Contributors: Ela Dudek, PhD; Brittany Fraser, MSc; Paige Grant, MSc; Iryna Ilienko, PhD; Diane Jeon, BCom; Laine Lysyk, MSc; Thuy Nguyen, PhD; Anyeld Ubeda, MSc; Simon Wu, MSc
In stem cell and therapeutic research, maintaining similarity to the physiological state is essential. Most proteins in the human body are naturally glycosylated, which impacts their function and interaction within cellular environments (Berger et al., Nadeem et al.)
Using glycosylated recombinant proteins for stem cell research more closely mimics this physiological environment, promoting more natural cellular responses, enhancing stem cell maintenance, and exerting greater control over growth and differentiation pathways (Berger et al).
Glycosylation plays a role in protein folding and quality control in the endoplasmic reticulum, ensuring proper protein structure. This leads to biologically relevant outcomes, as glycosylated proteins typically display increased stability, solubility, and activity over non-glycosylated versions. Moreover, glycosylation enables the study of complex cellular processes—such as cell adhesion, immune responses, and signaling pathways—that are mediated by glycan-protein interactions. Overall, the use of glycosylated proteins in research provides a more accurate representation of protein function in living systems, contributing to findings that are more likely to be translatable and clinically relevant.
Future Fields produces the first reported functional, glycosylated recombinant human prolactin (hPRL) in Drosophila melanogaster. Compared to unicellular hosts, the Drosophila melanogaster (D. mel) expression system can produce more complicated proteins and perform complex post-translational modifications, including glycosylation, similar to mammalian systems such as CHO and HEK293 cells. In our latest tech note, we dive deeper into the physico-chemical and biological characterization of D. mel-derived hPRL in comparison with hPRL recombinantly expressed in E. coli and HEK293 systems.
We characterized recombinant human prolactin (hPRL) produced in D. mel by assessing its purity through gel electrophoresis, verifying its identity with Western blotting, analyzing post-translational modifications using glycosidase digestion, and evaluating its activity via Surface Plasmon Resonance and Nb2-11 cell proliferation assays. Future Fields’ recombinant hPRL was compared to standard hPRL preparations derived from E. coli and HEK293 expression systems.