One-Step Synthesis of D-Amino Acids

Novick, Scott

Abstract

Many currently produced drugs such as beta-Iactam antibiotics, thrombin inhibitors and fertility treatments, and fine chemicals such as insecticides contain D-amino acids. Some of these compounds are produced on a scale greater than 5000 tons/yr. In addition, there are a number of D-amino acid containing drugs, either in the research or clinical test stages, used for the treatment of such illnesses as endometriosis and uterine fibroids in women, benign prostatic hypertrophy in men, and HIV. D-Amino acids have the advantage of often being more bioactive than their L-counterpart and they are also frequently more stable as they are less likely to undergo enzymatic degradation in the liver, kidney, and bloodstream. Clearly, there is significant demand for enantiomerically pure D-amino acids and this is expected to rise significantly in the near future as they become critical components of new pharmaceuticals. While current technologies exist for producing D-amino acids they all have drawbacks including poor yields and the requirement of multiple reaction steps. New methods to improve upon current technologies are needed. One method, as discussed in this proposal, is to produce D-amino acids from the 2-keto acid precursor using a D-amino acid dehydrogenase. This will allow for the single-step synthesis of D-amino acid from inexpensive starting materials. To this end, it is proposed to use direct evolution to generate broadrange D-amino acid dehydrogenases. The starting enzyme, meso-2,6-diaminopimelate-D-dehydrogenase (DAPDH) is highly selective for the synthesis of D-amino acids.
The specific aims of this proposal are to generate a library of mutant DAPDH, screen this library for activity towards important D-amino acids, characterize the best mutants, and use these mutants to perform a gram scale synthesis of one or more key D-amino acids.