Below, explore peer-reviewed journal articles related to ISS National Lab investigations. For a more extensive list of spaceflight-related publications (not limited to ISS National Lab sponsorship), see the International Space Station Research Results Citations on the NASA website.
The attenuation of sedimentation and convection in microgravity can sometimes decrease irregularities formed during macromolecular crystal growth. Current terrestrial protein crystal growth (PCG) capabilities are very different than those used during the Shuttle era and that are currently on the International Space Station (ISS). The focus of this experiment was to demonstrate the use of a commercial off-the-shelf, high throughput, PCG method in microgravity. Using Protein BioSolutions’ microfluidic Plug Maker™/CrystalCard™ system, we tested the ability to grow crystals of the regulator of glucose metabolism and adipogenesis: peroxisome proliferator-activated receptor gamma (apo-hPPAR-γ LBD), as well as several PCG standards. Overall, we sent 25 CrystalCards™ to the ISS, containing ~10,000 individual microgravity PCG experiments in a 3U NanoRacks NanoLab (1U = 103 cm.). After 70 days on the ISS, our samples were returned with 16 of 25 (64%) microgravity cards having crystals, compared to 12 of 25 (48%) of the ground controls. Encouragingly, there were more apo-hPPAR-γ LBD crystals in the microgravity PCG cards than the 1g controls. These positive results hope to introduce the use of the PCG standard of low sample volume and large experimental density to the microgravity environment and provide new opportunities for macromolecular samples that may crystallize poorly in standard laboratories.
Neutron macromolecular crystallography (NMC) is the prevailing method for the accurate determination of the positions of H atoms in macromolecules. As neutron sources are becoming more available to general users, finding means to optimize the growth of protein crystals to sizes suitable for NMC is extremely important. Historically, much has been learned about growing crystals for X-ray diffraction. However, owing to new-generation synchrotron X-ray facilities and sensitive detectors, protein crystal sizes as small as in the nano-range have become adequate for structure determination, lessening the necessity to grow large crystals. Here, some of the approaches, techniques and considerations for the growth of crystals to significant dimensions that are now relevant to NMC are revisited. These include experimental strategies utilizing solubility diagrams, ripening effects, classical crystallization techniques, microgravity and theoretical considerations.
MTAN (5?-methylthioadenosine nucleosidase) catalyzes the hydrolysis of the N-ribosidic bond of a variety of adenosine-containing metabolites. The Helicobacter pylori MTAN (HpMTAN) hydrolyzes 6-amino-6-deoxyfutalosine in the second step of the alternative menaquinone biosynthetic pathway. Substrate binding of the adenine moiety is mediated almost exclusively by hydrogen bonds, and the proposed catalytic mechanism requires multiple proton-transfer events. Of particular interest is the protonation state of residue D198, which possesses a pKa above 8 and functions as a general acid to initiate the enzymatic reaction. In this study we present three corefined neutron/X-ray crystal structures of wild-type HpMTAN cocrystallized with S-adenosylhomocysteine (SAH), Formycin A (FMA), and (3R,4S)-4-(4-Chlorophenylthiomethyl)-1-[(9-deaza-adenin-9-yl)methyl]-3-hydroxypyrrolidine (p-ClPh-Thio-DADMe-ImmA) as well as one neutron/X-ray crystal structure of an inactive variant (HpMTAN-D198N) cocrystallized with SAH. These results support a mechanism of D198 pKa elevation through the unexpected sharing of a proton with atom N7 of the adenine moiety possessing unconventional hydrogen-bond geometry. Additionally, the neutron structures also highlight active site features that promote the stabilization of the transition state and slight variations in these interactions that result in 100-fold difference in binding affinities between the DADMe-ImmA and ImmA analogs.
Huntington's disease is one of nine neurodegenerative diseases caused by a polyglutamine (polyQ)-repeat expansion. An anti-polyQ antigen-binding fragment, MW1 Fab, was crystallized both on Earth and on the International Space Station, a microgravity environment where convection is limited. Once the crystals returned to Earth, the number, size and morphology of all crystals were recorded, and X-ray data were collected from representative crystals. The results generally agreed with previous microgravity crystallization studies. On average, microgravity-grown crystals were 20% larger than control crystals grown on Earth, and microgravity-grown crystals had a slightly improved mosaicity (decreased by 0.03°) and diffraction resolution (decreased by 0.2 Å) compared with control crystals grown on Earth. However, the highest resolution and lowest mosaicity crystals were formed on Earth, and the highest-quality crystal overall was formed on Earth after return from microgravity.
Cerium oxide (CeO2) was prepared using a controlled-precipitation method under microgravity at the International Space Station (ISS). For comparison, ceria was also synthesized under normal-gravity conditions (referred as control). The Brunauer-Emmett-Teller (BET) surface area, pore volume and pore size analysis results indicated that the ceria particles grown in space had lower surface area and pore volume compared to the control samples. Furthermore, the space samples had a broader pore size distribution ranging from 30–600 Å, whereas the control samples consisted of pore sizes from 30–50 Å range. Structural information of the ceria particles were obtained using TEM and XRD. Based on the TEM images, it was confirmed that the space samples were predominantly nano-rods, on the other hand, only nano-polyhedra particles were seen in the control ceria samples. The average particle size was larger for ceria samples synthesized in space. XRD results showed higher crystallinity as well as larger mean crystal size for the space samples. The effect of sodium hydroxide concentration on synthesis of ceria was also examined using 1 M and 3 M solutions. It was found that the control samples, prepared in 1 M and 3 M sodium hydroxide solutions, did not show a significant difference between the two. However, when the ceria samples were prepared in a more basic medium (3 M) under microgravity, a decrease in the particle size of the nano-rods and appearances of nano-polyhedra and spheres were observed.
Calcium-independent phospholipase A 2 β (iPLA 2 β) regulates important physiological processes including inflammation, calcium homeostasis and apoptosis. It is genetically linked to neurodegenerative disorders including Parkinson?s disease. Despite its known enzymatic activity, the mechanisms underlying iPLA 2 β-induced pathologic phenotypes remain poorly understood. Here, we present a crystal structure of iPLA 2 βthat significantly revises existing mechanistic models. The catalytic domains form a tight dimer. They are surrounded by ankyrin repeat domains that adopt an outwardly flared orientation, poised to interact with membrane proteins. The closely integrated active sites are positioned for cooperative activation and internal transacylation. The structure and additional solution studies suggest that both catalytic domains can be bound and allosterically inhibited by a single calmodulin. These features suggest mechanisms of iPLA 2 β cellular localization and activity regulation, providing a basis for inhibitor development. Furthermore, the structure provides a framework to investigate the role of neurodegenerative mutations and the function of iPLA 2 β in the brain.
The International Space Station (ISS) is a marvel of international partnership and engineering. With more than 10 years of assembly led by five space agencies representing 15 countries, the ISS has now been operating and supporting a continuous human presence in space for nearly 20 years, hosting more than 200 people from 18 countries. In addition to serving as a crewed spacecraft and satellite in low Earth orbit (LEO), the ISS is a one-of-a-kind platform for research that takes advantage of the unique space environment to accelerate discovery and explore human limits (1). Research on the ISS helps NASA and international partners reduce the risk of human exploration beyond Earth’s orbit—but it also enables biomedical discovery with direct application to technologies and therapeutics that impact life on Earth. The vision to leverage space for Earth-based benefits was realized when the U.S. Congress designated the ISS as a U.S. National Laboratory in 2005, allowing for increased utilization of the ISS by a broad range of users focused on Earth-based benefits (Fig. 1). In this commentary, we briefly summarize how the ISS and its predecessors paved the way for drug discovery in space, discuss how the ISS National Laboratory is engaged in expanding this research, and provide a perspective on the future industrialization of space to improve medical innovation and drug discovery that benefits life on Earth. We focus on three examples to illustrate the benefits of in-orbit pharmaceutical research: rodent research, tissue chips, and macromolecular crystal growth.
On December 5, 2018, I watched the Falcon 9 rocket launch from Cape Canaveral, Florida, on SpaceX CRS-16 mission. A flash of fire, followed by an unrelenting rumbling sound that crescendoes to a deafening roar, shaking the ground and the air. As I watched the rocket ascending and gradually disappearing from my vision, I prayed for the safe docking of the capsule to the International Space Station (ISS), not just because I wanted to make sure the six astronauts on the ISS get their supply of food and water, but also because I had experiments in the capsule! In collaboration with the International Space Station National Laboratory, the NCI RAS Initiative launched a panel of RAS proteins in various conditions to promote crystal growth in microgravity on the International Space Station (ISS). Our goal was to solve protein structures by X-ray crystallography that we could not solve with crystals grown under Earth’s gravity.
A cost-effective capillary dialysis apparatus (Toledo Capillary Box, TCB) developed for biomacromolecule crystal growth in microgravity and unit gravity environments can provide slow equilibration between the precipitant reservoir and capillary solutions, nurturing growth of neutron-diffraction-quality crystals. Under microgravity conditions, mass transfer of precipitants and biomacromolecules occurs under diffusion-controlled conditions, promoting slow growth and suppressing defect formation. The equilibration of common precipitants (polyethylene glycol and salts such as ammonium sulfate) between capillary and reservoir solutions was measured for capillaries oriented horizontally or vertically with respect to the gravitational field at unit gravity. Precipitants equilibrated less rapidly in the vertical orientation when capillary solution densities were lower than those of the reservoir solutions. A plug filled with agarose gel was introduced in the TCB apparatus for salt precipitants since salts often exhibit relatively high free diffusion. Equilibration of the capillaries with reservoir solutions was significantly delayed for many of the salt precipitants tested. Analytical and semi-analytical models allow the prediction of precipitant equilibration of capillary and reservoir solutions under diffusion-controlled transport and show good agreement with experimental results.