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Doctoral Dissertation Announcement
Candidate: Thilak Kumara Mudalige
Doctor of Philosophy
Title: Bioengineering of Protein Nanotubes and Protein- Nanomaterial Composites
Dr. Subra Muralidharn, Chair
Dr. Brian C. Tripp
Dr. Dongil Lee
Dr. Yirong Mo
Dr. Nora Berrah
Date: Friday, May 25, 2007 3:00 p.m. – 5:00 p.m.
1260 Wood Hall
Flagella of Mesophilic bacteria such as E. coli are extremely ordered structures consisting of self-assemblies of flagellin 11-mers resulting in protein nanotubes of 2-3 nm inner diameter, 23 nm outer diameter, several microns in length and 5.4 nm separation between 11-mer segments. They are as a result attractive scaffolds and templates for the generation of uniform nanotubes and ordered array of nanoparticles. In this study fusion protein of bacterial flagellin (Fli C) and core thioredoxin (Trx), pFliTrx was used as peptide display on bacterial flagella. Various peptide loops such as cysteine, histidine, arginine-lysine, tyrosine-serine-lysine, and aspartic acid-glutamic acid were introduced in the multiple cloning site of the thioredoxin core of the FliTrx fusion protein by site directed mutagenesis and cassette mutagenesis. The flagella formed from these flagellin proteins expressed in E. coli bacteria were harvested, purified, and characterized. Various metal ions such as Cu(II), Cd(II), Ag(I), Au(I), Au(III) were bound to histidine and aspartic acid-glutamic acid peptide loops and reduced in a controlled manner to generate nanoparticle arrays and nanotubes. Slilicate and titanate ions were bound to arginine-lysine and tyrosine-serine-glycine peptide loops respectively and polymerized to obtained silica and titania nanotubes. Polyaniline nanotubes and hydroxyapatite nanoparticles were also generated on aspartic acid-glutamic acid peptide loops. The CdTe quantum dots of (3±0.3) nm diameter were bound to histidine peptide loops to generate an ordered array of quantum dots on the flagella. Exciton energy transfer between small (donor) and large (acceptor) CdTe quantum dots was demonstrated from a 5 nm (18 meV) red shift in the emission maximum for the flagella bound quantum dots compared to those free in solution. The flagella with cysteine loops aggregated through disulfide bond formation to form bundles which could be dissociated into single flagella nanotubes employing a reducing agent such as TCEP. These nanotube bundles exhibited an interesting behavior in optical traps generated with 1064 nm laser. They were repelled by the laser beam instead of being trapped resulting in their escape. This behavior which is opposite of spherical particles could be rationalized based on their shape and size. Significant results of these studies will be presented.