TMP-HTag

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Chemical structure of the TMP-HTag linker Trimethoprim-Halotag.svg
Chemical structure of the TMP-HTag linker

Trimethoprim-Halotag (TMP-HTag) is a small molecule chemical linker developed for the rapid and reversible control of protein localization in living cells (Ballister). TMP is an dihydrofolate reductase (DHFR) inhibitor chosen for its specificity in binding to the bacterial form of DHFR. [1] [2] The other half of the linker is a Halotag, a self labelling bacterial globular protein ligand that can bind covalently and irreversibly to the chloroalkane group of a Haloenzyme. [2] [3] Positioned between the TMP group and HaloTag is a flexible linker that can be modified to optimize protein linking efficiency. [2] [3] The modular structure of TMP-HaloTag makes it an ideal heterobifunctional tool for use in chemically induced dimerization (CID). [1] [2] Additionally, TMP- HTag can be modified to include photo-cleavable groups that allow for the control of CID using light. [1] [2]

Contents

TMP-HTag Examples

AbbrFull name
THTrimethoprim-Halotag
CTH7-(diethylamino)-coumarin-4-yl] methyl (DEACM)-TMP-Halo
TNHTMP-6- nitroveratryl carbamate (NVOC-HTag
TBHTMP- Benzamide-HTag
TFHTMP-Fluorobenzamide-HTag
AbbrStructural FunctionalityRef
THLinker length optimizable for efficient dimerization [1]
CTHDEACAM group on TMP end for photo cleavability that prevents off-target dimerization [3]
TNHNVOC group between TMP and HTag for photo cleavability. Requires light activation and use may result in photo toxicity. Light sensitive so must be stored and applied in controlled light. Slightly faster synthesis process than other dimerizer syntheses [3]
TBHAdditional aromatic rings for rigidity that improves permeability and stability of heterodimer protein complexes that form with dimerization. Not light sensitive but slower. Lengthier synthesis but simpler  process than TNH [2]
TFHAromatic fluorines lend lipophilicity and possibly “stickiness” that facilitates faster dimerization than with TBH. Minimizes photo-toxicity since light is not required. [2]

TMP-HTag Applications

Example ApplicationsReferences
Mitotic studies [4]
Alternative lengthening of telomeres (ALT) cancer  study [5] [6]
CRISPR based tool development [7]
Protein localization manipulation [2]

References

  1. 1 2 3 4 5 Ballister, Edward R.; Aonbangkhen, Chanat; Mayo, Alyssa M.; Lampson, Michael A.; Chenoweth, David M. (2014-11-17). "Localized light-induced protein dimerization in living cells using a photocaged dimerizer". Nature Communications. 5 (1): 5475. doi:10.1038/ncomms6475. ISSN   2041-1723. PMC   4308733 . PMID   25400104.
  2. 1 2 3 4 5 6 7 8 Lackner, Rachel M.; O'Connell, Will; Zhang, Huaiying; Chenoweth, David M. (2022-08-17). "A General Strategy for the Design and Evaluation of Heterobifunctional Tools: Applications to Protein Localization and Phase Separation" . ChemBioChem. 23 (16): e202200209. doi:10.1002/cbic.202200209. ISSN   1439-4227. PMID   35599237. S2CID   248987486.
  3. 1 2 3 4 Zhang, Huaiying; Chenoweth, David M.; Lampson, Michael A. (2018-01-01), Maiato, Helder; Schuh, Melina (eds.), "Chapter 7 - Optogenetic control of mitosis with photocaged chemical dimerizers" , Methods in Cell Biology, Mitosis and Meiosis Part A, 144, Academic Press: 157–164, doi:10.1016/bs.mcb.2018.03.006, ISBN   9780128141441, PMID   29804668 , retrieved 2023-04-03
  4. ↑ Zhang, Huaiying; Aonbangkhen, Chanat; Tarasovetc, Ekaterina V.; Ballister, Edward R.; Chenoweth, David M.; Lampson, Michael A. (October 2017). "Optogenetic control of kinetochore function". Nature Chemical Biology. 13 (10): 1096–1101. doi:10.1038/nchembio.2456. ISSN   1552-4469. PMC   5605432 . PMID   28805800.
  5. ↑ Zhao, Rongwei; Chenoweth, David M.; Zhang, Huaiying (2021-04-12). "Chemical Dimerization-Induced Protein Condensates on Telomeres". JoVE (Journal of Visualized Experiments) (170): e62173. doi:10.3791/62173. ISSN   1940-087X. PMC   8118565 . PMID   33900288.
  6. ↑ Zhang, Huaiying; Zhao, Rongwei; Tones, Jason; Liu, Michel; Dilley, Robert L.; Chenoweth, David M.; Greenberg, Roger A.; Lampson, Michael A. (2020-08-15). Misteli, Tom (ed.). "Nuclear body phase separation drives telomere clustering in ALT cancer cells". Molecular Biology of the Cell. 31 (18): 2048–2056. doi:10.1091/mbc.E19-10-0589. ISSN   1059-1524. PMC   7543070 . PMID   32579423.
  7. ↑ Li, Qingyang; Gao, Yanmin; Wang, Haifeng (December 2022). "CRISPR-Based Tools for Fighting Rare Diseases". Life. 12 (12): 1968. doi: 10.3390/life12121968 . ISSN   2075-1729. PMC   9787644 . PMID   36556333.
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