The industrial alkaline protease, alcalase, is stable and active in a high concentration of organic solvents and useful as a biocatalyst for (i) diastereoselective hydrolysis of peptide esters and preparation of racemization-free peptides; (ii) selective incorporation of esters of D-amino acid into peptides in t-butanol via a selective hydrolysis of esters of D,L-amino acid, followed by using the unhydrolyzed Dvesters as a nucleophile in a kinetically controlled peptide bond formation; (iii) resolution of esters of amino acid in 95% t-butanol/f % water, followed by saponification of the unreacted esters 10 offer both enantiomers with high yield and optical purity; (iv) completely resolve amino-acid esters with high yield and optical purity via in situ racemization of the unreacted antipode catalyzed by pyridoxal f-phosphate; (v) cryobioorganic synthesis of pep tides with increased yields 15%-40% of peptide bond formation by reaction at 5 "C instead of25-30 "C of a kinetically controlled enzymatic reaction in alcohols.
301Substrates (Ala-Ala-P2-PhEl-OMe) S,,·22 I