Recombinant expression, purification, and characterization of transmembrane domain of syndecan-4

Abstract

The transmembrane heparan sulfate proteoglycan, syndecan-4, plays potentially important roles that may influence tissue development and repair as well as the pathogenesis of numerous diseases. Recently, a number of studies have been conducted on the structural properties and biological functions of the syndecan-4 cytoplasmic domain or extracellular domain. However, previous attempts to understand the structure and function of the syndecan-4 transmembrane domain (Syd4-TM) have been marred by experimental adversities due to insufficient yields and low solubility. To investigate the structural and functional properties of Syd4-TM, an efficient method of preparing milligram quantities of the corresponding peptide is necessary. Herein, we report an efficient method for the recombinant expression and purification of Syd4-TM peptide. Peptide was released from the fusion protein, and then purified by semi-preparative reversed-phase HPLC. Finally, more than 5mg of Syd4-TM peptide was obtained with high purity from 1l of M9 minimal media under optimized conditions. The preliminary biophysical properties of recombinant Syd4-TM peptide were studied by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. The analysis of the CD spectrum shows that Syd4-TM adopts a stable α-helical structure in micelle environments. And solution NMR studies showed that Syd4-TM forms an asymmetric dimer in micelles.