Genetic engineering of bacteriophage and its applications for biomimetic materials

Abstract

Filamentous bacteriophage (M13) are excellent biological build block due to their multiple peptide display system including type 8 (complete peptide display at pVIII) and type 83 (complete peptide display at both pVIII and pIII) display systems. Unlike the phagemid systems, the advantage of these systems is that we can get homogenous peptide display on pVIII resulting in uniform placement of selected molecules as well as defined length and width. In this thesis, type 8 and type 83 phage were constructed and used as biological scaffolds to meet the following four specific aims. First, the self-assembly of engineered M13 bacteriophage as a template for Co-Pt crystals was demonstrated. An phage library with an octapeptide library on the major coat protein (pVIII) was used for selection of binders to cobalt ions. Fibrous structures with directionally ordered phage were obtained by interaction with cobalt ions. Co-Pt alloys were synthesized on the fibrous scaffold, and their magnetic properties were characterized. The mineralization showed organized nanoparticles on fibrous bundles with superparamagnetic properties. Second, an in vitro molecular selection method in non-biological conditions for inorganic synthesis was introduced.

(cont.) A phage display peptide library which is resistant to ethanol was constructed and used for selection against titania in 90% ethanol. The selected peptide, with a conserved basic amino acid sequence, promotes nanoparticle formation (- 60 nm) during titania synthesis by the traditional sol-gel method. Third, storage of proteins in smectically aligned phage film was demonstrated. [Beta]-galactosidase and a green fluorescent protein variant were stored in the phage film with increased stability. In addition, streptavidin conjugated phycoerythrin were aligned in a SI phage film, in which streptavidin binding peptides are displayed at the end of the phage particles. The alignment showed increased fluorescent intensity of phycoerythrin molecules. Finally, the potential of type 8 and type 83 phage as a nano-structural scaffold were studied 'or device application. An Au binding peptide was selected using type 8 phage library. Self-assembly of gold particles on phage was observed. In addition, type 83 phage which display both streptavidin binding peptide at pIII and Au binding peptide at pVIII were constructed for complex assembly of both Au nanoparticles and streptavidin conjugated nanoparticles. Genetically engineered bacteriophage show promise for application including biologically compatible materials and functional bio-inorganic hybrid materials.