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Preparation of an Amino Acid Intermediate for the Dipeptidyl Peptidase IV Inhibitor, Saxagliptin, using a Modified Phenylalanine Dehydrogenase
- Source :
- Advanced Synthesis & Catalysis. 349:1369-1378
- Publication Year :
- 2007
- Publisher :
- Wiley, 2007.
-
Abstract
- The non-proteinogenic amino acid 2-(3-hydroxy-1-adamantyl)-(2S)-aminoethanoic acid [2, (S)-3-hydroxyadamantylglycine], is a key intermediate required for the synthesis of Saxagliptin, a dipeptidyl peptidase IV inhibitor under development for treatment of type 2 diabetes mellitus. Keto acid 2-(3-hydroxy-1-adamantyl)-2-oxoethanoic acid (1) was converted to (S)-3-hydroxyadamantylglycine by reductive amination using a phenylalanine dehydrogenase from Thermoactinomyces intermedius expressed in a modified form in Pichia pastoris or Escherichia coli. NAD (nicotinamide adenine dinucleotide) produced during the reaction was recycled to NADH (reduced form of nicotinamide adenine dinucleotide) using formate dehydrogenase. Pichia pastoris produces an endogenous formate dehydrogenase when grown on methanol, and the corresponding gene was cloned and expressed in E. coli. The modified phenylalanine dehydrogenase contains two amino acid changes at the C-terminus and a 12-amino acid extension of the C-terminus. The modified enzyme is more effective with keto acid 1 than the wild-type enzyme, but less effective with the natural substrate, phenylpyruvate. Production of multi-kg batches was originally carried out with extracts of Pichia pastoris expressing the modified phenylalanine dehydrogenase from Thermoactinomyces intermedius and endogenous formate dehydrogenase, and further scaled up using a preparation of the two enzymes expressed in E. coli.
- Subjects :
- chemistry.chemical_classification
biology
General Chemistry
Nicotinamide adenine dinucleotide
Formate dehydrogenase
biology.organism_classification
Dipeptidyl peptidase
Pichia pastoris
Amino acid
Phenylalanine dehydrogenase
chemistry.chemical_compound
chemistry
Biochemistry
NAD+ kinase
Branched-chain alpha-keto acid dehydrogenase complex
Subjects
Details
- ISSN :
- 16154169 and 16154150
- Volume :
- 349
- Database :
- OpenAIRE
- Journal :
- Advanced Synthesis & Catalysis
- Accession number :
- edsair.doi...........a1277651cf8770f36ba50d61336a12df