https://scholar.dgist.ac.kr/handle/20.500.11750/58112 2026-04-09T22:12:26Z https://scholar.dgist.ac.kr/handle/20.500.11750/59165 Title: Intracristal space proteome mapping using super-resolution proximity labeling with isotope-coded probes Author(s): Kang, Myeong-Gyun; Shin, Sanghee; Jang, Dong-Gi; Kwon, Ohyeon; Lee, Song-Yi; Mishra, Pratyush Kumar; Jung, Minkyo; Mun, Ji Young; Kee, Jung-Min; Kim, Jong-Seo; Rhee, Hyun-Woo Abstract: Proximity labeling with engineered ascorbate peroxidase (APEX) has been widely used to identify proteomes within various membrane-enclosed subcellular organelles. However, constructing protein distribution maps between two non-partitioned proximal spaces remains challenging with the current proximity labeling tools. Here, we introduce a proximity labeling approach using isotope-coded phenol probes for APEX labeling (ICAX) that enables the quantitative analysis of the spatial proteome at nanometer resolution between two distinctly localized APEX enzymes. Using this technique, we identify the spatial proteomic architecture of the mitochondrial intracristal space (ICS), which is not physically separated from the peripheral space. ICAX analysis further reveals unexpected dynamics of the mitochondrial spatiome under mitochondrial contact site and cristae organizing system (MICOS) complex inhibition and mitochondrial uncoupling, respectively. Overall, these findings highlight the importance of ICS for mitochondrial quality control under dynamic stress conditions. 2025-07-31T15:00:00Z https://scholar.dgist.ac.kr/handle/20.500.11750/58971 Title: Directed evolution of LaccID for cell surface proximity labeling and electron microscopy Author(s): Lee, Song-Yi; Roh, Heegwang; Gonzalez-Perez, David; Mackey, Mason R.; Hoces, Daniel; Mclaughlin, Colleen N.; Lin, Chang; Adams, Stephen R.; Nguyen, Khanh; Kim, Keun-Young; Luginbuhl, David J.; Luo, Liqun; Udeshi, Namrata D.; Carr, Steven A.; Hernandez-Lopez, Rogelio A.; Ellisman, Mark H.; Alcalde, Miguel; Ting, Alice Y. Abstract: Enzymes that oxidize aromatic substrates have been harnessed for cell-based technologies including proximity labeling and electron microscopy; however, they are associated with drawbacks such as the need for toxic H2O2. Here, we explore multicopper oxidases (laccases) as a new enzyme class for proximity labeling and electron microscopy in mammalian cells. LaccID was generated through 11 rounds of directed evolution from an ancestral fungal laccase and catalyzes one-electron oxidation of diverse aromatic substrates using O2 instead of toxic H2O2. Surprisingly, we found that LaccID is selectively active at the surface plasma membrane of both living and fixed cells. We use LaccID proximity labeling and mass spectrometry to map the changing surface proteome of T cells that engage with tumor cells through antigen-specific T cell receptors. In addition, we use LaccID as a genetically encodable tag for EM visualization of cell surface features in mammalian cell culture and in the fly brain. Our study paves the way for future cell-based applications of LaccID. 2025-11-30T15:00:00Z https://scholar.dgist.ac.kr/handle/20.500.11750/11518 Title: Supra-blot: an accurate and reliable assay for detecting target proteins with a synthetic host molecule-enzyme hybrid Author(s): Sung, Gihyun; Lee, Song-Yi; Kang, Myeong-Gyun; Kim, Kyung Lock; An, Jaeyeon; Sim, Jaehwan; Kim, Sungwan; Kim, Seung Jun; Ko, Jaewon; Rhee, Hyun-Woo; Park, Kyeng Min; Kim, Kimoon Abstract: In accordance with the rapid increase in demand for selective and spatial chemical tagging, and accurate detection of proteins of interest, we develop a sensitive protein detection method, termed "Supra-blot" capitalizing on high-affinity host-guest interaction between cucurbit[7]uril (CB[7]) and adamantylammonium (AdA). The method can directly detect chemically tagged proteins without false-positive signals caused by endogenous biomolecules. Not only a single specific protein, but also spatially localized proteins in cells were labeled with AdA, and selectively detected by a host molecule-enzyme hybrid, CB[7]-conjugated horseradish peroxidase (CB[7]-HRP) generating amplified chemiluminescence signals. This study shows the great potential of Supra-blot for accurate and reliable detection of proteins of interest in cells. © 2020 The Royal Society of Chemistry. 2020-01-31T15:00:00Z