JAXA has long been engaged in the High-Quality Protein Crystal Growth (PCG) experiment on the International Space Station (ISS). This project has undergone extensive development for the last few years. This review describes a general outline of the project and services provided by JAXA.

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Mitogen-activated protein kinase kinase 7 (MAP2K7) regulates stress and inflammatory responses, and is an attractive drug discovery target for serious diseases such as arthritis and cardiac hypertrophy. A microgravity environment improved the crystal quality of MAP2K7 and improved the structural resolution to 1.3 Å. High resolution analysis structurally clarified the two regions, which were undefined in the previous low-resolution analysis and conferred structural insights for producing MAP2K7-specific inhibitors. The hinge region alternatively configures the canonical and atypical conformations. The latter could allow binding MAP2K7 inhibitors with a novel scaffold. The C-terminal region works as a negative regulator with intermolecular association, which implies the ability to produce highly selective MAP2K7 inhibitors.

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Human MTH1 hydrolyzes oxidized nucleoside triphosphates with broad substrate specificity and draws attention as a potential anticancer target. Recently, we determined the high resolution crystal structures of MTH1 and suggested that MTH1 recognizes different substrates via an exchange of the protonation state at Asp119 and Asp120. In order to validate this mechanism, it is essential to observe hydrogen atoms by ultra-high resolution X-ray crystallography and/or neutron crystallography using large high quality crystals. Here we carried out the crystallization of MTH1 in complex with a substrate, 8-oxo-dGTP, under microgravity in the Japanese Experiment Module ‘Kibo’. One of the crystals diffracted to 1.04-Å resolution, which is better than that we reported previously. We carried out bond length analysis of Asp119 and Asp120 using this updated data, which revealed the protonation state based on the bond lengths with higher accuracy and precision.

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Serum albumin is the most prominent plasma protein in the blood circulatory system and is the main transporter of various compounds. Although ligand binding ability and crystal structure of human serum albumin (HSA) have been widely studied, there are only few reports on serum albumins of companion animals, particularly dogs and cats. This review describes the synthesis, physicochemical properties, and crystal structures of recombinant canine serum albumin (rCSA) and feline serum albumin (rFSA). The rCSA and rFSA were produced in a gram quantity by genetic engineering procedure using Pichia yeast. The proteins show identical features to those of the native CSA and FSA derived from canine and feline plasma. Single crystals of rFSA were prepared under a microgravity environment in the International Space Station. The overall structures of rCSA and rFSA closely resemble to that of HSA. These recombinant serum albumins are anticipated for use as a therapeutic reagent that can be exploited in numerous veterinary medicine situations.

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The isoforms of jacalin-related lectins of Pteria penguin PPL3 are known to regulate biomineralization of pearl shell, although the molecular mechanisms are largely unknown. The PPL3 crystal structures were determined partly by utilizing microgravity environments on the International Space Station for three isoforms, namely, PPL3A, PPL3B, and PPL3C. The crystals grown in microgravity environments tended to diffract to higher resolutions. The crystal structures revealed the structure stabilization mechanism of PPL3 isoforms through disulfide bond formations. Also the crystal structures, in combination with docking simulations to calcite, suggested a regulatory mechanism of biomineralization by carbohydrate-binding to the PPL3 isoforms. Additionally, the N-terminal residues of PPL3 isoforms were found in pyroglutamate form in the high-resolution electron densities, which was partly explained by the post-translational modification implied from the discrepancy between amino acid and gene sequences of PPL3 isoforms.

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Dipeptidyl amino peptidase is well known as a drug target of diabetes. Therefore, the existence of many types of DPPs in many organisms has been often overlooked by clinical researchers. Human DPP4 is involved in the inactivation of incretin for the control of blood glucose levels. Meanwhile, some bacterial DPPs are involved in energy metabolism in Non-Fermenting Gram-Negative Rods (NFGNRs). NFGNRs utilize peptides or proteins as energy and carbon sources instead of carbohydrates. Bacterial DPPs are composed of two families (Clan SC S9, Clan PA S46). Clan SC S9 family DPPs are also seen in animals. By contrast, Clan PA S46 family DPPs are only found in bacteria. Additionally, substrate specificity of Clan PA S46 DPPs are completely different from Clan SC S9 DPPs. Considering these features together, bacterial DPPs may be potential target molecules for antimicrobials.

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JCB-SGT, which is the primary crystallization container used in JAXA PCG (Protein Crystal Growth) experiments, is simple, versatile and has a wide range of applications. Various crystallization methods can be applied in JCB-SGT: counter-diffusion method, dialysis method, vapor-diffusion method, osmosis-tube method and batch method. We have supported users of JAXA PCG for nearly 20 years and incrementally improved our technologies to obtain high-quality protein crystals in space. In this review, the outline of JCB-SGT is described and important insights and lessons learned from our experiences in optimizing crystallization conditions are explained. These are helpful for PCG users to optimize their protein samples for experimentation, contributing to both space-based experiments and also to laboratory experiments on the ground.

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CO2 adsorption and desorption experiments were conducted by using zeolite 5A to clarify the dependence of the CO2 removal efficiency of this device on the heater preset temperature in the range of 250–500 ℃ for use onboard the International Space Station. The CO2 concentration was almost independent of the heater preset temperature. High removal efficiency, indicated by the ratio of the exhausted volume of CO2 to that of the absorbed CO2, was obtained in the range of 350–450℃. When the heater preset temperature was higher than 350℃, the CO2 removal efficiency increased owing to the removal of other components. On the contrary, at a heater preset temperature of 500℃ the CO2 removal efficiency was low because the temperature did not reach room temperature within the cooling phase. The energy consumption can be expressed with a linear relationship using the heater preset temperature.

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