Asymmetric synthesis of L-phosphinothricin using thermostable alpha-transaminase mined from Citrobacter koseri

alpha-Transaminase (alpha-TA) responsible for catalyzing the reversible transfer of amino groups between amine donors and amine acceptors, is applicable to enzymatic route for asymmetric synthesis of herbicide L-phosphinothricin (L-PPT). In the search for alpha-TAs with better catalysis performance, three alpha-TAs were discovered by genome mining approach using a known sequence encoding Escherichia coli tyrosine TA (TyrB) as probe. Through detailed comparison of their expression amount, activities and characteristics, Citrobacter koseri TA (CkTA) exhibited better activity and thermostability, which retain 65.9% of initial activity after incubation at 57 degrees C for 4 h. The Km and kcat/Km values of CkTA were 36.75mM and 34.29 mM(-1) min(-1), respectively. In addition, recombinant CkTA cells were immobilized onto Celite 545 using tris(hydroxymethyl) phosphine as crosslinker. During five repetitive asymmetric synthesis of L-PPT from 20 g/L prostereogenic ketone using L-Glu as amine donor, all the yields of L-PPT reached up to 91.2% (> 99% ee). These characteristics made CkTA a valuable addition to the currently scarce a-TA library for stereospecific synthesis of L-PPT.