Efficient Preparation of (S)-N-Boc-3-Hydroxylpiperidine Through Bioreduction by a Thermostable Aldo-KetoReductase

Appl Biochem Biotechnol. 2017 Apr;181(4):1304-1313. doi: 10.1007/s12010-016-2285-3. Epub 2016 Nov 16.

Abstract

(S)-N-Boc-3-hydroxypiperidine ((S)-NBHP) is a key pharmaceutical intermediate and the chiral source in synthesizing Imbruvica, which is a newly approved drug in lymphoma therapy by targeting Bruton's tyrosine kinase (BTK). Current chemical synthesis of (S)-NBHP suffered from the need of noble metal catalyst and low yield. The single reported bioconversion of (S)-NBHP was achieved by using recombinant ketoreductase, but enzyme sequence was kept confidential and the catalytic process suffered from the thermodeactivation and substrate inhibition. In the current study, we presented a thermostable aldo-keto reductase (AKR)-AKR-43-which showed high activity toward N-Boc-3-piperidone (NBP) to produce (S)-NBHP, high enantioselectivity, and no substrate inhibition. The molecular simulations demonstrated the structural rationale for the enantioselectivity of AKR-43 toward NBP and supported the classic ordered two-step catalytic mechanism. The catalytic process was achieved by using glucose dehydrogenase (GDH) for cofactor recycling, and the optimal reaction conditions were determined to be 30 °C and pH 7.5. Within a reaction time of 16 h, the 16 % substrate concentration (w/w), over 99 % ee and under 3.5 % of enzyme loading (w/w) characterized a high efficiency process with promising industrial values.

Keywords: (S)-N-Boc-3-hydroxypiperidine; Aldo-keto reductase; Bioconversion; Molecular simulation; N-Boc-3-piperidone.

MeSH terms

  • Aldehyde Reductase / chemistry
  • Aldehyde Reductase / metabolism*
  • Aldo-Keto Reductases
  • Bacillus subtilis / enzymology
  • Biocatalysis
  • Enzyme Stability
  • Molecular Dynamics Simulation
  • Oxidation-Reduction
  • Piperidines / chemistry
  • Piperidines / metabolism*
  • Protein Conformation
  • Stereoisomerism
  • Substrate Specificity
  • Temperature*

Substances

  • N-tert-butoxycarbonyl-3-hydroxypiperidine
  • Piperidines
  • Aldo-Keto Reductases
  • Aldehyde Reductase