Thermococcus Sp. KS-1 PPIase As a Fusion Partner Improving Soluble Production of Aromatic Amino Acid Decarboxylase

Overview

Journal AMB Express

Date 2021 Dec 27

PMID 34958446

Authors

Takashi Koyanagi

Ayumi Hara

Kanako Kobayashi

Yuji Habara

Akira Nakagawa

Hiromichi Minami

Takane Katayama

Norihiko Misawa

Affiliations

Soon will be listed here.

Abstract

Peptidyl-prolyl cis-trans isomerase (PPIase, EC 5.2.1.8) catalyzes the racemization reaction of proline residues on a polypeptide chain. This enzyme is also known to function as a molecular chaperon to stabilize protein conformation during the folding process. In this study, we noted FK506 binding protein (FKBP)-type PPIase from a hyperthemophilic archaeon Thermococcus sp. strain KS-1 (PPIase ) to improve the solubility of Pseudomonas putida aromatic amino acid decarboxylase (AADC) that is an indispensable enzyme for fermentative production of plant isoquinoline alkaloids. AADC fused N-terminally with the PPIase (PPIase -AADC), which was synthesized utilizing Escherichia coli host, showed improved solubility and, consequently, the cell-free extract from the recombinant strain exhibited 2.6- to 3.4-fold elevated AADC activity than that from the control strain that expressed the AADC gene without PPIase . On the other hand, its thermostability was slightly decreased by fusing PPIase . The recombinant E. coli cells expressing the PPIase -AADC gene produced dopamine and phenylethylamine from L-dopa and phenylalanine by two- and threefold faster, respectively, as compared with the control strain. We further demonstrated that the efficacy of PPIase -AADC in solubility and activity enhancement was a little but obviously higher than that of AADC fused N-terminally with NusA protein, which has been assumed to be the most effective protein solubilizer. These results suggest that PPIase can be used as one of the best choices for producing heterologous proteins as active forms in E. coli.

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