The Photosensitive MNI Protecting Group

Published on 29/01/2025

Shining light on 4-methoxy-7-nitroindoline (MNI) photocaged aspartate and glutamate building blocks suitable for Fmoc-SPPS. Liberate the sidechain carboxylic acid by irradiation with UV-light of 365 nm.

The Photosensitive MNI Protecting Group

Photocaging is a technique where a light-sensitive protection group is attached to a functional group of a molecule. It may be removed by the user by controlled light irradiation. Thus, photocaging is a very useful method to control the spatiotemporal activity of molecules, esp. in conjunction with microscopes and lasers. It is frequently applied, e.g., in cellular signaling and protease research, in the controlled activation and liberation of drugs in photodynamic therapy, or for controlling the charge of polymer side chains. But photochemical reactions also may be carried out in dedicated batch or flow reactors. 

In this blog, we’ll introduce 4-methoxy-7-nitroindoline (MNI) derivatives of aspartate (Fmoc-Asp(MNI)-OH; FAA9555) and glutamate (Fmoc-Glu(MNI)-OH; FAA9560) suitable for Fmoc-SPPS. The MNI group acts as photosensitive protecting group and can be removed by irradiation with UV-light of 365 nm and 2-photon absorption in the double-wavelength near-infrared (NIR) range, to reveal the carboxy side chain. 

Applications of MNI-protected amino acids are, e.g., in UV-mediated gel-sol transition in peptide hydrogels (used for the stabilization of blood vessel anastomosis in microsurgical procedures) or for receptor studies. MNI has also been used as one of multiple dyes in wavelength-multiplexed photo-uncaging and in photoactivable caspase inhibitors.

 


Illustration of the MNI protection in SPPS: shown is the excerpt of a peptide with the sequence -Ala-Asp/Glu(MNI)-Gly-, (for Asp: n= 1, for Glu: n= 2). The MNI-protected building blocks are stable in standard Fmoc-SPPS. Upon irradiation with UV-light of 365 nm, the carboxy function is deprotected and 4-methoxy-7-nitroindoline is released.

 

The MNI photocage has excellent photochemical properties: The quantum yield for MNI-protected Glu is 0.065; the two-photon cross-section is 0.06 GM (Goeppert-Mayer units). MNI is rapidly removed, the photo-release half-time is less than 0.26 ms, allowing for complete deprotection of batches within minutes. The nitroindolinyl moiety is stable under aqueous conditions and in buffers which are commonly used for biochemical and physiological experiments (PBS, HEPES), while ester-based protection groups (e.g., nitrophenyl moieties) are prone to hydrolysis by esterases which are present in biological samples. 

In addition, the MNI group prevents the formation of aspartimide (aminosuccinyl) side products and pyrrolidones during Fmoc-SPPS! 

 

→ MNI may also serve as protection group for alcohols and amines. For more details, please get in contact! 

→ Struggling with aspartimide formation? See our flyer and find ways to prevent/reduce this undesired side reaction! 

 

References:

An Fmoc-compatible method for synthesis of peptides containing photocaged aspartic acid or glutamic acid; S. Tang, J.-Y. Cheng, J.-S. Zheng; Tetrahedron Lett. 2015; 56(31): 4582-4585. https://doi.org/10.1016/j.tetlet.2015.06.016

Controlled Inhibition of Apoptosis by Photoactivatable Caspase Inhibitors; S. Chakrabarty, S. H. L. Verhelst; Cell Chem. Biol. 2020; 27: 1434-1440. https://doi.org/10.1016/j.chembiol.2020.08.001 

A multiphase transitioning peptide hydrogel for suturing ultrasmall vessels; D. J. Smith, G. A. Brat, S. H. Medina, D. Tong, Y. Huang, J. Grahammer, G. J. Furtmüller, B. C. Oh, K. J. Nagy-Smith, P. Walczal, G. Brandacher, J. P. Schneider; Nat. Nanotech. 2016; 11: 95-102. https://doi.org/10.1038/nnano.2015.238

Biomimetic Heterodimerization of Tetrapeptides to Generate Liquid Crystalline Hydrogel in A Two-Component System; B. Wu, S. Zhao, X. Yang, L. Zhou, Y. Ma, H. Zhang, W. Li, H. Wang; ACS Nano 2022; 16(3): 4126-4138. https://doi.org/10.1021/acsnano.1c09860

Wavelength-selective caged surfaces: how many functional levels are possible? V. SanMiguel, C. G. Bochet, A. del Campo. J. Am. Chem. Soc. 2011; 133(14): 5380-5388. https://doi.org/10.1021/ja110572j 

Cooperative Veratryle and Nitroindoline Cages for Two-Photon Uncaging in the NIR; Chem. Eur. J. 2016; 22(31): 10848-10859. https://doi.org/10.1002/chem.201601109 

Photolysis of a peptide with N-peptidyl-7-nitroindoline units using two-photon absorption, K. A Hatch, A. Ornelas, K. N Williams, T. Boland, K. Michael, C. Li; Biomed. Opt. Express 2016; 7(11): 4654-4659. https://doi.org/10.1364/boe.7.004654 

Comparative one- and two-photon uncaging of MNI-glutamate and MNI-kainate on hippocampal CA1 neurons; S. Passlick, G. C. R. Ellis-Davies; J. Neurosci. Meth. 2018; 293: 321-328. https://doi.org/10.1016/j.jneumeth.2017.10.013 

Synthesis and characterization of a photocleavable collagen-like peptide; A. Ornelas, K. N. Williams, K. A. Hatch, A. Paez, A. C. Aguilar, C. C. Ellis, N. Tasnim, S. Ray, C. W. Dirk, T. Boland, B. Joddar, C. Li, K. Michael; Org. Biomol. Chem. 2018; 16(6): 1000-1013. https://doi.org/10.1039/c7ob02198d 

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