AsiaChem | Chemistry in Japan | December 2021 Volume 2 Issue 1

www.asiachem.news December 2021 | 43 By Kousuke Tsuchiya and Keiji Numata https://doi.org/10.51167/acm00022 amino acid combinations offer complex periodic sequences through chemoenzymatic polymerization. Our targets are functional polypeptides for a wide range of applications, from biomimetic structural materials to bioactive peptides for biotechnology. Proteases cleave amide bonds of specific proteins. The catalytic center in proteases attacks an amide bond to form an activated tetrahedral intermediate followed by hydrolytic reaction, resulting in scission of proteins. If we can exploit the reverse reaction of enzymatic hydrolysis, polymerization of amino acids proceeds in a chemoselective manner regulated by spatial information for the catalytic pocket in enzymes.3 Although the equilibrium for enzymatic hydrolysis is biased toward cleavage, moderate activation of amino acids by modification with ester groups on the C-terminus kinetically facilitates protease-mediated formation of the tetrahedral intermediate and subsequent aminolysis reaction. The catalytic pocket in proteins consists of a series of subsites, each of which has distinct specificity to amino acid substrates. We harnessed chemoenzymatic polymerization by selecting a reasonable combination of substrate-specific proteins and amino acidmonomers, which was assisted by theoretical predictions using several techniques, including molecular dynamics simulations.4 Even an amino acid that mismatches the substrate specificity of proteases can be incorporated into polypeptides when it is inserted into an oligopeptide monomer with an elaborate sequence design to mitigate the mismatch. This technique motivates us to design and synthesize novel artificial polypeptides with more complicated sequences for further functionalization. Structural proteins Proteins and polypeptides play critical roles in living bodies. Versatile functions of proteins are determined by sequential variety, which are assembled into functional higher-order structures, and proteins are roughly categorized into structural proteins and globular functional proteins such as enzymes. In particular, structural proteins often possess long repetitive sequences to fold into specific higher-order structures that build up supportive frameworks in body tissues.5 To mimic such repetitive sequences as found in silk fibroin, collagen, elastin, and resilin, our synthetic technique for designing and controlling periodic sequences via chemoenzymatic polymerization is useful to offer artificial biomimetic polypeptide materials with physical and physiological functionality. We designed polypeptides containing specific short peptide motifs that are thought to govern the physical properties of structural proteins Figure 1. Chemoenzymatic polymerization catalyzed by proteases (papain). The amino acid ester monomer is activated by acylation with a catalytic cysteine residue in papain to produce a tetrahedral intermediate, which subsequently undergoes aminolysis with another amino acid ester monomer.

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