Biology subjects

Nam, K. H.

Publications and source records attributed to Nam, K. H..

6 recordsLinked to original sources

bioRxiv2024

Evaluation of AlphaFold3 for the fatty acids docking to human fatty acid-binding proteins

Human fatty acid-binding proteins (FABPs) are involved in many aspects of lipid metabolism, such as the uptake, transport, and storage of lipophilic molecules, as well as cellular functions. Understanding how FABPs recognize fatty acids (FAs) is crucial for identifying FABP function and applications, such as in inhibitor design or biomarker development. The recently developed AlphaFold3 (AF3) demonstrates significantly higher accuracy than other prediction tools, particularly in predicting protein-ligand interactions with state-of-the-art docking tools. Studies on whether AF3 can be used to identify the FAs of FABP are lacking. To assess the accuracy of FA docking to FABPs using AF3, models of FA docked into FABP generated using AF3 were compared with experimentally determined FA-bound FABP structures. FA ligands in AF3 structures docked reliably into the FA-binding pocket of FABPs; however, the detailed binding configuration of most FA ligands docked into FABPs and the interaction between FA and FABP determined using AF3 and experimentally differed. These results will aid in understanding FA docking to FABPs and other FA-binding proteins using AF3.

bioinformatics↗

bioRxiv2022

Disruption of lysosomal proteolysis in astrocytes facilitates midbrain proteostasis failure in an early-onset PD model.

Accumulation of advanced glycation end products (AGEs) on biopolymers accompany cellular aging and drives poorly understood disease processes. Here, we studied how AGEs contribute to development of early on-set Parkinsons Disease (PD) caused by loss-of-function of DJ1, a protein deglycase. In induced pluripotent stem cell (iPSC)-derived midbrain organoid models deficient for DJ1 activity, we find that lysosomal proteolysis is impaired, causing AGEs to accumulate, -synuclein (-syn) phosphorylation to increase, and proteins to aggregate. These processes are at least partly driven by astrocytes, as DJ1 loss reduces their capacity to provide metabolic support and triggers acquisition of a pro-inflammatory phenotype. Consistently, in co-cultures, we find that DJ1-expressing astrocytes are able to reverse the proteolysis deficits of DJ1 knockout midbrain neurons. In conclusion, astrocytes capacity to clear toxic damaged proteins is critical to preserve neuronal function and their dysfunction contributes to the neurodegeneration observed in PD.

neuroscience↗

bioRxiv2022

Combination of an inject-and-transfer system for serial femtosecond crystallography

Serial femtosecond crystallography (SFX) enables the determination of the room-temperature crystal structure of macromolecules without causing radiation damage and provides time-resolved molecular dynamics by pump-probe experiments. In the SFX experiment, the injector-based sample delivery method continuously provides fresh crystals to X-rays, and the fixed-target scanning method can be programmed to move the crystals to the desired location. This study introduces a combination of the inject-and-transfer system (BITS) method for sample delivery for SFX experiments, a hybrid injection, and a fixed-target scanning method. BITS allows solution samples to be reliably deposited on an ultraviolet ozone (UVO)-treaed polyimide films at flow rates as low as 1 nl/min. In application of BITS in SFX experiment, the lysozyme crystal samples were embedded in a viscous lard medium and injected at a 50-100 nl/min flow rate through a syringe needle onto an UVO-treated polyimide film mounted on a fixed-target scan stage. The deposited crystal sample on film were raster scanned to XFEL by motion stage in the horizontal and vertical directions. Using this method, we successfully determined the room-temperature structure of lysozyme at 2.1 [A] resolution. This method can be applied to the SFX experiments.

biophysics↗

bioRxiv2020

Viscous medium-based crystal support in sample holder for fixed-target serial femtosecond crystallography

Serial femtosecond crystallography (SFX) enables the determination of a room-temperature crystal structure of macromolecules without causing radiation damage, as well as provides time-resolved molecular dynamics data in pump-probe experiments. Fixed-target SFX (FT-SFX) can minimize sample consumption and physical effects to crystals during sample delivery. Various types of sample holders have been developed and applied in FT-SFX; however, no sample holder has been developed that can universally mount crystals of various sizes and shapes. Here, we introduce a viscous media-based crystal support in a sample holder for FT-SFX. Crystal samples were embedded in viscous media such as gelatin and agarose, which were enclosed in a polyimide film. In the vertically placed sample holder, the viscous medium stably supported crystals between the two polyimide films without crystal sinking due to gravity. Using this method, we performed FT-SFX experiments with glucose isomerase and lysozyme embedded in gelatin and agarose, respectively. The room-temperature crystal structures of glucose isomerase and lysozyme were successfully determined at 1.75 and 1.80 [A] resolutions, respectively. Viscous media used in this experiment showed negligible background scattering in data processing. This method is useful for delivering crystal samples of various sizes and shapes in FT-SFX experiments.

biophysics↗

bioRxiv2019

Sample delivery using capillary and delivery medium for serial crystallography

Serial crystallography (SX) is an innovative technology in structural biology that enables the visualization of molecular dynamics of macromolecules at room temperature. SX experiments always require a considerable amount of effort to deliver a crystal sample to the X-ray interaction point continuously and reliably. Here, a sample delivery method using a capillary and a delivery medium is introduced. The crystals embedded in the delivery medium can pass through the capillary tube, which is aligned with the X-ray beam, at very low flow rates without requiring elaborate delivery techniques and drastically reducing sample consumption. This simple but highly efficient sample delivery method can allow researchers to deliver crystals precisely to X-rays in SX experiments.

biophysics↗

bioRxiv2019

Shortening injection matrix for serial crystallography

Serial crystallography (SX) allows crystal structures to be observed at room temperature through the steady delivery of crystals to the X-ray interaction point. Viscous delivery media are advantageous because they afford efficient sample delivery from an injector or syringe at a low flow rate. Hydrophobic delivery media, such as lipidic cubic phase (LCP) or grease, provide a very stable injection stream and are widely used. The development of new hydrophobic delivery materials can expand opportunities for future SX studies with various samples. Here, I introduce fat-based shortening as a delivery medium for SX experiments. This material is commercially available at low cost and is straightforward to handle because its phase (i.e., solid or liquid) can be controlled by temperature. Shortening was extruded from a syringe needle in a very stable injection stream even below 200 nl/min. X-ray exposed shortening produced several background scattering rings, which have similar or lower intensities than those of LCP and contribute negligibly to data processing. Serial millisecond crystallography was performed using two shortening delivery media, and the room temperature crystal structures of lysozyme and glucose isomerase were successfully determined at resolutions of 1.5-2.0 [A]. Therefore, shortening can be used as a sample delivery medium in SX experiments.

biophysics↗