Chemical synthesis of transmembrane peptide and its application for research on the transmembrane–juxtamembrane region of membrane protein

Abstract

Membrane proteins possess one or more hydrophobic regions that span the membrane and interact with the lipids that constitute the membrane. The interactions between the transmembrane (TM) region and lipids affect the structure and function of these membrane proteins. Molecular characterization of synthetic TM peptides in lipid bilayers helps to understand how the TM region participates in the formation of the structure and in the function of membrane proteins. The use of synthetic peptides enables site-specific labeling and modification and allows for designing of an artificial TM sequence. Research involving such samples has resulted in significant increase in the knowledge of the mechanisms that govern membrane biology. In this review, the chemical synthesis of TM peptides has been discussed. The preparation of synthetic TM peptides is still not trivial; however, the accumulated knowledge summarized here should provide a basis for preparing samples for spectroscopic analyses. The application of synthetic TM peptides for gaining insights into the mechanism of signal transduction by receptor tyrosine kinase (RTK) has also been discussed. RTK is a single TM protein and is one of the difficult targets in structural biology as crystallization of the full-length receptor has not been successful. This review describes the structural characterization of the synthetic TM-juxtamembrane sequence and proposes a possible scheme for the structural changes in this region for the activation of ErbBs, the epidermal growth factor receptor family. © 2015 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 613-621, 2016.

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

“…A great advantage of chemical synthesis displays the possibility to custom design of the desired sequence. As soon as the successful synthetic protocol is elaborated, the integration of unnatural amino acids, mutations at arbitrary positions, post-translational modifications (PTMs) and sitespecific labels for e.g., solid-state/solution NMR spectroscopy or fluorescence microscopy experiments is readily possible (Sato, 2016). Furthermore, product in the multi-milligram range can be obtained making numerous analytical experiments possible, leading to a better structural and functional understanding.…”

Chemical synthesis of membrane proteins: a model study on the influenza virus B proton channel

“…The synthetic production of highly hydrophobic peptides, such as medium-sized fragments of membrane proteins representing functional parts, offers many advantages over other production strategies, such as protein expression, mainly because synthetic peptides can be customized and derivatized depending on the purpose of the research. 7 Moreover, the production of multi-milligram amounts of membrane protein fragments by chemical synthesis might be even more important with respect to the development of bio-inspired materials. 8 …”

“… 11 – 14 The work up, including purification steps, for those peptides is usually challenging, since these peptides contain a high number of amino acids with hydrophobic side chains, which causes their aggregation in conventional solvents. 7 , 15 The first attempt of the synthesis of influenza A virus M2 membrane protein was performed by Kochendoerfer et al The authors used a Boc-based SPPS, and for the ligation step they used 6 M guanidinium chloride containing 20% TFE in order to solubilize the peptide, resulting in 65% product formation. 12 …”

Coupling of sterically demanding peptides by β-thiolactone-mediated native chemical ligation

“…This challenge is particularly relevant for the synthesis of membrane proteins in which amino acids such as Val, Ile, Leu, and Ala constitute more than one-third of the transmembrane domains. 5 Thus, the need to develop methodologies for the ligation of these residues is tremendous. Although considerable efforts have been directed towards improving the ligation efficiency at sterically hindered sites, 6 , 7 such as the elegant report by Dong 8 on the internal activation of thioesters promoting the occurrence of ligation at proline sites, the problem remains and continues to place severe limitations on the ability to access membrane proteins by NCL.…”