School of Science Division of Life Science 15 Cryo-EM Study of Membrane Proteins Supervisor: DANG Shangyu / LIFS Student: LAM Yuk Ki / BCB Course: UROP 1100, Summer Human transmembrane protein 168 (TMEM168) is a protein compromise of 697 amino acids, localized on the nuclear membrane. It has been reported to be involved in various cellular regulation and pathological progress. However, the molecular mechanism of TMEM168 is elusive. Previously, we have determined the cryo-EM structure of TMEM168, which shows an extra transmembrane helix compared to the predicted structure by AlphaFold. Due to the local resolution limitation, it remains elusive the belonging of this helix. In this study, we aim to verify if this extra transmembrane helix belongs to the C-terminal of TMEM168 by protein engineering. Using the molecular cloning method, we generated a TMEM168 construct with Cterminus deletion, which will be further used to generate the virus for test expression of target protein based on the bac-to-bac system. We have produced sufficient recombinant bacmid in the past several weeks for further research. Cryo-EM Study of Membrane Proteins Supervisor: DANG Shangyu / LIFS Student: TEMIRBEKOV Yerassyl / BCB-IRE Course: UROP 2100, Fall UROP 3100, Spring TMEM168 is an evolutionary conserved eukaryotic membrane protein with an unknown structure and unclear function. It appears in a diverse set of research contexts and has been shown to have many loosely related interactions. This protein has no apparent homologs outside the TMEM168 family. Sequence analysis shows no defined known domains, suggesting that TMEM168 may have novel functional mechanisms. Our objective is to purify the target protein and resolve the structure using single-particle cryo-EM. We aim to characterize TMEM168 using its structure and additional biochemical assays. Understanding the structure and function of TMEM168 can lead to the discovery of new signal pathways, regulatory mechanisms, and other previously unknown molecular interactions. Cryo-EM Study of Membrane Proteins Supervisor: DANG Shangyu / LIFS Student: ZHEGLOV David / BIBU Course: UROP 1100, Spring UROP 2100, Summer There are two main types of sodium-dependent phosphate transporters in the human body - SLC20 and SLC34 families. In this work, we employed cryo-electron microscopy (cryo-EM) technology to elucidate the structural characteristics of proteins from the SLC34 family. Various strategies were explored to optimize protein stability in the detergent buffer and enhance soluble region features for alignment. The SLC34A1's oligomerization, caused by the cellular stress, and its connection to Pi transport efficiency were also investigated. Understanding the structure of SLC34 will enhance our knowledge of regulatory processes, uncover new molecular interactions, and offer promise for creating innovative treatments for kidney-related diseases and associated disorders.
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