Interplay between Disulfide Bonding and N-Glycosylation Defines SLC4 Na+-coupled Transporter Extracellular Topography
Year: 2015
Type: article
Abstract: The extracellular loop 3 (EL-3) of SLC4 Na + -coupled transporters contains 4 highly conserved cysteines and multiple N -glycosylation consensus sites. In the electrogenic Na + -HCO 3 − cotransporter NBCe1-A, EL-3 is the largest extracellular loop and is predicted to consist of 82 amino acids. To determine the structural-functional importance of the conserved cysteines and the N -glycosylation sites in NBCe1-A EL-3, we analyzed the potential interplay between EL-3 disulfide bonding and N -glycosylation and their roles in EL-3 topological folding. Our results demonstrate that the 4 highly conserved cysteines form two intramolecular disulfide bonds, Cys 583 -Cys 585 and Cys 617 -Cys 642 , respectively, that constrain EL-3 in a folded conformation. The formation of the second disulfide bond is spontaneous and unaffected by the N -glycosylation state of EL-3 or the first disulfide bond, whereas formation of the first disulfide bond relies on the presence of the second disulfide bond and is affected by N -glycosylation. Importantly, EL-3 from each monomer is adjacently located at the NBCe1-A dimeric interface. When the two disulfide bonds are missing, EL-3 adopts an extended conformation highly accessible to protease digestion. This unique adjacent parallel location of two symmetrically folded EL-3 loops from each monomer resembles a domain-like structure that is potentially important for NBCe1-A function in vivo . Moreover, the formation of this unique structure is critically dependent on the finely tuned interplay between disulfide bonding and N -glycosylation in the membrane processed NBCe1-A dimer. Background Cysteines in extracellular loop 3 (EL-3) are highly conserved in SLC4 Na + -coupled transporters. Results Disulfide bonding and N -glycosylation uniquely determine the topography of NBCe1-A EL-3. Conclusion A domain-like structure formed by EL-3 is present at the NBCe1-A dimeric interface. Significance The interplay between disulfide bonding and N -glycosylation may play an important role in transmembrane protein maturation.
Source: Journal of Biological Chemistry
Authors: Quansheng Zhu, Liyo Kao, Rustam Azimov, Natalia Abuladze, Debra Newman, Ira Kurtz
Institution: University of California, Los Angeles
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: hybrid
Estimated APC paid: $3,430
OpenAlex ID: W2005507704