Journal
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
Volume 97, Issue 2, Pages 649-659Publisher
SPRINGER
DOI: 10.1007/s00253-012-4324-5
Keywords
Ginsenoside; Biotransformation; Glycoside hydrolase; Actinosynnema mirum
Categories
Funding
- Next-Generation BioGreen 21 Program, Rural Development Administration [PJ008193]
- Intelligent Synthetic Biology Center of Global Frontier Project
- Ministry of Education, Science and Technology, Republic of Korea [2011-0031967]
- National Research Foundation of Korea [2011-0031967, 2011-0031955] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
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This study focused on the cloning, expression, and characterization of ginsenoside-transforming recombinant beta-glucosidase from Actinosynnema mirum KACC 20028(T) in order to biotransform ginsenosides efficiently. The gene, termed as bglAm, encoding a beta-glucosidase (BglAm) belonging to the glycoside hydrolase family 3 was cloned. bglAm consisted of 1,830 bp (609 amino acid residues) with a predicted molecular mass of 65,277 Da. This enzyme was overexpressed in Escherichia coli BL21(DE3) using a GST-fused pGEX 4T-1 vector system. The recombinant BglAm was purified with a GST center dot bind agarose resin and characterized. The optimum conditions of the recombinant BglAm were pH 7.0 and 37 A degrees C. BglAm could hydrolyze the outer and inner glucose moieties at the C3 and C20 of the protopanaxadiol-type ginsenosides (i.e., Rb-1 and Rd, gypenoside XVII) to produce protopanaxadiol via gypenoside LXXV, F-2, and Rh-2(S) with various pathways. BglAm can effectively transform the ginsenoside Rb-1 to gypenoside XVII and Rd to F-2; the K (m) values of Rb-1 and Rd were 0.69 A +/- 0.06 and 0.45 A +/- 0.02 mM, respectively, and the V (max) values were 16.13 A +/- 0.29 and 51.56 A +/- 1.35 mu mol min(-1) mg(-1) of protein, respectively. Furthermore, BglAm could convert the protopanaxatriol-type ginsenoside Re and Rg(1) into Rg(2)(S) and Rh-1(S) hydrolyzing the attached glucose moiety at the C6 and C20 positions, respectively. These various ginsenoside-hydrolyzing pathways of BglAm may assist in producing the minor ginsenosides from abundant major ginsenosides.
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