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Biotechnology Letters

, Volume 41, Issue 1, pp 171–180 | Cite as

Enzyme-catalyzed regio-selective demethylation of papaverine by CYP105D1

  • Chen Shen
  • Wanli Zhao
  • Xuming Liu
  • Jihua Liu
Original Research Paper

Abstract

Objectives

To investigate the regio-selective demethylation of papaverine by CYP105D1 and develop a whole-cell biocatalytic system for the preparative synthesis of 6-O-demethyl-papaverine.

Results

CYP105D1 from Streptomyces griseus ATCC 13273 was used for the regioselective demethylation of papaverine at C-6 using putidaredoxin reductase (PDR) and putidaredoxin (Pdx) as the electron transport system. The Km value of CYP105D1 towards papaverine was estimated to be 92.24 μM. Furthermore, a CYP105D1-based whole-cell system was established in E. coli BL21(DE3). The whole cell biotransformation condition was optimized as 25 °C, pH 7.5, 8 g (cell dry weight) L−1 whole cell biomass and 3% (v/v) PEG-200 as cosolvent. Under the optimal condition, the conversion yield of papaverine reached to 61.15% within 24 h.

Conclusions

The selective demethylation of papaverine by CYP105D1 was accomplished. The CYP105D1-based whole-cell biocatalyst has a potential used for the efficient synthesis of 6-O-demethyl-papaverine.

Keywords

Biocatalysis Cytochrome P450 O-demethylation Papaverine Whole-cell biotransformation 

Notes

Acknowledgements

This work was supported by the Major Scientific and Technological Specialized Project for ‘New Drugs Development’ (No. 2012ZX09J12110-06B), and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).

Supporting information

Supplementary Table 1—Primers used in this study.

Supplementary Table 2—Strains and plasmids used in this study.

Supplementary Table 3—HPLC conditions for analysis of papaverine biocatalytic profile.

Supplementary Fig. 1—Expression and purification of CYP105D1, Pdx and PDR.

Supplementary Fig. 2—The codon-optimized sequence of the gene encoding putidaredoxin reductase (PDR).

Supplementary Fig. 3—The codon-optimized sequence of the gene encoding putidaredoxin (Pdx).

Supplementary Fig. 4—1H NMR analysis spectrum of the product 6-O-demethylpapaverine (MeOD, 500 MHz).

Supplementary Fig. 5—13C NMR analysis spectrum of the product 6-O-demethylpapaverine (MeOD, 126 MHz).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

10529_2018_2626_MOESM1_ESM.doc (1 mb)
Electronic supplementary material 1 (DOC 1065 kb)

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Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  1. 1.Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese PharmacyChina Pharmaceutical UniversityNanjingChina
  2. 2.School of Life Science and TechnologyChina Pharmaceutical UniversityNanjingChina
  3. 3.State Key Laboratory of Natural MedicinesChina Pharmaceutical UniversityNanjingChina

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