Abstract 3238: The transmembrane domain of prostate-specific membrane antigen: Its role in oligomerization and activation in prostate cancer and nonprostatic neovasculature

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Prostate cancer (PCa) is the second leading cause of cancer-related deaths among American men. The high expression of the membrane protein called prostate specific membrane antigen (PSMA) is implicated in PCa invasiveness as well as neovasculature of nonprostatic solid tumors. Little is known about the exact role of PSMA in PCa progression albeit it is one of the most validated biomarker for the diagnosis and detection of PCa. The functional form of PSMA occurs as a non-covalent homodimer, which is a common motif among bitopic membrane proteins. Further, it is widely recognized that many membrane proteins participate in intracellular signal transduction through the lateral association of their transmembrane domains (TMDs), but this has not been directly demonstrated in PSMA. We hypothesize that PSMA TMD participates in oligomerization and subsequent activation of PSMA. Thus, to gain insight into the non-covalent assembly of PSMA, we aim to determine if the isolated PSMA TMD peptide forms a dimer and if the treatment of full-length PSMA with exogenous PSMA TMD peptide will perturb its properties. Studies using isolated TMDs have been widely used to probe protein-protein interactions and provide useful information on protein assembly under membrane mimetic systems. We identified the amino acid sequence and designed the isolated PSMA TMD peptide based on a known human PSMA sequence (UniProt accession code: [Q04609][1]). Two Lysine residues ware appended at both the N- and C-termini to increase the solubility of this highly hydrophobic peptide in polar solvents. Our initial studies revealed that the TMD has twenty-four residues and contains two repeat units of the Small-XXX-Small motif. This sequence is known to be crucial in inducing a strong self-assembly as observed in the bitopic protein Glycophorin A. We prepared the PSMA TMD peptide by microwave-assisted solid phase synthesis and purified by high performance liquid chromatography. The peptide was analyzed by MALDI-TOF mass spectrometry to establish homogeneity and by circular dichroism to examine its secondary structure in membrane mimetic environment. We investigated the oligomerization of the peptide using SDS-PAGE gel shift and fluorescence self-quenching assays. We tested the effect of the peptide against PSMA using an established assay. Herein we report on the oligomerization and PSMA binding properties of PSMA TMD. Our findings may shed light on the role of PSMA TMD in PSMA oligomerization and its potential as a model peptide that target transmembrane helices. It may also aid in the rational design and development of peptide probes for protein-protein interactions as well as therapeutics that target cancer- and clinically-relevant membrane proteins. Citation Format: Brianna S. Berg, Brandan M. Cook, Nathan Beattie, Cliff E. Berkman, Jonel P. Saludes. The transmembrane domain of prostate-specific membrane antigen: Its role in oligomerization and activation in prostate cancer and nonprostatic neovasculature. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3238. doi:10.1158/1538-7445.AM2014-3238 [1]: /lookup/external-ref?link_type=GEN&access_num=Q04609&atom=%2Fcanres%2F74%2F19_Supplement%2F3238.atom