Research Article Volume 7 • Issue 4 670 Affiliation: Department of Chemistry, University of Toronto, 80 St. George Street, Toronto ON M5S 3H6 *Corresponding author: Michael Thompson, Department of Chemistry, University of Toronto, 80 St. George Street, Toronto ON M5S 3H6, Canada. Citation: Navina Lotay, Cynthia Maria Suarez, Brian De La Franier, Rebecca Ann Jockusch and Michael Thompson. Synthesis and Chemistry of Recombinant Gelsolin 1-3 as A Probe for The Ovarian Cancer Biomarker Lysophosphatidic Acid. Fortune Journal of Health Sciences. 7 (2024): 670-675. Received: October 25, 2024 Accepted: November 01, 2024 Published: November 15, 2024 Synthesis and Chemistry of Recombinant Gelsolin 1-3 as A Probe for The Ovarian Cancer Biomarker Lysophosphatidic Acid Navina Lotay, Cynthia Maria Suarez, Brian De La Franier, Rebecca Ann Jockusch and Michael Thompson * Abstract Gelsolin is an actin-binding protein that is competitively bound by lysophosphatidic acid (LPA), a possible biomarker for the early detection of ovarian cancer. Our group has previously shown the usage of gelsolin 1-3 (a fragment of gelsolin composed of the first three of its six subdomains) along with actin to detect the presence of lysophosphatidic acid. This histidine-tagged gelsolin 1-3 fragment is synthesized in our lab using bl21 Rosetta cells but required further characterization. This study aims to provide sufficient characterization of gelsolin 1-3 synthesized in our lab via both sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry (MS) using a Fourier-transform ion cyclotron resonance mass spectrometer (FTICR-MS). These techniques show the presence of protein at the expected mass of approximately 41 kDa. Additionally, the dissociation of a smaller fragment of the complex when introduced into the gas phase which was equivalent to the approximate mass of one subdomain of gelsolin 1-3 further supports this conclusion. Keywords: Gelsolin, SDS-PAGE, FTICR-MS, Mass spectrometry, Protein characterization Introduction Gelsolin is an 82 kDa calcium-dependent actin-binding protein that regulates cell motility and morphology through actin filament assembly and disassembly. Actin is a protein that forms microfilaments in the cell cytoskeleton and is therefore the most abundant protein in most eukaryotic cells [1]. Gelsolin binds actin monomers, severs actin filaments, and nucleates actin polymerization to form new filaments. For this to occur, actin can bind to gelsolin at three different sites [2]. The affinity for this binding is heavily dependent on the particular binding site and the presence and type of salts, with K d measured to be as low as 4.5 pM and as high as 400 μM [3–5]. Gelsolin consists of six homologous domains, 1-6, each containing a five- stranded β-sheet flanked by two α-helices: one parallel to the strands and one perpendicular [6]. The subdomains form two identical complexes (domains 1-3 and 4-6) [7–9] (Figure 1). Lysophosphatidic acid, a promising ovarian cancer biomarker, is a regulator of actin-gelsolin binding. It binds to the PIP 2 -binding domain of gelsolin with a K d of 6 nM, causing the release of actin [7,11]. Interestingly, each half of gelsolin can individually bind to LPA [7,11]. LPA has been found to have a sensitivity of 98% and specificity of 90% in the detection of ovarian cancer, with levels shown to increase with disease progression