Document Type

Honors Capstone Project

Date of Submission

Spring 5-1-2013

Capstone Advisor

Dr. Ivan Korendovych

Honors Reader

Dr. James Hougland

Capstone Major

Biology

Capstone College

Arts and Science

Audio/Visual Component

no

Capstone Prize Winner

no

Won Capstone Funding

yes

Honors Categories

Sciences and Engineering

Subject Categories

Biochemistry | Biochemistry, Biophysics, and Structural Biology

Abstract

Abstract

Metalloenzymes constitute a large subclass of proteins with diverse functions including metal-sensing, signal transduction pathways, and metal transport. In this report, I aim to study the intricate relationship between structure and function as it relates to metalloproteins in the fields of catalysis and bioimaging. For catalysis, I used existing platforms in addition to de novo design to study small scaffolds with varying secondary structure, and their function in catalyzing the hydrolysis of a p-nitrophenol ester. For bioimaging, I used an existing enzyme known as human carbonic anhydrase I, hCA, to develop a bioimaging agent that can be fused to other biological molecules. After subjecting hCA to mutagenesis, I altered its metal binding site to accommodate a rhenium based complex; these results could eventually be applied to Tc99m due to the two metals’ similar reactivity, and used as a bioimaging technique based on technetium’s gamma radiation, which aids in visualization. Similar approaches are under current development but they are based on synthetic peptide probes. The hCA-Re complex is based on a naturally occurring enzyme, which could eliminate any immunogenic response. Finally, all of the described genes were cloned using ligation-independent cloning, LIC, which is an advantageous although uncommonly used cloning method. LIC does not usually require the purchase of a commercial kit, and it does not require DNA ligase, hence the name of the method. In addition, virtually any gene can be cloned using LIC as the gene and vector sequences do not play major roles in LIC design.

Creative Commons License

Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.

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Biochemistry Commons

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