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

Abstract

NCL results in the quantitative yield of a membrane protein, where a thioester peptide is formed from an oxo-ester with an in situ cleavable solubilizing tag.

Challenges and Perspectives in Chemical Synthesis of Highly Hydrophobic Peptides

“…However, this method is hampered by impossible incorporation of removable C-terminal solubilizing tag to the hydrazide moiety. Lately, Baumruck et al (2018) presents an oxo-ester mediated NCL with almost quantitative NCL yields introducing an interesting prospect by attaching removable solubilizing tag to the oxoester. This strategy benefits from the easier handling of highly hydrophobic thioester-forming peptide due to its solubility while purification and analysis prior to NCL.…”

“…A brilliant example of using an arginine tag [phenylacetamidomethyl (Phacm) attached to cysteine] and selective orthogonal removal of the Cys masking group 1,3-thiazolidine-4-carboxylic acid (Thz) without affecting this tag was demonstrated by Jbara et al (2018). Moreover, most of the presented synthetic strategies include addition of organic solvents such as TFE Valiyaveetil et al, 2002;Baumruck et al, 2018) or surfactants like OG and DPC (Bianchi et al, 1999;Clayton, 2005;Lahiri et al, 2011;Zheng et al, 2014) to the ligation solution. Finally, Tang et al presented a synthesis and ligation route toward BM2 that was published as a protocol, establishing a basis for generation of general approach.…”

“…The last decades have witnessed an unprecedented progress in chemical peptide synthesis, however there were still sequences which were difficult or even impossible to synthesize by standard SPPS belonging to class of "difficult sequences, " i.e., amylin (Cooper et al, 1987;Harris et al, 2013), Aβ(1-42) (Bacsa et al, 2010;Kasim et al, 2019), and BM2(1-51) (Baumruck et al, 2018). With respect to SPPS "difficult sequences" are defined as peptides that are poorly solvated while attached to the solid support thus preventing complete deprotection and coupling steps (Tickler and Wade, 2007).…”

Development of Trityl Group Anchored Solubilizing Tags for Peptide and Protein Synthesis

“…Consequently, we improved the synthesis of 4 . The preparation of [Cys(Trt‐K 5 )11, Leu21]BM2(1–51) ( 8 ) was achieved without significant side reactions in the absence of any organic solvents, whereas the preparation of 4 through NCL–desulfurization without the aid of the Trt‐K tag required the addition of organic solvents, such as HFIP, to solubilize hydrophobic peptide segments …”

Smoothing membrane protein structure determination by initial upstream stage improvements

“…To overcome the limitations displayed by these in vivo expression systems-toxicity, limited membrane space for MP functional folding and inefficient transport, and membrane insertion mechanisms-CFE systems have been reported, which rely on the use of prokaryotic and eukaryotic protein synthesis machinery and related elements to direct protein synthesis from added DNA or mRNA templates (He et al 2011;Henrich et al 2015;Zheng et al 2014). In a different way, the preparation of highly hydrophobic peptides representing functional parts of MPs foreseeing their application onto structural and functional studies can be attained via chemical synthesis (Baumruck et al 2018). Previously, Fernández and Vega (2016) reported some recommendations on which expression host use for a particular protein.…”