Browsing by Subject "R5 peptide"

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Open Access
Genetically programmed engineered cells for biomaterials synthesis
(2021-01) Kırpat, Büşra Merve
Several organisms can process nanomaterials and producing in various sizes and morphologies in mild conditions by utilizing specific proteins. In sea sponges, silicatein proteins play a key role in synthesizing silica nanoparticles the precursor silicic acid. Silaffin proteins in diatoms can also biomineralize silica. One subunit of silaffin called R5 peptide has a key role for nucleation and initiation of the nanoparticle formation and it has been shown that bacteria synthesized R5 peptide has ability to precipitate silica structures. These silica nanostructures can be utilized in many areas. Silica-based cements take attentions to make them useful in restorative dentistry and endodontics. In this work, a synthetic cell system has reprogrammed autotransporter (Ag43) system to display R5 peptide fused with fluorescent proteins. After displaying the fused proteins on the surface of bacteria or secreting them into environment, whole cell or the proteins are used to precipitate silica in the presence of precursor such as tetramethyl orthosilicate (TMOS). These silica structures are used to evaluate their in vitro effects on the proliferation of dental pulp stem cells (DPSCs) and their osteogenesis.
Open Access
A synthetic biology approach for nanomaterial design, synthesis and functionalization
(2017-11) Ölmez, Tolga Tarkan
Biological formation of inorganic material occurs in most organisms in nature. Various biomolecules such as polypeptides, lipids and metabolites are responsible for biomineralization in cells and tissues. Biological synthesis of biohybrid materials is a recently emerged discipline that uses these biomolecules in synthetic biological systems. Synthetic biology is one of most promising approaches for the development of biohybrid systems, and stands at the intersection of computer science, engineering and molecular genetics. Synthetic biology tools allow the design of programmable genetic toolkits that can compete with natural biosynthesis systems. The present thesis elaborates on the formation of well-controlled genetic systems that can synthesize and functionalize biological materials. Artificial peptides were fused to various genes through molecular genetics techniques, allowing the production of designer proteins. One aspect concerns the fusion of the 19 amino acid-long R5 motif of silaffin protein to three distinct fluorescent proteins. The R5 peptide motif can nucleate silica precursor ions to synthesize silica nanostructures. Therefore, fusion of fluorescent proteins with the R5 motif allows the synthesis and encapsulation of fluorescent silica nanoparticles. Due to its affinity to silica, R5 tag was also shown to be a candidate tag for silica resin-based affinity chromatography purification. Using synthetic biology tools, production of autonomously formed biotemplating platforms can be achieved. A bacterial functional amyloid fiber biosystem called curli can be utilized as a biotemplating platform for nanomaterials synthesis in this context. The major curli subunit CsgA was fused to artificial peptides that can nucleate and synthesize various nanomaterials. Inducible systems were also integrated into the genetic design system to confer temporal control over curli synthesis. These designs were improved through the incorporation of material-sensitive transcription factors and their cognate promoters for ions of cadmium, gold and iron. First, these material sensitive pairs were used in the development of microbial whole cell sensors that produce a fluorescence output upon induction by material precursor ions. Later, material-sensitive pairs were integrated into a modified curli nanofiber display biosystem to produce living autonomous whole cell nanomaterial synthesizers. These systems recognize precursor ions in the environment and synthesize modified curli nanofibers that can nuclate precursor ions to form functional nanomaterials.