Injectable, photoresponsive hydrogels for delivering neuroprotective proteins enabled by metal-directed protein assembly

Abstract

Axon regeneration constitutes a fundamental challenge for regenerative neurobiology, which necessitates the use of tailor-made biomaterials for controllable delivery of cells and biomolecules. An increasingly popular approach for creating these materials is to directly assemble engineered proteins into high-order structures, a process that often relies on sophisticated protein chemistry. Here, we present a simple approach for creating injectable, photoresponsive hydrogels via metal-directed assembly of His6-tagged proteins. The B12-dependent photoreceptor protein CarHC can complex with transition metal ions through an amino-terminal His6-tag, which can further undergo a sol-gel transition upon addition of AdoB12, leading to the formation of hydrogels with marked injectability and photodegradability. The inducible phase transitions further enabled facile encapsulation and release of cells and proteins. Injecting the Zn2+-coordinated gels decorated with leukemia inhibitory factor into injured mouse optic nerves led to prolonged cellular signaling and enhanced axon regeneration. This study illustrates a powerful strategy for designing injectable biomaterials.

Light-Triggered Carotenogenesis in Myxococcus xanthus: New Paradigms in Photosensory Signaling, Transduction and Gene Regulation

“…Satisfyingly, CarH has now been exploited as one of the few green-light responsive optogenetic tools for light-controlled: (a) gene expression in M. x anthus and transgene expression in mammalian and plant cells; (b) receptor interactions and signaling in human cells and zebra fish embryos; (c) generation of protein hydrogels that enable facile encapsulation and release of cells and proteins, and cell adhesions [ 123 , 124 , 125 , 126 , 127 , 128 ]. Notably, this last application was very recently adapted to address challenges in regenerative neurobiology to engineer metal-coordinated protein hydrogels for sustained delivery of neuroprotective cytokines aimed at neuronal survival and axon regeneration in vivo [ 126 ].…”

Extracellular Optogenetics at the Interface of Synthetic Biology and Materials Science

“…Spatial ( Wu et al (2018) ; Hörner et al (2019) ; Liu L. et al (2018) ) and temporal ( Hörner et al (2019) ; Wu et al (2018) ; Liu L. et al (2018) ; Narayan et al (2021) ; Jiang et al (2020) ; Xiang et al (2020) ; Wang et al (2017) ; Hopkins et al (2021) ; Duan et al (2021) ; Yang et al (2022) ; Emig et al (2022) ) control of OptoGel stiffness has been demonstrated using atomic force microscopy (AFM) and/or rheology ( Figure 3A,B ). Two commonly reported rheological quantities are storage modulus ( G ′) and loss modulus ( G ″ ) describing elastic and viscous responses of a material, respectively.…”

“…Advancing property and structure changes a step further, a material can undergo phase transition and enter states with new useful functionalities. Many opto-enabled materials have demonstrated light-induced gel-sol phase transitions ( Xiang et al (2020) ; Narayan et al (2021) ; Wang et al (2017) ; Lyu et al (2017) ; Jiang et al (2020) ; Zhang et al (2015) ) with rheology ( Figure 3G ), some even reversible ( Zhang et al (2015) ; Lyu et al (2017) ). The transition from gel (solid phase) to sol (liquid phase) is defined when the loss modulus G ″ exceeds the storage modulus G ′, and occurred after 1.5 h (UV, around 300 nm) for UVR8-based OptoGels ( Zhang et al (2015) ), 17 min (violet light, 405 nm, 468.6 mW ⋅ cm −2 ) for PhoCl-based OptoGels ( Xiang et al (2020) ), 2 h for Dronpa-based OptoGels (cyan light, 500 nm, nine 3 W LED beads) ( Lyu et al (2017) ).…”

“…The transition from gel (solid phase) to sol (liquid phase) is defined when the loss modulus G ″ exceeds the storage modulus G ′, and occurred after 1.5 h (UV, around 300 nm) for UVR8-based OptoGels ( Zhang et al (2015) ), 17 min (violet light, 405 nm, 468.6 mW ⋅ cm −2 ) for PhoCl-based OptoGels ( Xiang et al (2020) ), 2 h for Dronpa-based OptoGels (cyan light, 500 nm, nine 3 W LED beads) ( Lyu et al (2017) ). Various compositions of OptoGels with cobalamin-binding domains from myxococcus xanthus (MxCBD) and thermus thermophilus (TtCBD) undergo gel-sol transitions following white light exposures of 5 min (90 klx) ( Wang et al (2017) ), 8 min (35 klx) ( Jiang et al (2020) ), 20 min (30 klx) ( Wang et al (2017) ), and 30 min (30 klx) ( Narayan et al (2021) ). Light-induced phase transitions in OptoGels further extends the utility of these materials to applications such as cell elution, further discussed in Section 4.3 .…”

Interplay between chromophore binding and domain assembly by the B12-dependent photoreceptor protein, CarH

“…Mechanistic details about its function are now beginning to emerge 4,[6][7][8][9][10][11] and it is already showing great promise and versatility as the basis of photoactivated, biomolecular tools. [12][13][14][15][16] The size and structural complexity of AdoCbl reect a long and expensive biosynthetic pathway 17 and mean that its uptake 18 and subsequent binding to its dependent enzymes 19 are oen tightly regulated. The importance of these pathways is highlighted by the genetic disorders that are caused by mutations to the regulatory proteins.…”