complexity and low yields relative to their prokaryotic counterparts, despite providing a number of advantageous properties. The yeast Pichia pastoris (also known as Komagataella phaffii) is a particularly attractive eukaryotic host from which to generate cell‐free extracts, due to its ability to grow to high cell densities with high volumetric productivity, genetic tractability for strain engineering, and ability to perform post‐translational modifications. Here, we describe methods for conducting cell‐free protein synthesis using P. pastoris as the host, from preparing the cell lysates to protocols for both coupled and linked transcription‐translation reactions. By providing these methodologies, we hope to encourage the adoption of the platform by new and experienced users alike

Theoretical analysis of electronic processes occurring during ultrafast demagnetization of cobalt triggered by X-ray photons tuned to Co L3_3 resonance

Magnetization dynamics triggered with ultrashort laser pulses has been attracting significant attention, with strong focus on the dynamics excited by VIS/NIR pulses. Only recently, strong magnetic response in solid materials induced by intense X-ray pulses from free-electron lasers (FELs) has been observed. The exact mechanisms that trigger the X-ray induced demagnetization are not yet fully understood. They are subject of on-going experimental and theoretical investigations. Here, we present a theoretical analysis of electronic processes occurring during demagnetization of Co multilayer system irradiated by X-ray pulses tuned to L$_3$-absorption edge of cobalt. We show that, similarly as in the case of X-ray induced demagnetization at M-edge of Co, electronic processes play a predominant role in the demagnetization until the pulse fluence does not exceed the structural damage threshold. The impact of electronic processes can reasonably well explain the available experimental data, without a need to introduce the mechanism of stimulated elastic forward scattering.Comment: 10 pages, 4 figures (7 panels), 57 references; pdfRevTeX class; double column formatting; two appendices and 18 references added; author-created version submitted to and accepted in Physical Review B journal. arXiv admin note: text overlap with arXiv:2202.1384

Direct 2D spatial coherence determination using the Fourier analysis method Multi parameter characterization of the P04 beamline at PETRA III

We present a systematic 2D spatial-coherence analysis of the soft-X-ray beamline P04 at PETRA III for various beamline configurations. The influence of two different beam-defining apertures on the spatial coherence properties of the beam is discussed and optimal conditions for coherence-based experiments are found. A significant degradation of the spatial coherence in the vertical direction has been measured and sources of this degradation are identified and discussed. The Fourier-analysis method, which gives fast and simple access to the 2D spatial coherence function of the X-ray beam, is used for the experiment. Here, we exploit the charge scattering of a disordered nanodot sample allowing the use of arbitrary X-ray photon energies with this method

A Low-Diversity Microbiota Inhabits Extreme Terrestrial Basaltic Terrains and Their Fumaroles : Implications for the Exploration of Mars

A major objective in the exploration of Mars is to test the hypothesis that the planet hosted life. Even in the absence of life, the mapping of habitable and uninhabitable environments is an essential task in developing a complete understanding of the geological and aqueous history of Mars and, as a consequence, understanding what factors caused Earth to take a different trajectory of biological potential. We carried out the aseptic collection of samples and comparison of the bacterial and archaeal communities associated with basaltic fumaroles and rocks of varying weathering states in Hawai'i to test four hypotheses concerning the diversity of life in these environments. Using high-throughput sequencing, we found that all these materials are inhabited by a low-diversity biota. Multivariate analyses of bacterial community data showed a clear separation between sites that have active fumaroles and other sites that comprised relict fumaroles, unaltered, and syn-emplacement basalts. Contrary to our hypothesis that high water flow environments, such as fumaroles with active mineral leaching, would be sites of high biological diversity, alpha diversity was lower in active fumaroles compared to relict or nonfumarolic sites, potentially due to high-temperature constraints on microbial diversity in fumarolic sites. A comparison of these data with communities inhabiting unaltered and weathered basaltic rocks in Idaho suggests that bacterial taxon composition of basaltic materials varies between sites, although the archaeal communities were similar in Hawai'i and Idaho. The taxa present in both sites suggest that most of them obtain organic carbon compounds from the atmosphere and from phototrophs and that some of them, including archaeal taxa, cycle fixed nitrogen. The low diversity shows that, on Earth, extreme basaltic terrains are environments on the edge of sustaining life with implications for the biological potential of similar environments on Mars and their exploration by robots and humans.Peer reviewe

Deep-Sea Exploration of the US Gulf of Mexico with NOAA Ship Okeanos Explorer

Oceanography articles are licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution, and reproduction in any medium or format as long as users cite the materials appropriately (e.g., authors, Oceanography, volume number, issue number, page number[s], figure number[s], and DOI for the article), provide a link to the Creative Commons license, and indicate the changes that were made to the original content

Friction reduction and zero wear for 52100 bearing steel by high‐dose implantation of carbon

Ion implantation of carbon in the AISI 52100 bearing steel yields a distinct reduction in friction and wear. This improvement is strongly dependent on the implanted fluence. The coefficient of friction decreases from 0.6 to 0.2 for doses >1×1018 cm-2 (energy 100 keV) and a wear reduction to nearly ‘‘zero wear’’ was obtainable even under severe wear conditions. The counterpart (unimplanted AISI 52100 steel ball) shows a similar behavior, which demonstrates that the tribological system is totally changed. Mössbauer spectroscopy and x-ray diffraction revealed that hexagonal ¿-carbide is formed on implantation. On the other hand, Rutherford backscattering spectrometry shows that for high doses a large fraction of the implanted carbon is not contained in this carbide

Overview of NASA Finesse (Field Investigations to Enable Solar System Science and Exploration) Science and Exploration Project

NASA's FINESSE (Field Investigations to Enable Solar System Science and Exploration) project was selected as a research team by NASA's Solar System Exploration Research Virtual Institute (SSERVI). SSERVI is a joint Institute supported by NASA's Science Mission Directorate (SMD) and Human Exploration and Operations Mission Directorate (HEOMD). As such, FINESSE is focused on a science and exploration field-based research program to generate strategic knowledge in preparation for human and robotic exploration of other planetary bodies including our Moon, Mars moons Phobos and Deimos, and near-Earth asteroids. FINESSE embodies the philosophy that "science enables exploration and exploration enables science"