THESIS
2021
1 online resource (xv, 92 pages) : illustrations (some color)
Abstract
Biomolecules such as fluorescent proteins (e.g. GFP and its homologs) have long
served as versatile tools in a broad range of research areas and, in couple with proper
protein engineering, have also generated numerous genetically encoded devices and
materials for biomedical applications. However, many of these resulting tools are
constrained in their abilities to sense and respond to long-wavelength (red or near-infrared;
600 nm) light, which is important for biomedical applications as far as deep-tissue
penetration is concerned. It is therefore necessary to seek for alternative protein
molecules or mechanisms to overcome this limitation. The water soluble chlorophyll
binding protein (WSCP) derived from plants, which is characterized by its marked
stability and responsiveness to 650 nm...[
Read more ]
Biomolecules such as fluorescent proteins (e.g. GFP and its homologs) have long
served as versatile tools in a broad range of research areas and, in couple with proper
protein engineering, have also generated numerous genetically encoded devices and
materials for biomedical applications. However, many of these resulting tools are
constrained in their abilities to sense and respond to long-wavelength (red or near-infrared;
> 600 nm) light, which is important for biomedical applications as far as deep-tissue
penetration is concerned. It is therefore necessary to seek for alternative protein
molecules or mechanisms to overcome this limitation. The water soluble chlorophyll
binding protein (WSCP) derived from plants, which is characterized by its marked
stability and responsiveness to >650 nm light , is deemed with such potential but has yet
to receive any attention. This thesis aims to develop WSCP into a versatile tool for
designing photoresponsive materials, as well as for bioimaging.
Using the WSCP from Lepidium virginicum, genetically encoded click chemistry,
and mussel foot proteins (MFP), an entirely protein-based photoresponsive hydrogel was
developed. In this material, the WSCP acted as a singlet oxygen generator on exposure
to red light, leading to oxidative crosslinking of tyrosine in MFP and strengthening the
gel mechanics. Thanks to the photothermal effect of the WSCP, the resulting hydrogel
showed the potential for photoacoustic imaging.
The WSCP was also converted to a new class of near infrared fluorescent proteins
through the combined use of protein engineering and ligand replacement with
bacteriochlorophylls. A key residue that controlled the spectral properties of WSCP was
identified and mutated, red-shifting the emission of the protein by >100 nm. These
proteins were expressed in mammalian cells and reconstituted with bacteriochlorophylls
in vitro.
Together, these studies demonstrated the WSCP as a versatile platform for creating
new materials and molecular tools, pointing to great potential in biomedical research.
Post a Comment