Modulation of enzyme activity is a powerful means of probing cellular function and can be exploited for diverse applications. Here, we explore a method of enzyme activation where covalent tethering ...of a small molecule to an enzyme can increase catalytic activity (
k
cat
/
K
M
) up to 35-fold. Using a bacterial glycoside hydrolase, BtGH84, we demonstrate how small molecule "fragments", identified as activators in free solution, can be covalently tethered to the protein using Michael-addition chemistry. We show how tethering generates a constitutively-activated enzyme-fragment conjugate, which displays both improved catalytic efficiency and increased susceptibility to certain inhibitor classes. Structure guided modifications of the tethered fragment demonstrate how specific interactions between the fragment and the enzyme influence the extent of activation. This work suggests that a similar approach may be used to modulate the activity of enzymes such as to improve catalytic efficiency or increase inhibitor susceptibility.
Structure-guided tethering of a fragment activator significantly increases enzyme activity.
Structure-guided tethering of a fragment activator significantly increases enzyme activity.
Modulation of enzyme activity is a powerful means of probing cellular function and can be exploited for ...diverse applications. Here, we explore a method of enzyme activation where covalent tethering of a small molecule to an enzyme can increase catalytic activity (
k
cat
/
K
M
) up to 35-fold. Using a bacterial glycoside hydrolase, BtGH84, we demonstrate how small molecule “fragments”, identified as activators in free solution, can be covalently tethered to the protein using Michael-addition chemistry. We show how tethering generates a constitutively-activated enzyme-fragment conjugate, which displays both improved catalytic efficiency and increased susceptibility to certain inhibitor classes. Structure guided modifications of the tethered fragment demonstrate how specific interactions between the fragment and the enzyme influence the extent of activation. This work suggests that a similar approach may be used to modulate the activity of enzymes such as to improve catalytic efficiency or increase inhibitor susceptibility.
Pulmonary epithelial cell responses can enhance type 2 immunity and contribute to control of nematode infections. An important epithelial product is the collectin Surfactant Protein D (SP-D). We ...found that SP-D concentrations increased in the lung following Nippostrongylus brasiliensis infection; this increase was dependent on key components of the type 2 immune response. We carried out loss and gain of function studies of SP-D to establish if SP-D was required for optimal immunity to the parasite. N. brasiliensis infection of SP-D-/- mice resulted in profound impairment of host innate immunity and ability to resolve infection. Raising pulmonary SP-D levels prior to infection enhanced parasite expulsion and type 2 immune responses, including increased numbers of IL-13 producing type 2 innate lymphoid cells (ILC2), elevated expression of markers of alternative activation by alveolar macrophages (alvM) and increased production of the type 2 cytokines IL-4 and IL-13. Adoptive transfer of alvM from SP-D-treated parasite infected mice into naïve recipients enhanced immunity to N. brasiliensis. Protection was associated with selective binding by the SP-D carbohydrate recognition domain (CRD) to L4 parasites to enhance their killing by alvM. These findings are the first demonstration that the collectin SP-D is an essential component of host innate immunity to helminths.
Decision-makers want to perform risk-based cost-benefit prioritization of security investments. However, strong nonlinearities in the most common physical security performance metric make it ...difficult to use for cost-benefit analysis. This paper extends the definition of risk for security applications and embodies this definition in a new but related security risk metric based on the degree of difficulty an adversary will encounter to successfully execute the most advantageous attack scenario. This metric is compatible with traditional cost-benefit optimization algorithms, and can lead to an objective risk-based cost-benefit method for security investment option prioritization. It also enables decision-makers to more effectively communicate the justification for their investment decisions with stakeholders and funding authorities.
Abstract
Metal additive manufacturing provides a path to optimized component design with significant realized advantages in the medical and aerospace industries. Limitations to expansion to other ...industries, e.g. automotive, and to enabling supply chain relief is the limited number of materials available and the ability to produce material on demand. Current additive manufacturing powder feedstock is produced at large, remote atomization facilities with long lead times. Here we identify a new “on-demand” powder production technology, cold mechanically derived, able to produce non-spherical powder for additive manufacturing, with high efficiency, and wrought equivalent material properties. We analyze the powder flow characteristics and mechanical properties comparing typical gas atomized with the new process demonstrating wrought property equivalence despite power sourcing. This research will enable expansion of additional alloy systems as well as encourage the processing of non-spherical powders to expand the available supply base of new alloys for additive manufacturing.
To perform an updated systematic review of the anatomy, biomechanics, function of the anterolateral ligament (ALL), and the clinical outcomes of anterolateral ligament reconstruction (ALLR) when ...performed in conjunction with anterior cruciate ligament reconstruction (ACLR).
A systematic search of the literature was performed by searching PubMed, the Cochrane Library, and Embase with the search phrase anterolateral ligament for articles published from February 2017 to May 2020. Inclusion criteria included studies that evaluated the anatomy, function, or biomechanics of the ALL; surgical technique articles on ALLR; clinical articles reporting outcomes of ALLR; studies published in English; and full-text articles. Exclusion criteria included studies published before February 2017. A subjective synthesis was performed, in which ranges were reported, and individual study data were presented in forest plots.
Overall, 40 articles were included in this systematic review, with 11 articles describing ALL anatomy, 14 articles analyzing ALL function and biomechanics, 7 articles discussing the surgical technique of combined ACLR and ALLR (ACLR/ALLR), and 8 articles describing the clinical outcomes of ACLR/ALLR. The addition of ALLR in combination with ACLR (ACLR+) results in lower graft failure rates for ACLR/ALLR (0.0%–15.7%) when compared with isolated ACLR (I-ACLR) patients (7.4%–21.7%). Three of 5 studies using the Subjective International Knee Documentation Committee score, 2 of 5 studies using the Lysholm score, and 1 of 2 studies using the Tegner score reported significantly better scores at latest follow-up among ACLR+ patients compared with I-ACLR (P < .05).
The ALL acts as a secondary stabilizer to the anterior cruciate ligament and helps resist internal knee rotation and anterior tibial translation. Based on the current literature, combined ACLR with ALLR may result in lower graft failure rates and improved patient-reported outcomes when compared with I-ACLR in patients with specific indications, although several studies have shown equivalent outcomes between these 2 cohorts.
The contents of this review provide great insight for orthopaedic surgeons who are performing ACLR and considering additional procedures to increase overall knee stability and decrease likeliness for re-rupture. The postoperative functional and clinical outcomes shown in patients undergoing ACLR+ compared with I-ACLR should be given proper consideration when evaluating available treatment courses.
The ChemCam instrument suite on the Mars Science Laboratory (MSL) rover
Curiosity
provides remote compositional information using the first laser-induced breakdown spectrometer (LIBS) on a planetary ...mission, and provides sample texture and morphology data using a remote micro-imager (RMI). Overall, ChemCam supports MSL with five capabilities: remote classification of rock and soil characteristics; quantitative elemental compositions including light elements like hydrogen and some elements to which LIBS is uniquely sensitive (e.g., Li, Be, Rb, Sr, Ba); remote removal of surface dust and depth profiling through surface coatings; context imaging; and passive spectroscopy over the 240–905 nm range. ChemCam is built in two sections: The mast unit, consisting of a laser, telescope, RMI, and associated electronics, resides on the rover’s mast, and is described in a companion paper. ChemCam’s body unit, which is mounted in the body of the rover, comprises an optical demultiplexer, three spectrometers, detectors, their coolers, and associated electronics and data handling logic. Additional instrument components include a 6 m optical fiber which transfers the LIBS light from the telescope to the body unit, and a set of onboard calibration targets. ChemCam was integrated and tested at Los Alamos National Laboratory where it also underwent LIBS calibration with 69 geological standards prior to integration with the rover. Post-integration testing used coordinated mast and instrument commands, including LIBS line scans on rock targets during system-level thermal-vacuum tests. In this paper we describe the body unit, optical fiber, and calibration targets, and the assembly, testing, and verification of the instrument prior to launch.
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