Development of methodologies to identify polysialylated proteins and to visualize receptors for polysialic acid in biological samples

• Author / Creator

  Idrees, Muhammad

• Polysialic acid (polySia) is a homo polymer made of Sialic acid monomers linked to each other through ⍺-2-8 linkages in humans. Its expression on cellular surfaces is limited to nervous,
  immune, and reproductive systems in a healthy adult human. It is overexpressed in certain tumors and results in higher invasion and metastasis. It increases cancer progression and severity
  and is associated with higher mortality and decreased survival rates. Only a handful of polysialylated proteins have been discovered so far. Willis’s lab has confirmed the presence of
  more such proteins that need to be identified. Research has also shown that polySia is an immunomodulatory ligand, but additional studies are required to identify its receptor(s). To
  explore the polySia role further, the development of reliable and sensitive methods is essential as limited methodologies are available to work with polySia.
  We propose two tactics to enhance polySia research. Firstly, we have developed a biotin-modified sialic acid that can be incorporated onto the polySia chain of polysialylated protein and
  subsequently extracted using streptavidin beads. Secondly, we propose employing a polysialylated fluorescent protein as a ligand to detect the presence of polySia receptors on
  immune cells.
  The modified sialic acid with an attached biotin though disulfide linkage (CMP-Sia-S-S-Biotin) has been synthesized and successfully added to polySia chains on small molecules and
  cell surface polySia. We could also effectively extract biotin-polySia-protein with streptavidin beads and reduce the disulfide bond. To confirm the presence of polySia receptor(s), a fluorescent ligand was designed and incubated with Jurkat T cells. Later, the ligand was modified to enhance its avidity. These methods lay the groundwork for future research and development.

• Subjects / Keywords

  • Polysialic acid
  • Sialic acid
  • Overexpression
  • immune cells

• Graduation date

  Fall 2024

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/r3-mrfn-5788

• License

  This thesis is made available by the University of Alberta Library with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.