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Conformational states dynamically populated by a kinase determine its function

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A moving target

Abl kinase is an important signaling protein that is dysregulated in leukemia and other cancers and is the target of inhibitors such as imatinib. Like other kinases, Abl kinase is dynamic, and regulating conformational dynamics is key to regulating activity. Xie et al. used nuclear magnetic resonance to show that the Abl kinase domain interconverts between one active and two inactive states. Imatinib stabilizes an inactive conformation, and several resistance mutations act by destabilizing this conformation. In a construct that includes the regulatory domain, depending on the relative arrangement of the kinase and regulatory domains, the kinase domain is stabilized in either the active state or one of the inhibited states. Understanding the conformational dynamics of kinases can be leveraged to design selective drugs.

Science, this issue p. eabc2754

Structured Abstract

INTRODUCTION

Protein kinases mediate many cell signaling processes. Central to their physiological function is the regulation of their binding and enzymatic activities, which is typically achieved by conformational transitions between active and inactive states. Dysregulation of kinase activity by deletions or mutations often results in disease. Protein kinases are dynamic molecules that intrinsically sample a number of conformational states. However, it has been challenging to experimentally access their conformational ensemble and structurally characterize the discrete conformations associated with distinct activities. Such information could advance our understanding of activation and inhibition mechanisms in this protein family and aid in the development of selective inhibitors.

RATIONALE

We used nuclear magnetic resonance spectroscopy to monitor in atomic-level detail how Abl kinase transitions between distinct conformational states and to elucidate how the conformational ensemble is exploited by mutants, ligands, posttranslational modifications, and inhibitors to regulate the kinase activity and function. We combined structural and energetic approaches to quantitate the contribution of key structural elements such as the activation loop, the Asp-Phe-Gly (DFG) motif, the regulatory spine, and the gatekeeper residue to kinase regulation and provide the mechanistic basis for drug resistance.

RESULTS

We found that the Abl kinase domain interconverts between an active and two, transiently populated, conformational states that adopt discrete inactive structures. There are extensive differences in key structural elements between the conformational states that reveal multiple intrinsic regulatory mechanisms. The small energy difference between active and inactive states allows oncogenic mutations in the regulatory spine or the gatekeeper position to counteract inhibitory mechanisms and constitutively activate the kinase. By capturing and structurally characterizing the conformational state to which the cancer drug imatinib selectively binds, we explain a number of drug-resistance variants isolated in patients. These mutants confer resistance by depleting, through various mechanisms, the conformation to which imatinib binds. To determine the basis for allosteric regulation, we studied a construct that includes the kinase domain and the regulatory domains and that can adopt an assembled and an extended conformation. In the assembled conformation, in which the regulatory domains dock onto the back of the kinase domain, one of the inactive states is selectively stabilized, thereby suppressing catalytic activity. In the extended conformation, wherein the regulatory domains dock on top of the N-lobe, the inactive state is eliminated, thus explaining the increased leukemogenic activity associated with this conformational state. Only one of the detected inactive states appears to be physiologically relevant. The inactive state with no apparent biological function can nevertheless be leveraged for the design of selective inhibitors. Targeting nonphysiological conformational states may be an effective strategy in the design of drugs with increased selectivity and reduced selection pressure for the occurrence of drug-resistance mutations. Although the structure of inactive states can, in principle, vary considerably among kinases, structural comparison of the Abl inactive states with those previously determined for other kinases reveals that there may be a limited number of structurally divergent inactive states intrinsic to kinases.

CONCLUSION

Our data demonstrate that the detection and structural characterization of the distinct conformational states populated by a kinase, coupled to the energetic dissection of the contribution of key structural elements to the selective stability of these states, are essential to advance our understanding of the mechanisms underpinning kinase regulation and function. The approaches presented here can be extended to other kinases to characterize transiently populated conformational states, with the goal of revealing the full repertoire of regulatory and drug-resistance mechanisms in the kinome.

Transitions of Abl kinase between conformational states.

The Abl kinase domain adopts predominantly (~90%) an active state in solution, but it transiently switches between two low-populated (~5%) states that adopt distinct inactive conformations. Key structural elements that rearrange in the various states are highlighted. The conformational equilibrium is exploited by physiological and pathological stimuli to alter the function of Abl.

Abstract

Protein kinases intrinsically sample a number of conformational states with distinct catalytic and binding activities. We used nuclear magnetic resonance spectroscopy to describe in atomic-level detail how Abl kinase interconverts between an active and two discrete inactive structures. Extensive differences in key structural elements between the conformational states give rise to multiple intrinsic regulatory mechanisms. The findings explain how oncogenic mutants can counteract inhibitory mechanisms to constitutively activate the kinase. Energetic dissection revealed the contributions of the activation loop, the Asp-Phe-Gly (DFG) motif, the regulatory spine, and the gatekeeper residue to kinase regulation. Characterization of the transient conformation to which the drug imatinib binds enabled the elucidation of drug-resistance mechanisms. Structural insight into inactive states highlights how they can be leveraged for the design of selective inhibitors.

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After historic OSIRIS-REx asteroid sample collection, Lockheed Martin VP Lisa Callahan will join us at TC Sessions: Space

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This December 16 and 17, we’re hosting our very first TC Sessions: Space event. It’ll be a virtual, live-streamed two-day show, including conversations with some of the best and brightest in the space industry. We’re thrilled to be hosting Lisa Callahan, Vice President and General Manager of Commercial Civil Space at Lockheed Martin, at the event, and she’ll join us to discuss her company’s history-making work in robotic space exploration – including the asteroid mining sample collection at asteroid Bennu that happened today – as well as the future of human space exploration.

Callahan’s work at Lockheed covers all the work they do to support NASA and other civil exploration efforts of space, including both robotic and human transportation and science investigations. That includes OSIRIS-REx, the asteroid study and sample return mission that earlier today made a historic descent to the surface of rocky solar system visitor Bennu, an asteroid that’s over 200 thousand miles from Earth.

OSIRIS-REx already made plenty of history, including becoming the closest orbit to an asteroid ever conducted by a spacecraft. But today it topped all of that with a ‘tap-and-go’ descent to the rocky surface, scooping samples that it will now attempt to return to Earth for direct study by scientists. That’s exactly the kind of ambitious extra-planetary robotic research that Callahan and her division at Lockheed have made possible with their work in advanced spacecraft and robotics design.

Callahan is also directly involved in NASA’s plans to return humans to the surface of the Moon – including sending a woman on a lunar landing mission for the first time ever. Lockheed Martin is the manufacturing partner for NASA’s Orion lander, which will transport the first American woman and the next American man to the Moon for their historic mission in 2024.

We’ll talk about what these achievements mean for the space industry, and the future of space exploration – and human spaceflight – in December with Callahan.

You can get Early-Bird tickets right now, and save $150 before prices go up on November 13 — and you can even get a fifth person free if you bring a group of four from your company. Special discounts for current members of the government/military/nonprofit and student tickets are also available directly on the website. And if you are an early-stage space startup looking to get exposure to decision makers, you can even exhibit for the day for just $2,000.

Is your company interested in partnering at TC Sessions: Space 2020? Click here to talk with us about available opportunities.

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How to ‘watch’ NASA’s OSIRIS-REx snatch a sample from near-Earth asteroid Bennu

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NASA’s OSIRIS-REx probe is about to touch down on an asteroid for a smash-and-grab mission, and you can follow its progress live — kind of. The craft is scheduled to perform its collection operation this afternoon, and we’ll know within minutes if all went according to plan.

OSIRIS-REx, which stands for Origins Spectral Interpretation Resource Identification Security – Regolith Explorer, was launched in September of 2016 and since arriving at its destination, the asteroid Bennu, has performed a delicate dance with it, entering an orbit so close it set records.

Today is the culmination of the team’s efforts, the actual “touch and go” or TAG maneuver that will see the probe briefly land on the asteroid’s surface and suck up some of its precious space dust. Just a few seconds later, once sampling is confirmed, the craft will jet upwards again to escape Bennu and begin its journey home.

Image Credits: NASA

Image Credits: NASA

While there won’t be live HD video of the whole attempt, NASA will be providing both a live animation of the process informed by OSIRIS-REx’s telemetry, and presumably any good images that are captured as it descends.

We know for certain this is both possible and very cool because Japan’s Hayabusa-2 asteroid mission did something very similar last year, but with the added complexity (and coolness) of firing a projectile into the surface to stir things up and get a more diverse sample.

NASA’s coverage starts at 2 PM Pacific, and the touchdown event is planned to take place an hour or so later, at 3:12 if all goes according to plan. You can watch the whole thing take place in simulation at this Twitch feed, which will be updated live, but NASA TV will also have live coverage and commentary on its YouTube channel. Images may come back from the descent and collection, but they’ll be delayed (it’s hard sending lots of data over a million-mile gap) so if you want the latest, listen closely to the NASA feeds.

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Intel is providing the smarts for the first satellite with local AI processing on board

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Intel detailed today its contribution to PhiSat-1, a new tiny small satellite that was launched into sun-synchronous orbit on September 2. PhiSat-1 has a new kind of hyperspectral-thermal camera on board, and also includes a Movidius Myriad 2 Vision Processing Unit. That VPU is found in a number of consumer devices on Earth, but this is its first trip to space – and the first time it’ll be handling large amounts of local data, saving researchers back on Earth precious time and satellite downlink bandwidth.

Specifically, the AI on board the PhiSat-1 will be handling automatic identification of cloud cover – images where the Earth is obscured in terms of what the scientists studying the data actually want to see. Getting rid of these images before they’re even transmitted means that the satellite can actually realize a bandwidth savings of up to 30%, which means more useful data is transmitted to Earth when it is in range of ground stations for transmission.

The AI software that runs on the Intel Myriad 2 on PhiSat-1 was created by startup Ubotica, which worked with the hardware maker behind the hyperspectral camera. It also had to be tuned to compensate for the excess exposure to radiation, though a bit surprisingly testing at CERN found that the hardware itself didn’t have to be modified in order to perform within the standards required for its mission.

Computing at the edge takes on a whole new meaning when applied to satellites on orbit, but it’s definitely a place where local AI makes a ton of sense. All the same reasons that companies seek to handle data processing and analytics at the site of sensors hear on Earth also apply in space – but magnified exponentially in terms of things like network inaccessibility and quality of connections, so expect to see a lot more of this.

PhiSat-1 was launched in September as part of Arianspace’s first rideshare demonstration mission, which it aims to use to show off its ability to offer launch services to smaller startups for smaller payloads at lower costs.

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