A Characterization of the Quadruple Mutant of the FGF1-FGF2 Heterodimer

Author

Kenlee ClabornFollow

Date of Graduation

5-2023

Document Type

Thesis

Degree Name

Bachelor of Science in Chemistry

Degree Level

Undergraduate

Department

Chemistry & Biochemistry

Advisor/Mentor

Thallapuranam, Suresh

Committee Member/Reader

Kayser, Casey

Committee Member/Second Reader

Dong, Bin

Committee Member/Third Reader

Potra, Adriana

Abstract

Human Fibroblast Growth Factor (FGF) was first identified as a protein whose function was to aid in cell proliferation, but now, FGF has been determined to consist of over 23 different proteins in 7 different subfamilies with three different types of cell-signaling purposes.¹ In the first subfamily are the two most notable and studied proteins, Fibroblast Growth Factor 1 (FGF1) and Fibroblast Growth Factor 2 (FGF2). Respectively, they are known for their unique capabilities of cell proliferation/mitogenesis and angiogenesis.² To induce the cell-signaling pathway that leads to the function of these proteins, FGF1 and FGF2 binds to their cofactor, heparin. This induces a conformational change in each protein, allowing them to bind to their target receptor, leading to the actions of cell proliferation, cell differentiation, or angiogenesis.³ These properties are especially significant in terms of wound healing because they translate to the creation of new, specialized cells and the formation of new blood vessels. This has led to the hypothesis that a heterodimer of FGF1 and FGF2 (FGF1-2) will have cell proliferative and angiogenic properties, but with mutations, will be more thermally stable, resistant to cleavage, and more bioactive. This project investigated the characteristics of the quadruple mutant of FGF1-2 with the R136E, K126N, Q54P, and S61L mutations. Through protein expression in recombinant E. coli, the mutated protein was purified using an affinity column with heparin-Sepharose linked beads. After analysis with fluorescence and circular dichroism spectroscopy, the protein was determined to be folded correctly and with increased stability in its structure with the added mutations. Through analysis of thermal denaturation, the mutated protein was found to have an increased resistance to thermal denaturation of about 6°C from the heterodimer.

Keywords

Fibroblast Growth Factors, heterodimer, characterization, FGF1, FGF2

Citation

Claborn, K. (2023). A Characterization of the Quadruple Mutant of the FGF1-FGF2 Heterodimer. Chemistry & Biochemistry Undergraduate Honors Theses Retrieved from https://scholarworks.uark.edu/chbcuht/48