Molecularly, collagen exists as a triple helical structure that is stabilized by the presence of the amino acid hydroxyproline. These triple helices further assemble linearly and laterally to form fibers on length-scales of micro- to millimeters (Figure 1) to achieve the desired mechanical properties and biological functions of the collagen in tissues. Chemical synthesis of short collagen-mimetic peptides sequences to date have utilized hydroxyproline-based, thermally stable sequences because simple proline-based sequences do not deliver stability and higher order assemblies. While chemical techniques are severely limited to producing mainly short sequences, bio-synthetic (natural) routes can be employed to produce very long and modular collagen sequences, which would far more closely mimic the natural collagens. However, the simple biosynthetic production of such thermally stable sequences has not been possible due to the lack of a specific enzyme inside the bacterial host for producing hydroxyproline, which thus results in collagens of low thermal stability.

COLLAGEN LIKE PEPTIDES FOR BIOMIMETIC FOLDING, ASSEMBLY AND BIOLOGICAL FUNCTIONS

Kristi L. Liick, University of Delaware, DMR 0239744

Figure 1. Schematic of collagen assembly

Figure 2. Design of hydroxyproline-lacking collagen-mimetic peptide sequence, that is thermally stable and forms fibrils, like native collagen.

Collagens are one of the most abundant fibrous proteins found in body tissues and organs, endowing structural integrity, mechanical strength and multiple biological functions. Destabilization or deficiencies of collagens in the body lead to various diseases such as arthritis, corneal opacity resulting in blurry vision, arteriosclerosis, limited joint flexibility, and others. Owing to its prevalence in the body, there have been many applications developed to use collagen-based materials, such as in wound healing, as surgical glues, and in the food processing industry. However, collagens extracted from bovine or other sources suffer the risk of adverse immunological and pathological consequences. Therefore, collagen-mimetic peptides and polypeptides synthesized chemically or biosynthetically represent useful alternatives in the production of collagen-based materials.

Addressing these challenges, in this program we have designed and synthesized a collagen-mimetic peptide sequence which is thermally stable under physiological conditions even in the absence of hydroxyproline (melting temperature ~45 ˚C), shows higher-order assembly to form fibrils, and contains a biologically relevant peptide motif (Figure 2). Spectroscopic characterization of the peptides confirms that they form triple helical structures like those adopted by native collagens, and that these triple helical structures are thermally stable under physiological conditions. Additional characterization of the peptides confirms that they form higher order structures on the nanometer and micrometer length scale (Figure 2), despite the fact that they contain no assembly-directing motifs, in contrast to other collagen-mimetic systems. These results suggest the potential for their biosynthetic production in bacterial hosts, yielding collagen-like polypeptides with improved thermal stability and biological activity over those currently available. The production of modular polypeptides and block polymers capable of sequential assembly into complex and responsive structures is a current focus of our work.

Triple helix assembly

Higher-order assembly

In addition to the training and development activities of the PI, this program supports outreach efforts at both the undergraduate and secondary school levels. At the undergraduate level, the PI continues to participate in undergraduate research

discussions and poster sessions, and has served as a thesis advisor and reader for several UD undergraduates completing honors theses in the

chemical sciences. These students and their work have been included on several published manuscripts and presentations at regional and national

meetings, and the students have continued employment in industry as well as in graduate school.

An outreach program, “The Science of Art”, developed by Kiick in collaboration with an art teacher (Karen Kiick) at Haddon Township High School in Westmont,

New Jersey, has also been continued over the course of this grant program. The outreach activities have included UD graduate students in Materials

Science and Engineering and at least 30-40 students, over 90% of whom are female, in the arts and crafts curriculum at Haddon Township. Activities have been focused on the science of materials manipulation and have focused on three main areas to date: ceramic glaze formulation (Figure 1), metal surface

modification, and titanium anodization (Figure 2).

Based on the results of this program, students in this curriculum now create their own glazes and have incorporated a knowledge of metal patinas in their artwork. They have gathered an appreciation for the chemical

and materials properties of the materials with which they work, and have hopefully gained increased familiarity and confidence with the manipulation of

materials via application of chemical methods. The outreachefforts (Figure 3) are currently directed at the production of a manual

describing the glazes, patinas, and outreach activities, and have beenexpanded to include other materials science demonstrations

and experiments. We intend to integrate these materials into demonstrations regionally, as well as into curriculum materials at the high school level.

PROTEINS CONTAINING NON-NATURAL AMINO ACIDS AS BUILDING BLOCKS FOR NOVEL MATERIALS

Kristi L. Liick, University of Delaware, DMR 0239744

Figure 2. Anodized titanium pieces (left) and metal surfaces (right, Ni shown) prepared for an outreach module. The image on the right is shown as observed under the optical microscope at a magnification=200x.

Figure 1. Examples of the new glaze formulations employed by high school students and teachers at Haddon Township High School in Westmont, New Jersey.

Figure 3. Currentoutreach participantsat the University of Delaware.