Zinc finger domains are complexes found within a group of proteins that contain certain transcription factors allowing for the regulation of specific genes, or portions of cellular DNA. If specific genes in prostate cancer cells can be regulated – either turned on or turned off – the transcriptional function of the genome could be controlled. Potentially, this could be used for targeted apoptosis (or cell death) of cells containing those specific genes. This study involved the use of BL21 competent cells to express AreA protein, a ninety-two amino acid zinc finger domain-containing protein with a molecular weight of 10038 Da. For purification purposes, a six-histidine tag was included. Sodium Dodecyl Sulfate Polyacrylamaide Gel Electrophoresis (SDS-PAGE) verified expression of the protein, which was then purified utilizing a Nickel NTA affinity resin. Histidines bind strongly to this affinity resin whereascellular proteins, which do not typically have a histidine tag, do not bind. This allowed for easy purification of the target AreA protein. After the purification procedure was complete, SDS-PAGE was used once more to verify that the eluate had yielded pure protein. Fluorescence spectroscopy qualitatively indicated that the purified protein interacts with its target GATA sequence of DNA. Current research includes quantitative analysis of the binding of the protein to the target DNA sequence. In addition, we are currently in the process of designing a gene to code for a protein that targets a sequence of DNA found only in certain prostate cancer cells. This protein will be expressed and purified as above and analyzed for binding to its target sequence using fluorescence spectroscopy. Our overall goal is to design a protein that can target prostate cancer cells for apoptosis.

Analyzing the binding of a zinc-finger protein to its target DNA

2019 Spring National Meeting

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